Vinit Mahajan, MD, PhD

All Publications

Molecular Surgery: Proteomics of a Rare Genetic Disease Gives Insight into Common Causes of Blindness. iScience Velez, G., Mahajan, V. B. 2020; 23 (11): 101667
Abstract

Rare diseases are an emerging global health priority. Although individually rare, the prevalence of rare "orphan" diseases is high, affecting approximately 300 million people worldwide. Treatments for these conditions are often inadequate, leaving the disease to progress unabated. Here, we review the clinical features and pathophysiology of neovascular inflammatory vitreoretinopathy (NIV), a rare inflammatory retinal disease caused by mutations in the CAPN5 gene. Although the prevalence of NIV is low (1 in 1,000,000 people), the disease mimics more common causes of blindness (e.g. uveitis, retinitis pigmentosa, proliferative diabetic retinopathy, and proliferative vitreoretinopathy) at distinct clinical stages. There is no cure for NIV to date. We highlight how personalized proteomics helped identify potential stage-specific biomarkers and drug targets in liquid vitreous biopsies. The NIV vitreous proteome revealed enrichment of molecular pathways associated with common retinal pathologies and implicated superior targets for therapeutic drug repositioning. In addition, we review our pipeline for collecting, storing, and analyzing ophthalmic surgical samples. This approach can be adapted to treat a variety of rare genetic diseases.

View details for DOI 10.1016/j.isci.2020.101667

View details for PubMedID 33134897
Retinal Manifestations of Mitochondrial Oxidative Phosphorylation Disorders. Investigative ophthalmology & visual science Oh, J. K., Lima de Carvalho, J. R., Nuzbrokh, Y., Ryu, J., Chemudupati, T., Mahajan, V. B., Sparrow, J. R., Tsang, S. H. 2020; 61 (12): 12
Abstract

Purpose: The purpose of this paper was to discuss manifestations of primary mitochondrial dysfunctions and whether the retinal pigment epithelium or the photoreceptors are preferentially affected.Methods: A retrospective analysis was performed of patients with clinically and laboratory confirmed diagnoses of maternally inherited diabetes and deafness (MIDD) or Kearns-Sayre syndrome (KSS). Patients underwent full ophthalmic examination, full-field electroretinogram, and multimodal imaging studies, including short-wavelength autofluorescence, spectral domain-optical coherence tomography, and color fundus photography.Results: A total of five patients with MIDD and four patients with KSS were evaluated at two tertiary referral centers. Mean age at initial evaluation was 50.3 years old. Nascent outer retinal tubulations corresponding with faint foci of autofluorescence were observed in two patients with MIDD. Characteristic features of this cohort included a foveal sparing phenotype observed in 13 of 18 eyes (72%), global absence of intraretinal pigment migration, and preserved retinal function on full-field electroretinogram testing in 12 of 16 eyes (75%). One patient diagnosed with MIDD presented with an unusual pattern of atrophy surrounding the parapapillary region and one patient with KSS presented with an atypical choroideremia-like phenotype.Conclusions: MIDD and KSS are phenotypically heterogeneous disorders. Several features of disease suggest that primary mitochondrial dysfunction may first affect the retinal pigment epithelium followed by secondary photoreceptor loss. Similarities between primary mitochondrial degenerations and retinal disorders, such as age-related macular degeneration may suggest a primary role of mitochondria in the pathogenesis of these oligogenic disorders.

View details for DOI 10.1167/iovs.61.12.12

View details for PubMedID 33049060
Untitled Reply RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Mahajan, V. B., Wood, E. 2020; 40 (8): E37
View details for Web of Science ID 000559752400006
Whole-Exome Sequencing of Patients with Posterior Segment Uveitis. American journal of ophthalmology Li, A. S., Velez, G., Darbro, B., Toral, M. A., Yang, J., Tsang, S. H., Ferguson, P. J., Folk, J. C., Bassuk, A. G., Mahajan, V. B. 2020
Abstract

OBJECTIVE: To elucidate molecular risk factors for posterior segment uveitis using a functional genomics approach.DESIGN: Genetic Association Cohort Study.METHODS: SETTING: Single-center study at an academic referral center.STUDY POPULATION: 164 patients with clinically diagnosed uveitis of the posterior segment.MAIN OUTCOME MEASURES: Exome sequencing was used to detect variants identified in 164 patients with posterior segment uveitis. A phenotype-driven analysis, protein structural modeling and in silico calculations were then used to rank and predict the functional consequences of key variants.RESULTS: A total of 203 single nucleotide variants, in 23 genes across 164 patients, were included in this study. Both known and novel variants were identified in genes previously implicated in specific types of syndromic uveitis - such as NOD2 (Blau Syndrome) and CAPN5 NIV (Neovascular Inflammatory Vitreoretinopathy) - as well as variants in genes not previously linked to posterior segment uveitis. Based on a ranked list and protein-protein-interaction network, missense variants in NOD-like receptor family genes (NOD2, NLRC4, NLRP3, and NLRP1), CAPN5, and TYK2 were characterized via structural modeling and in silico calculations to predict how specific variants might alter protein structure and function. The majority of analyzed variants were notably different from wild type.CONCLUSIONS: This study implicates new pathways and immune signaling proteins that may be associated with posterior segment uveitis susceptibility. A larger cohort and functional studies will help validate the pathogenicity of the mutations identified. In specific cases, whole exome sequencing can help diagnose non-syndromic uveitis patients harboring known variants for syndromic inflammatory diseases.

View details for DOI 10.1016/j.ajo.2020.07.021

View details for PubMedID 32707200
Novel PRPF31 gene loss of function for retinitis pigmentosa 11 Chemudupati, T., Sun, Y., Yang, J., Ng, B., Mahajan, V. B. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2020
View details for Web of Science ID 000554528300228
Structural Insights into the Unique Activation Mechanisms of a Non-classical Calpain and Its Disease-Causing Variants. Cell reports Velez, G., Sun, Y. J., Khan, S., Yang, J., Herrmann, J., Chemudupati, T., MacLaren, R. E., Gakhar, L., Wakatsuki, S., Bassuk, A. G., Mahajan, V. B. 2020; 30 (3): 881
Abstract

Increased calpain activity is linked to neuroinflammation including a heritable retinal disease caused by hyper-activating mutations in the calcium-activated calpain-5 (CAPN5) protease. Although structures for classical calpains are known, the structure of CAPN5, a non-classical calpain, remains undetermined. Here we report the 2.8A crystal structure of the human CAPN5 protease core (CAPN5-PC). Compared to classical calpains, CAPN5-PC requires high calcium concentrations for maximal activity. Structure-based phylogenetic analysis and multiple sequence alignment reveal that CAPN5-PC contains three elongated flexible loops compared to its classical counterparts. The presence of a disease-causing mutation (c.799G>A, p.Gly267Ser) on the unique PC2L2 loop reveals a function in this region for regulating enzymatic activity. This mechanism could be transferred to distant calpains, using synthetic calpain hybrids, suggesting an evolutionary mechanism for fine-tuning calpain function by modifying flexible loops. Further, the open (inactive) conformation of CAPN5-PC provides structural insight into CAPN5-specific residues that can guide inhibitor design.

View details for DOI 10.1016/j.celrep.2019.12.077

View details for PubMedID 31968260
Metabolite therapy guided by liquid biopsy proteomics delays retinal neurodegeneration. EBioMedicine Wert, K. J., Velez, G., Kanchustambham, V. L., Shankar, V., Evans, L. P., Sengillo, J. D., Zare, R. N., Bassuk, A. G., Tsang, S. H., Mahajan, V. B. 2020; 52: 102636
Abstract

Neurodegenerative diseases are incurable disorders caused by progressive neuronal cell death. Retinitis pigmentosa (RP) is a blinding neurodegenerative disease that results in photoreceptor death and progresses to the loss of the entire retinal network. We previously found that proteomic analysis of the adjacent vitreous served as way to indirectly biopsy the retina and identify changes in the retinal proteome.We analyzed protein expression in liquid vitreous biopsies from autosomal recessive (ar)RP patients with PDE6A mutations and arRP mice with Pde6ɑ mutations. Proteomic analysis of retina and vitreous samples identified molecular pathways affected at the onset of photoreceptor death. Based on affected molecular pathways, arRP mice were treated with a ketogenic diet or metabolites involved in fatty-acid synthesis, oxidative phosphorylation, and the tricarboxylic acid (TCA) cycle.Dietary supplementation of a single metabolite, ɑ-ketoglutarate, increased docosahexaeonic acid levels, provided neuroprotection, and enhanced visual function in arRP mice. A ketogenic diet delayed photoreceptor cell loss, while vitamin B supplementation had a limited effect. Finally, desorption electrospray ionization mass spectrometry imaging (DESI-MSI) on ɑ-ketoglutarate-treated mice revealed restoration of metabolites that correlated with our proteomic findings: uridine, dihydrouridine, and thymidine (pyrimidine and purine metabolism), glutamine and glutamate (glutamine/glutamate conversion), and succinic and aconitic acid (TCA cycle).This study demonstrates that replenishing TCA cycle metabolites via oral supplementation prolongs retinal function and provides a neuroprotective effect on the photoreceptor cells and inner retinal network.NIH grants [R01EY026682, R01EY024665, R01EY025225, R01EY024698, R21AG050437, P30EY026877, 5P30EY019007, R01EY018213, F30EYE027986, T32GM007337, 5P30CA013696], NSF grant CHE-1734082.

View details for DOI 10.1016/j.ebiom.2020.102636

View details for PubMedID 32028070
Reply. Retina (Philadelphia, Pa.) Mahajan, V. B., Wood, E. 2020
View details for DOI 10.1097/IAE.0000000000002828

View details for PubMedID 32343099
Phenotypic variance in Calpain-5 retinal degeneration. American journal of ophthalmology case reports Tang, P. H., Chemudupati, T., Wert, K. J., Folk, J. C., Mahajan, M., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2020; 18: 100627
Abstract

To characterize the phenotype of patients with mild calpain-5 Neovascular Inflammatory Vitreoretinopathy (ADNIV).The CAPN5 p.R243L mutation is typically associated with onset in the twenties and severe, progressive uveitis, retinal neovascularization, and intraocular fibrosis. Two subjects with this CAPN5 variant only showed mild peripheral retinal pigmentary degeneration and loss of the ERG b-wave at age 45 and 69, respectively, without signs of uveitis or neovascularization.The phenotypic penetrance of a specific variant in CAPN5-vitreoretinopathy may vary significantly in severity. Patients with pigmentary retinal dystrophy may consider CAPN5 gene testing.

View details for DOI 10.1016/j.ajoc.2020.100627

View details for PubMedID 32274441

View details for PubMedCentralID PMC7132063
Proteomic analysis of intermediate uveitis suggests myeloid cell recruitment and implicates IL-23 as a therapeutic target. American journal of ophthalmology case reports Sepah, Y. J., Velez, G., Tang, P. H., Yang, J., Chemudupati, T., Li, A. S., Nguyen, Q. D., Bassuk, A. G., Mahajan, V. B. 2020; 18: 100646
Abstract

To profile vitreous protein expression of intermediate uveitis (IU) patients.We identified a mean of 363 ± 41 unique proteins (mean ± SD) in IU vitreous and 393 ± 69 unique proteins in control samples using liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis of liquid vitreous biopsies collected during pars plana vitrectomy. A total of 233 proteins were differentially expressed among control and IU samples, suggesting a protein signature that could distinguish the two groups. Pathway analysis identified 22 inflammatory mediators of the interleukin-12 (IL-12) signaling pathway in IU vitreous. Upstream regulator analysis identified downstream mediators of IL-23 and myeloid differentiation primary response protein (MYD88), both of which are involved in the recruitment and differentiation of myeloid cells. Taken together, our results suggest the recruitment of myeloid cells as an upstream pathway in the pathogenesis of IU.This study provides insights into proteins that will serve as biomarkers and therapeutic targets for IU. These biomarkers will help design future clinical trials using rational molecular therapeutics.

View details for DOI 10.1016/j.ajoc.2020.100646

View details for PubMedID 32274442

View details for PubMedCentralID PMC7132169
Modulation of post-traumatic immune response using the IL-1 receptor antagonist anakinra for improved visual outcomes. Journal of neurotrauma Evans, L. P., Woll, A. W., Wu, S., Todd, B. P., Hehr, N. N., Hedberg-Buenz, A., Anderson, M. G., Newell, E. A., Ferguson, P. J., Mahajan, V. B., Harper, M. M., Bassuk, A. G. 2020
Abstract

The purpose of this study was to characterize acute changes in inflammatory pathways in the mouse eye following a blast-mediated TBI (bTBI) model, and to determine if modulation of these pathways could protect the structure and function of retinal ganglion cells (RGC). bTBI was induced in C57BL/6J male mice by exposure to three 20 PSI blast waves directed towards the head with the body shielded, with an inter-blast interval of one hour. Acute cytokine expression in retinal tissue was measured through real-time quantitative polymerase chain reaction (RT-qPCR) 4 hours post-blast. Increased retinal expression of IL-1α, IL-1β, IL-6, and TNFα was observed in bTBI mice exposed to blast when compared to shams, which was associated with activation of microglia and macroglia reactivity, assessed via immunohistochemistry with IBA-1 and GFAP, respectively, 1 week post-blast. Blockade of the IL-1 pathway was accomplished using anakinra, an IL-1RI antagonist, administered intraperitoneally for 1 week prior to injury and continuing for 3 weeks post-injury. Retinal function and RGC layer thickness were evaluated 4 weeks post-injury using pattern electroretinogram (PERG) and optical coherence tomography (OCT), respectively. After bTBI, anakinra treatment resulted in a preservation of RGC function and RGC structure when compared to saline treated bTBI mice. Optic nerve integrity analysis demonstrated a trend of decreased damage suggesting that IL-1 blockade also prevents axonal damage after blast. Blast exposure results in increased retinal inflammation including upregulation of pro-inflammatory cytokines and activation of resident microglia and macroglia. This may partially explain the RGC loss we observed in this model as blockade of the acute inflammatory response after injury with the IL-1R1 antagonist anakinra resulted in preservation of RGC function and structure. Key Words: Blast; IL-1; anakinra; retina; visual function.

View details for DOI 10.1089/neu.2019.6725

View details for PubMedID 32056479
Sex Does Not Influence Visual Outcomes After Blast-Mediated Traumatic Brain Injury but IL-1 Pathway Mutations Confer Partial Rescue. Investigative ophthalmology & visual science Evans, L. P., Boehme, N., Wu, S., Burghardt, E. L., Akurathi, A., Todd, B. P., Newell, E. A., Ferguson, P. J., Mahajan, V. B., Dutca, L. M., Harper, M. M., Bassuk, A. G. 2020; 61 (12): 7
Abstract

In a mouse model of blast-mediated traumatic brain injury (bTBI), interleukin-1 (IL-1)-pathway components were tested as potential therapeutic targets for bTBI-mediated retinal ganglion cell (RGC) dysfunction. Sex was also evaluated as a variable for RGC outcomes post-bTBI.Male and female mice with null mutations in genes encoding IL-1α, IL-1β, or IL-1RI were compared to C57BL/6J wild-type (WT) mice after exposure to three 20-psi blast waves given at an interblast interval of 1 hour or to mice receiving sham injury. To determine if genetic blockade of IL-1α, IL-1β, or IL-1RI could prevent damage to RGCs, the function and structure of these cells were evaluated by pattern electroretinogram and optical coherence tomography, respectively, 5 weeks following blast or sham exposure. RGC survival was also quantitatively assessed via immunohistochemical staining of BRN3A at the completion of the study.Our results showed that male and female WT mice had a similar response to blast-induced retinal injury. Generally, constitutive deletion of IL-1α, IL-1β, or IL-1RI did not provide full protection from the effects of bTBI on visual outcomes; however, injured WT mice had significantly worse visual outcomes compared to the injured genetic knockout mice.Sex does not affect RGC outcomes after bTBI. The genetic studies suggest that deletion of these IL-1 pathway components confers some protection, but global deletion from birth did not result in a complete rescue.

View details for DOI 10.1167/iovs.61.12.7

View details for PubMedID 33030508
Quantitative Autofluorescence Following Gene Therapy With Voretigene Neparvovec. JAMA ophthalmology Levi, S. R., Oh, J. K., de Carvalho, J. R., Mahajan, V. B., Tsang, S. H., Sparrow, J. R. 2020
View details for DOI 10.1001/jamaophthalmol.2020.2018

View details for PubMedID 32556084
Optical Gap Biomarker in Cone-Dominant Retinal Dystrophy. American journal of ophthalmology Oh, J. K., Ryu, J., Lima de Carvalho, J. R., Levi, S. R., Lee, W., Tsamis, E., Greenstein, V. C., Mahajan, V. B., Allikmets, R., Tsang, S. H. 2020
Abstract

To characterize the progression of optical gaps and expand the known etiologies of this phenotype.Retrospective cohort study.Thirty-six patients were selected based on the identification of an optical gap on spectral-domain optical coherence tomography (SD-OCT) from a large cohort of patients (n=746) with confirmed diagnoses of inherited retinal dystrophy. The width and height of the gaps in 70 eyes of 36 patients were measured using the caliper tool on Heidelberg Explorer by two independent graders. Measurements of outer and central retinal thickness were also evaluated and correlated with gap dimensions.Longitudinal analysis confirmed the progressive nature of optical gaps in patients with Stargardt disease, achromatopsia, occult macular dystrophy, and cone dystrophies (p<0.003). Larger changes in gap width were noted in patients with Stargardt disease (78.1μm/year) and cone dystrophies (31.9μm/year) as compared to patients with achromatopsia (16.2μm/year) and occult macular dystrophy (15.4μm/year). Gap height decreased in patients with Stargardt disease (6.5μm/year) (p=0.02), but increased in patients with achromatopsia (3.3μm/year) and occult macular dystrophy (1.2μm/year). Gap height correlated with measurements of central retinal thickness at the fovea (r=0.782, p=0.00012). Interocular discordance of the gap was observed in 7 patients. Finally, a review of all currently described etiologies of optical gap was summarized.The optical gap is a progressive phenotype seen in an increasing number of etiologies. This progressive nature suggests a use as a biomarker in the understanding of disease progression. Interocular discordance of the phenotype may be a feature of Stargardt disease and cone dystrophies.

View details for DOI 10.1016/j.ajo.2020.05.016

View details for PubMedID 32445700
Intravitreal methotrexate and fluocinolone acetonide implantation for Vogt-Koyanagi-Harada uveitis. American journal of ophthalmology case reports Park, J. G., Callaway, N. F., Ludwig, C. A., Mahajan, V. B. 2020; 19: 100859
Abstract

To report a case of intravitreal methotrexate treatment and fluocinolone acetonide (Retisert®) implantation in a patient with Vogt-Koyanagi-Harada syndrome (VKH).A 34-year-old male was referred for worsening vision and bilateral panuveitis consistent with VKH. He was treated with prednisone, mycophenolate mofetil, prednisolone acetate eye drops, and injections of triamcinolone and adalimumab. He failed to improve with these therapies and developed multiple adverse effects, including hepatotoxicity, severe eye pain, cataracts, and cystoid macular edema. We treated him with intravitreal methotrexate injections in both eyes, which rapidly improved his eye pain, inflammation, and vision. He subsequently underwent fluocinolone acetonide (Retisert®) implantation, cataract extraction with intraocular lens insertion, and Ahmed tube placement for long-term intraocular pressure control. His vision improved from hand motions to 20/30, intraocular pressure remained stable at 17, there was complete resolution of his panuveitis and uveitic macular edema, and his systemic medications were able to be discontinued./Importance: This case demonstrates intravitreal methotrexate may successfully treat intraocular inflammation, pain, and macular edema in VKH. Excellent long-term vision and reduction of adverse effects of systemic medications were also achieved with subsequent fluocinolone acetonide implantation. Combining these two targeted therapies may be an effective strategy in treating VKH in patients who have severe pain and cannot tolerate systemic therapy.

View details for DOI 10.1016/j.ajoc.2020.100859

View details for PubMedID 32793845

View details for PubMedCentralID PMC7415829
Sex Differences in the Repair of Retinal Detachments in the United States. American journal of ophthalmology Callaway, N. F., Vail, D., Al-Moujahed, A., Ludwig, C., Ji, M. H., Mahajan, V. B., Pershing, S., Moshfeghi, D. M. 2020
Abstract

To determine differences between women and men in the repair of rhegmatogenous retinal detachments (RRD) in the United States.Retrospective cohort study.SETTING: A large insurance claims database.Subjects with an incident RRD between 2007-2015.Demographic data, comorbid ocular conditions associated with RRD,systemic comorbidities,and surgical intervention (pneumatic retinopexy (PR), pars plana vitrectomy (PPV), laser barricade, or scleral buckle (SB)) were collected.Odds of receipt of surgical intervention for incident RRD, time to repair, type of intervention, and the rate of reoperation by sex.The study period included 133 million eligible records with 61,071 cases of incident RRD meeting inclusion criteria among which 43% (n = 26,289) were women. Women had 34% reduced odds of receipt of surgical repairof an RRD (OR 0.66, 95% CI 0.59 - 0.73, p<0.001) after adjusting for confounders. This effect persisted in all sensitivity models. Among patients that received repair, women were more often delayed (0.17 days, p = 0.04). Women were more likely to undergo primary laser barricade (RRR 1.68, p < 0.001), primary SB (RRR 1.15, p < 0.001), and PR (RRR 1.07, p < 0.04) than men. The odds of reoperation were lower in women (OR 0.91, 95% CI 0.85 - 0.96, p=0.002) after adjustment.Insured women are less likely than insured men to receive surgical intervention for an RRD. If the odds of repair were equal between women and men in the U.S. then 781 more women would receive surgery each year, or 7,029 more during the study period. Women are more likely to have the repair performed with scleral buckle and laser barricade. The reason for thesesex differences in RRD repair remains unknown and requires further investigation.

View details for DOI 10.1016/j.ajo.2020.06.039

View details for PubMedID 32640255
Fundoscopy-directed genetic testing to re-evaluate negative whole exome sequencing results. Orphanet journal of rare diseases Cho, A., Lima de Carvalho, J. R., Tanaka, A. J., Jauregui, R., Levi, S. R., Bassuk, A. G., Mahajan, V. B., Tsang, S. H. 2020; 15 (1): 32
Abstract

Whole exome sequencing (WES) allows for an unbiased search of the genetic cause of a disease. Employing it as a first-tier genetic testing can be favored due to the associated lower incremental cost per diagnosis compared to when using it later in the diagnostic pathway. However, there are technical limitations of WES that can lead to inaccurate negative variant callings. Our study presents these limitations through a re-evaluation of negative WES results using subsequent tests primarily driven by fundoscopic findings. These tests included targeted gene testing, inherited retinal gene panels, whole genome sequencing (WGS), and array comparative genomic hybridization.Subsequent genetic testing guided by fundoscopy findings identified the following variant types causing retinitis pigmentosa that were not detected by WES: frameshift deletion and nonsense variants in the RPGR gene, 353-bp Alu repeat insertions in the MAK gene, and large exonic deletion variants in the EYS and PRPF31 genes. Deep intronic variants in the ABCA4 gene causing Stargardt disease and the GUCY2D gene causing Leber congenital amaurosis were also identified.Negative WES analyses inconsistent with the phenotype should raise clinical suspicion. Subsequent genetic testing may detect genetic variants missed by WES and can make patients eligible for gene replacement therapy and upcoming clinical trials. When phenotypic findings support a genetic etiology, negative WES results should be followed by targeted gene sequencing, array based approach or whole genome sequencing.

View details for DOI 10.1186/s13023-020-1312-1

View details for PubMedID 32000842
Compound heterozygous novel frameshift variants in the PROM1 gene result in Leber congenital amaurosis. Cold Spring Harbor molecular case studies Ragi, S. D., Lima de Carvalho, J. R., Tanaka, A. J., Park, K. S., Mahajan, V. B., Maumenee, I. H., Tsang, S. H. 2019; 5 (6)
Abstract

The PROM1 (prominin 1) gene encodes an 865-amino acid glycoprotein that is expressed in retinoblastoma cell lines and in the adult retina. The protein is localized to photoreceptor outer segment disc membranes, where it plays a structural role, and in the retinal pigment epithelium (RPE), where it acts as a cytosolic protein that mediates autophagy. Mutations in PROM1 are typically associated with cone-rod dystrophy 12 (OMIM#3612657), autosomal dominant retinal macular dystrophy 2 (OMIM#608051), autosomal recessive retinitis pigmentosa 41 (OMIM#612095), and Stargardt disease 4 (OMIM#603786). Here we describe the first case of PROM1-associated Leber congenital amaurosis (LCA) in a 12-yr-old Asian male, caused by two not previously described deleterious frameshift variants in the compound heterozygous state. Clinical features include the presence of bull's eye maculopathy, pendular horizontal nystagmus, and photodysphoria consistent with the clinical diagnosis of LCA. The patient was evaluated using ophthalmic imaging, electroretinography, and whole-exome sequencing. Electroretinography revealed extinguished retinal activity.

View details for DOI 10.1101/mcs.a004481

View details for PubMedID 31836589
A Reversible Silicon Oil-Induced Ocular Hypertension Model in Mice. Journal of visualized experiments : JoVE Zhang, J., Fang, F., Li, L., Huang, H., Webber, H. C., Sun, Y., Mahajan, V. B., Hu, Y. 2019
Abstract

Elevated intraocular pressure (IOP) is a well-documented risk factor for glaucoma. Here we describe a novel, effective method for consistently inducing stable IOP elevation in mice that mimics the post-operative complication of using silicone oil (SO) as a tamponade agent in human vitreoretinal surgery. In this protocol, SO is injected into the anterior chamber of the mouse eye to block the pupil and prevent inflow of aqueous humor. The posterior chamber accumulates aqueous humor and this in turn increases the IOP of the posterior segment. A single SO injection produces reliable, sufficient, and stable IOP elevation, which induces significant glaucomatous neurodegeneration. This model is a true replicate of secondary glaucoma in the eye clinic. To further mimic the clinical setting, SO can be removed from the anterior chamber to reopen the drainage pathway and allow inflow of aqueous humor, which is drained through the trabecular meshwork (TM) at the angle of the anterior chamber. Because IOP quickly returns to normal, the model can be used to test the effect of lowering IOP on glaucomatous retinal ganglion cells. This method is straightforward, does not require special equipment or repeat procedures, closely simulates clinical situations, and may be applicable to diverse animal species. However, minor modifications may be required.

View details for DOI 10.3791/60409

View details for PubMedID 31789319
Limbal Trocar-Cannulas for Complex Vitrectomy Surgery RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Mears, K. A., Mahajan, V. B. 2019; 39: 119–22
View details for DOI 10.1097/IAE.0000000000001781

View details for Web of Science ID 000507481300028
Hypoxic drive caused Type 3 neovascularization in a preclinical model of exudative age-related macular degeneration. Human molecular genetics Zhang, L., Cui, X., Han, Y., Park, K. S., Gao, X., Zhang, X., Yuan, Z., Hu, Y., Hsu, C., Li, X., Bassuk, A. G., Mahajan, V. B., Wang, N., Tsang, S. H. 2019
Abstract

Hypoxia associated with the high metabolic demand of rods has been implicated in the pathology of age-related macular degeneration (AMD), the most common cause of adult blindness in the developed world. The majority of AMD-associated severe vision loss cases are due to exudative AMD, characterized by neovascularization. To further investigate the causes and histopathology of exudative AMD, we conditionally induced hypoxia in a novel preclinical AMD model (Pde6gcreERT2/+;Vhl-/-) by targeting Vhl and used multimodal imaging and immunohistochemistry to track the development of hypoxia-induced neovascularization. In addition to developing a preclinical model that phenocopies exudative AMD, our studies revealed that the photoreceptor hypoxic response initiates and drives Type 3 neovascularization, mainly in the outer retina. Activation of the VHL-HIF1a-VEGF-EPO pathway in the adult retina led to long-term neovascularization, retinal hemorrhages, and compromised retinal layers. Our novel preclinical model would accelerate the testing of therapies that use metabolomic approaches to ameliorate AMD.

View details for DOI 10.1093/hmg/ddz159

View details for PubMedID 31518400
Mechanisms of neurodegeneration in a preclinical autosomal dominant retinitis pigmentosa knock-in model with a Rho(D190N) mutation CELLULAR AND MOLECULAR LIFE SCIENCES Sancho-Pelluz, J., Cui, X., Lee, W., Tsai, Y., Wu, W., Justus, S., Washington, I., Hsu, C., Park, K., Koch, S., Velez, G., Bassuk, A. G., Mahajan, V. B., Lin, C., Tsang, S. H. 2019; 76 (18): 3657–65
View details for DOI 10.1007/s00018-019-03090-9

View details for Web of Science ID 000481801300012
CAPN5 genetic inactivation phenotype supports therapeutic inhibition trials. Human mutation Wert, K. J., Koch, S. F., Velez, G., Hsu, C., Mahajan, M., Bassuk, A. G., Tsang, S. H., Mahajan, V. B. 2019
Abstract

Small molecule pharmacological inhibition of dominant human genetic disease is a feasible treatment that does not rely on the development of individual, patient-specific gene therapy vectors. However, the consequences of protein inhibition as a clinical therapeutic are not well-studied. In advance of human therapeutic trials for CAPN5 vitreoretinopathy, genetic inactivation can be used to infer the effect of protein inhibition in vivo. We created a photoreceptor-specific knockout mouse for Capn5 and compared the retinal phenotype to both wild-type and an existing Capn5 knockout mouse model. In humans, CAPN5 loss-of-function gene variants were ascertained in large exome databases from 60,706 unrelated subjects without severe disease phenotypes. Ocular examination of the retina of Capn5 knockout mice by histology and electroretinography showed no significant abnormalities. In humans, there were 22 loss-of-function CAPN5 variants located throughout the gene and in all major protein domains. Structural modeling of coding variants showed these loss-of-function variants were nearby known disease-causing variants within the proteolytic core and in regions of high homology between human CAPN5 and 150 homologs, yet the loss-of-function of CAPN5 was tolerated as opposed to gain-of-function disease-causing variants. These results indicate that localized inhibition of CAPN5 is a viable strategy for hyperactivating disease alleles. This article is protected by copyright. All rights reserved.

View details for DOI 10.1002/humu.23894

View details for PubMedID 31403230
Ophthalmic specimens from bedside to bench: An integrated biorepository platform to bridge the gap between the operating room and laboratory Chemudupati, S., Mahajan, V. B., Mruthyunjaya, P. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2019
View details for Web of Science ID 000488800705218
Comparison of structural progression between ciliopathy and non-ciliopathy associated with autosomal recessive retinitis pigmentosa Xu, C. L., Takahashi, V., Ben Apatoff, M., Takiuti, J., Duong, J. K., Mahajan, V. B., Tsang, S. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2019
View details for Web of Science ID 000488800702264
Visual Function Questionnaire Responses in the Voretigene Neparvovec Phase 3 Trial Russell, S. R., Maguire, A. M., Bennett, J., Falabella, P., Sohn, E. H., Mahajan, V. B., Chung, D. C., Reape, K. Z., High, K. A. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2019
View details for Web of Science ID 000488800703306
Characterization of primary cilia in mouse retina during retinal development Ning, K., Kowal, T., Chang, K., Alvarado, J., Silva, R., Kreymerman, A., Mahajan, V. B., Hu, Y., Sun, Y. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2019
View details for Web of Science ID 000488800706182
CAPN5 genetic inactivation phenotype supports therapeutic inhibition trials Wert, K., Koch, S. F., Velez, G., Hsu, C., Mahajan, M., Bassuk, A. G., Tsang, S. H., Mahajan, V. B. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2019
View details for Web of Science ID 000488628100371
A novel inducible and reversible mouse glaucoma model: Silicone Oil-Induced Ocular Hypertension Under-detected (SOHU) Zhang, J., Li, L., Huang, H., Webber, H., Li, S., Tang, P., Mahajan, V. B., Sun, Y., Zhang, M., Hu, Y. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2019
View details for Web of Science ID 000488628101260
Therapeutic Window for Phosphodiesterase 6-Related Retinitis Pigmentosa JAMA OPHTHALMOLOGY Wang, N., Mahajan, V. B., Tsang, S. H. 2019; 137 (6): 679–80
View details for DOI 10.1001/jamaophthalmol.2018.6381

View details for Web of Science ID 000471750500017
Traumatic chorioretinitis sclopetaria: Risk factors, management, and prognosis. American journal of ophthalmology case reports Ludwig, C. A., Shields, R. A., Do, D. V., Moshfeghi, D. M., Mahajan, V. B. 2019; 14: 39–46
Abstract

Purpose: To describe new cases of sclopetaria and evaluate the risk factors, management, and visual prognosis of all reported cases in the literature.Observations: We performed a retrospective, observational case series. This study included six cases (median age 23, interquartile range 33) of sclopetaria. Additionally, literature searches were conducted in the PubMed and Cochrane Library databases to uncover risk factors associated with all published cases of sclopetaria. Main outcome measure was best corrected visual acuity (BCVA) worse than 20/20. Sixty-seven cases (71 eyes) of sclopetaria have been reported, of which 59 cases (61 eyes) met inclusion criteria in this study. Most were young (median age 19.5 years) men (51/59, 88.1%). Thirty-seven eyes were observed while 24 underwent immediate surgery including six pars plana vitrectomies and three scleral buckles. Compared to initial presentation, BCVA improved in 31/48 (64.6%) eyes, remained stable in 12/48 eyes (25.0%), and worsened in 5/48 eyes (10.4%). Ten patients (16.4%) achieved a final BCVA of 20/20 with median follow up time of seven months. In a multivariate model, location of sclopetaria in the macula, temporal retina, or immediate orbital foreign body removal predicted poor final BCVA with an area under receiver operating characteristic curve of 0.767.Conclusions and importance: Traumatic chorioretinitis sclopetaria is rare, but reports have increased dramatically over the past two decades. While pars plana vitrectomy may be required for the management of retinal detachments and non-clearing vitreous hemorrhage, close observation is appropriate in most cases. Visual prognosis is poor with most patients attaining 20/200 vision or worse.

View details for PubMedID 30834355
Gain-of-function mutations in a member of the Src family kinases cause autoinflammatory bone disease in mice and humans. Proceedings of the National Academy of Sciences of the United States of America Abe, K., Cox, A., Takamatsu, N., Velez, G., Laxer, R. M., Tse, S. M., Mahajan, V. B., Bassuk, A. G., Fuchs, H., Ferguson, P. J., Hrabe de Angelis, M. 2019
Abstract

Autoinflammatory syndromes are characterized by dysregulation of the innate immune response with subsequent episodes of acute spontaneous inflammation. Chronic recurrent multifocal osteomyelitis (CRMO) is an autoinflammatory bone disorder that presents with bone pain and localized swelling. Ali18 mice, isolated from a mutagenesis screen, exhibit a spontaneous inflammatory paw phenotype that includes sterile osteomyelitis and systemic reduced bone mineral density. To elucidate the molecular basis of the disease, positional cloning of the causative gene for Ali18 was attempted. Using a candidate gene approach, a missense mutation in the C-terminal region of Fgr, a member of Src family tyrosine kinases (SFKs), was identified. For functional confirmation, additional mutations at the N terminus of Fgr were introduced in Ali18 mice by CRISPR/Cas9-mediated genome editing. N-terminal deleterious mutations of Fgr abolished the inflammatory phenotype in Ali18 mice, but in-frame and missense mutations in the same region continue to exhibit the phenotype. The fact that Fgr null mutant mice are morphologically normal suggests that the inflammation in this model depends on Fgr products. Furthermore, the levels of C-terminal negative regulatory phosphorylation of Fgr Ali18 are distinctly reduced compared with that of wild-type Fgr. In addition, whole-exome sequencing of 99 CRMO patients including 88 trios (proband and parents) identified 13 patients with heterozygous coding sequence variants in FGR, including two missense mutant proteins that affect kinase activity. Our results strongly indicate that gain-of-function mutations in Fgr are involved in sterile osteomyelitis, and thus targeting SFKs using specific inhibitors may allow for efficient treatment of the disease.

View details for DOI 10.1073/pnas.1819825116

View details for PubMedID 31138708
Silicone oil-induced ocular hypertension and glaucomatous neurodegeneration in mouse ELIFE Zhang, J., Li, L., Huang, H., Fang, F., Webber, H. C., Zhuang, P., Liu, L., Dalal, R., Tang, P. H., Mahajan, V. B., Sun, Y., Li, S., Zhang, M., Goldberg, J. L., Hu, Y. 2019; 8
View details for DOI 10.7554/eLife.45881

View details for Web of Science ID 000468863200001
Silicone oil-induced ocular hypertension and glaucomatous neurodegeneration in mouse. eLife Zhang, J., Li, L., Huang, H., Fang, F., Webber, H. C., Zhuang, P., Liu, L., Dalal, R., Tang, P. H., Mahajan, V. B., Sun, Y., Li, S., Zhang, M., Goldberg, J. L., Hu, Y. 2019; 8
Abstract

Understanding the molecular mechanism of glaucoma and development of neuroprotectants are significantly hindered by the lack of a reliable animal model that accurately recapitulates human glaucoma. Here we sought to develop a mouse model for the secondary glaucoma that is often observed in humans after silicone oil (SO) blocks the pupil or migrates into the anterior chamber following vitreoretinal surgery. We observed significant intraocular pressure (IOP) elevation after intracameral injection of SO, and that SO removal allows IOP to return quickly to normal. This simple, inducible and reversible mouse ocular hypertension model shows dynamic changes of visual function that correlate with progressive RGC loss and axon degeneration. It may be applicable with only minor modifications to a range of animal species in which it will generate stable, robust IOP elevation and significant neurodegeneration that will facilitate selection of neuroprotectants and investigating the pathogenesis of ocular hypertension-induced glaucoma.

View details for PubMedID 31090540
Therapeutic Window for Phosphodiesterase 6-Related Retinitis Pigmentosa. JAMA ophthalmology Wang, N., Mahajan, V. B., Tsang, S. H. 2019
View details for PubMedID 30998807
Early Onset Neovascular Inflammatory Vitreoretinopathy Due to a De Novo CAPN5 Mutation: Report of a Case OCULAR IMMUNOLOGY AND INFLAMMATION O'Keefe, G., Hanif, A. M., Mahajan, V. B., Jain, N. 2019
View details for DOI 10.1080/09273948.2019.1582783

View details for Web of Science ID 000466103700001
Early Onset Neovascular Inflammatory Vitreoretinopathy Due to a De Novo CAPN5 Mutation: Report of a Case. Ocular immunology and inflammation O'Keefe, G., Hanif, A. M., Mahajan, V. B., Jain, N. 2019: 1–3
Abstract

Autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV) is a rare autoimmune condition that typically presents as progressive uveitis and vitreoretinal degeneration between the second and third decades of life. Though traditionally attributed to inherited mutations of the CAPN5 gene, few reports of de novo variants exist. This report of vision and hearing loss in a 3 year-old girl describes the youngest documented case of ADNIV due to a de novo pathogenic c.865C>T (p.Arg289Trp) CAPN5 variant, illustrating the early stages of this enigmatic disease process.

View details for PubMedID 30986125
Fundus autofluorescence and ellipsoid zone (EZ) line width can be an outcome measurement in RHO-associated autosomal dominant retinitis pigmentosa GRAEFES ARCHIVE FOR CLINICAL AND EXPERIMENTAL OPHTHALMOLOGY Takahashi, V. L., Takiuti, J. T., Carvalho-, J. L., Xu, C. L., Duong, J. K., Mahajan, V. B., Tsang, S. H. 2019; 257 (4): 725–31
View details for DOI 10.1007/s00417-018-04234-6

View details for Web of Science ID 000462942600008
SCAPER-associated nonsyndromic autosomal recessive retinitis pigmentosa AMERICAN JOURNAL OF MEDICAL GENETICS PART A Jauregui, R., Thomas, A. L., Liechty, B., Velez, G., Mahajan, V. B., Clark, L., Tsang, S. H. 2019; 179 (2): 312–16
View details for DOI 10.1002/ajmg.a.61001

View details for Web of Science ID 000459032300024
Optical Coherence Tomography Angiography of RPGR- Associated Retinitis Pigmentosa Suggests Foveal Avascular Zone is a Biomarker for Vision Loss OPHTHALMIC SURGERY LASERS & IMAGING RETINA Tang, P. H., Jauregui, R., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2019; 50 (2): E44–E48
Abstract

RPGR-associated retinitis pigmentosa (RP) is a progressive disease with retina degeneration. Optical coherence tomography angiography (OCTA) is an imaging technique that provides novel insights. The authors report two affected male siblings who underwent OCTA imaging. The area of the foveal avascular zone (FAZ) was measured. Although the younger sibling exhibited more advanced clinical disease, his visual acuity was superior to his older sibling. OCTA imaging revealed a better preserved FAZ in the younger sibling as the reason for this. It also highlighted attenuation of choriocapillaris / choroid layers as biomarkers for disease severity. This provides new insights into retinal degeneration in RPGR-associated RP. [Ophthalmic Surg Lasers Imaging Retina. 2019;50:e44-e48.].

View details for DOI 10.3928/23258160-20190129-18

View details for Web of Science ID 000458843200007

View details for PubMedID 30768229
Fundus autofluorescence and ellipsoid zone (EZ) line width can be an outcome measurement in RHO-associated autosomal dominant retinitis pigmentosa. Graefe's archive for clinical and experimental ophthalmology = Albrecht von Graefes Archiv fur klinische und experimentelle Ophthalmologie Takahashi, V. K., Takiuti, J. T., Carvalho-Jr, J. R., Xu, C. L., Duong, J. K., Mahajan, V. B., Tsang, S. H. 2019
Abstract

PURPOSE: To evaluate the progression of retinitis pigmentosa (RP) due to mutations in rhodopsin (RHO) by measuring the short-wavelength autofluorescence (SW-AF) increased autofluorescence ring and ellipsoid zone (EZ)-line width.METHODS: Fundus autofluorescence (FAF) and spectral domain optical coherence tomography (SD-OCT) images were obtained from 10 patients with autosomal dominant RP due to mutations in the RHO gene. Measurements of ring area on FAF images, as well as the EZ line width on SD-OCT images and horizontal, vertical diameter, were performed by two independent masked graders.RESULTS: The ring area decreased by a rate of 0.6±0.2mm2 per year. We observed that the EZ line width decreased by an average of 152±37mum per year, while the horizontal and vertical diameters decreased by 106±35mum and 125±29mum per year, respectively. Progression rates were similar between eyes.CONCLUSIONS: We observed SW-AF ring constriction and a progressive loss of EZ line width over time.

View details for PubMedID 30635721
Viral Delivery Systems for CRISPR. Viruses Xu, C. L., Ruan, M. Z., Mahajan, V. B., Tsang, S. H. 2019; 11 (1)
Abstract

The frontiers of precision medicine have been revolutionized by the development of Clustered Regularly-Interspaced Short Palindromic Repeats (CRISPR)/Cas9 as an editing tool. CRISPR/Cas9 has been used to develop animal models, understand disease mechanisms, and validate treatment targets. In addition, it is regarded as an effective tool for genome surgery when combined with viral delivery vectors. In this article, we will explore the various viral mechanisms for delivering CRISPR/Cas9 into tissues and cells, as well as the benefits and drawbacks of each method. We will also review the history and recent development of CRISPR and viral vectors and discuss their applications as a powerful tool in furthering our exploration of disease mechanisms and therapies.

View details for PubMedID 30621179
Bilateral Endophthalmitis after Immediately Sequential Bilateral Cataract Surgery. Ophthalmology. Retina Callaway, N. F., Ji, M. H., Mahajan, V. B., Moshfeghi, D. M. 2019
View details for DOI 10.1016/j.oret.2019.04.007

View details for PubMedID 31153851
Novel REEP6 gene mutation associated with autosomal recessive retinitis pigmentosa. Documenta ophthalmologica. Advances in ophthalmology Lin, Y., Xu, C. L., Velez, G., Yang, J., Tanaka, A. J., Breazzano, M. P., Mahajan, V. B., Sparrow, J. R., Tsang, S. H. 2019
Abstract

This study reports the ophthalmic and genetic findings of a Cameroonian patient with autosomal recessive retinitis pigmentosa (arRP) caused by a novel Receptor Expression Enhancing Protein 6 (REEP6) homozygous mutation.A 33-year-old man underwent comprehensive ophthalmic examinations, including visual acuity measurements, dilated fundus imaging, electroretinography (ERG), and spectral-domain optical coherence tomography (SD-OCT). Short-wavelength fundus autofluorescence (SW-AF) and near-infrared fundus autofluorescence (NIR-AF) were also evaluated. Whole exome sequencing (WES) was used to identify potential pathogenic variants.Fundus examination revealed typical RP findings with additional temporal ten micron yellow dots. SD-OCT imaging revealed cystoid macular edema and perifoveal outer retinal atrophy with centrally preserved inner segment ellipsoid zone (EZ) bands. Hyperreflective spots were seen in the inner retinal layers. On SW-AF images, a hypoautofluorescent area in the perifoveal area was observed. NIR-AF imaging revealed an irregularly shaped hyperautofluorescent ring. His visual acuity was mildly affected. ERG showed undetectable rod responses and intact cone responses. Genetic testing via WES revealed a novel homozygous mutation (c.295G>A, p.Glu99Lys) in the gene encoding REEP6, which is predicted to alter the charge in the transmembrane helix.This report is not only the first description of a Cameroonian patient with arRP associated with a REEP6 mutation, but also this particular genetic alteration. Substitution of p.Glu99Lys in REEP6 likely disrupts the interactions between REEP6 and the ER membrane. NIR-AF imaging may be particularly useful for assessing functional photoreceptor cells and show an "avocado" pattern of hyperautofluorescence in patients with the REEP6 mutation.

View details for DOI 10.1007/s10633-019-09719-1

View details for PubMedID 31538292
Opposing T cell responses in experimental autoimmune encephalomyelitis. Nature Saligrama, N., Zhao, F., Sikora, M. J., Serratelli, W. S., Fernandes, R. A., Louis, D. M., Yao, W., Ji, X., Idoyaga, J., Mahajan, V. B., Steinmetz, L. M., Chien, Y. H., Hauser, S. L., Oksenberg, J. R., Garcia, K. C., Davis, M. M. 2019
Abstract

Experimental autoimmune encephalomyelitis is a model for multiple sclerosis. Here we show that induction generates successive waves of clonally expanded CD4+, CD8+ and γδ+ T cells in the blood and central nervous system, similar to gluten-challenge studies of patients with coeliac disease. We also find major expansions of CD8+ T cells in patients with multiple sclerosis. In autoimmune encephalomyelitis, we find that most expanded CD4+ T cells are specific for the inducing myelin peptide MOG35-55. By contrast, surrogate peptides derived from a yeast peptide major histocompatibility complex library of some of the clonally expanded CD8+ T cells inhibit disease by suppressing the proliferation of MOG-specific CD4+ T cells. These results suggest that the induction of autoreactive CD4+ T cells triggers an opposing mobilization of regulatory CD8+ T cells.

View details for DOI 10.1038/s41586-019-1467-x

View details for PubMedID 31391585
VCAN Canonical Splice Site Mutation is Associated With Vitreoretinal Degeneration and Disrupts an MMP Proteolytic Site. Investigative ophthalmology & visual science Tang, P. H., Velez, G., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2019; 60 (1): 282–93
Abstract

To gain insight into the pathophysiology of vitreoretinal degeneration, the clinical course of three family members with Versican Vitreoretinopathy (VVR) is described, and a canonical splice site mutation in the gene encoding for versican (VCAN) protein was biochemically analyzed.A retrospective chart review, human eye histopathology, Sanger DNA sequencing, protein structural modeling, and in vitro proteolysis assays were performed.The proband (II:1), mother (I:2), and younger sibling (II:2) suffered retinal degeneration with foveal sparing and retinal detachments with proliferative vitreoretinopathy, features that were confirmed on histopathologic analysis. All affected members carried a heterozygous adenine to guanine variant (c.4004-2A>G) predicted to result in exon 8 skipping or the deletion of 13 amino acids at the beginning of the GAGβ chain (VCAN p.1335-1347). This deleted region corresponded to a putative MMP cleavage site, validated using fluorescence resonance energy transfer (FRET)-based proteolysis assays. Proteomic network analysis identified 10 interacting partners in the human vitreous and retina linked to retinal detachment and degeneration.VVR causes significant ocular disease, including retinal detachment and retinal dystrophy. The intronic VCAN mutation removes an MMP cleavage site, which alters versican structure and results in abnormal vitreous modeling. Disruption of a versican protein network may underlie clinicopathologic disease features and point to targeted therapies.

View details for PubMedID 30657523
In Vivo Expression of Mutant Calpains in the Eye Using Lentivirus. Methods in molecular biology (Clifton, N.J.) Wert, K. J., Mahajan, V. B. 2019; 1915: 233–47
Abstract

Exome sequencing has identified many candidate genes and mutations for human diseases, but the functional validation of these candidates is a time-consuming and costly process. Here, we describe a method which uses lentiviruses to overexpress calpain mutations that may play a role in dominant diseases such as autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV). The use of lentivirus to deliver the mutant calpain allows for a cost-effective, rapid, and efficient approach to test whether or not a candidate gene mutation from exome sequencing acts as the disease-causing allele for a human disorder. This method also provides for a comparison of different candidate mutations from a single gene identified by exome sequencing, as well as elucidating the mechanisms underlying these complex human disorders. Furthermore, this chapter focuses on two different methods to deliver mutant calpain to the cells of the eye, using either a subretinal or an intravitreal injection of the lentivirus into the mouse eye.

View details for PubMedID 30617808
Mechanisms of neurodegeneration in a preclinical autosomal dominant retinitis pigmentosa knock-in model with a RhoD190N mutation. Cellular and molecular life sciences : CMLS Sancho-Pelluz, J., Cui, X., Lee, W., Tsai, Y. T., Wu, W. H., Justus, S., Washington, I., Hsu, C. W., Park, K. S., Koch, S., Velez, G., Bassuk, A. G., Mahajan, V. B., Lin, C. S., Tsang, S. H. 2019
Abstract

D190N, a missense mutation in rhodopsin, causes photoreceptor degeneration in patients with autosomal dominant retinitis pigmentosa (adRP). Two competing hypotheses have been developed to explain why D190N rod photoreceptors degenerate: (a) defective rhodopsin trafficking prevents proteins from correctly exiting the endoplasmic reticulum, leading to their accumulation, with deleterious effects or (b) elevated mutant rhodopsin expression and unabated signaling causes excitotoxicity. A knock-in D190N mouse model was engineered to delineate the mechanism of pathogenesis. Wild type (wt) and mutant rhodopsin appeared correctly localized in rod outer segments of D190N heterozygotes. Moreover, the rhodopsin glycosylation state in the mutants appeared similar to that in wt mice. Thus, it seems plausible that the injurious effect of the heterozygous mutation is not related to mistrafficking of the protein, but rather from constitutive rhodopsin activity and a greater propensity for chromophore isomerization even in the absence of light.

View details for PubMedID 30976840
In trans variant calling reveals enrichment for compound heterozygous variants in genes involved in neuronal development and growth. Genetics research Cox, A. J., Grady, F., Velez, G., Mahajan, V. B., Ferguson, P. J., Kitchen, A., Darbro, B. W., Bassuk, A. G. 2019; 101: e8
Abstract

Compound heterozygotes occur when different variants at the same locus on both maternal and paternal chromosomes produce a recessive trait. Here we present the tool VarCount for the quantification of variants at the individual level. We used VarCount to characterize compound heterozygous coding variants in patients with epileptic encephalopathy and in the 1000 Genomes Project participants. The Epi4k data contains variants identified by whole exome sequencing in patients with either Lennox-Gastaut Syndrome (LGS) or infantile spasms (IS), as well as their parents. We queried the Epi4k dataset (264 trios) and the phased 1000 Genomes Project data (2504 participants) for recessive variants. To assess enrichment, transcript counts were compared between the Epi4k and 1000 Genomes Project participants using minor allele frequency (MAF) cutoffs of 0.5 and 1.0%, and including all ancestries or only probands of European ancestry. In the Epi4k participants, we found enrichment for rare, compound heterozygous variants in six genes, including three involved in neuronal growth and development - PRTG (p = 0.00086, 1% MAF, combined ancestries), TNC (p = 0.022, 1% MAF, combined ancestries) and MACF1 (p = 0.0245, 0.5% MAF, EU ancestry). Due to the total number of transcripts considered in these analyses, the enrichment detected was not significant after correction for multiple testing and higher powered or prospective studies are necessary to validate the candidacy of these genes. However, PRTG, TNC and MACF1 are potential novel recessive epilepsy genes and our results highlight that compound heterozygous variants should be considered in sporadic epilepsy.

View details for DOI 10.1017/S0016672319000065

View details for PubMedID 31190668
Viral Delivery Systems for CRISPR VIRUSES-BASEL Xu, C. L., Ruan, M. C., Mahajan, V. B., Tsang, S. H. 2019; 11 (1)
View details for DOI 10.3390/v11010028

View details for Web of Science ID 000459132000028
VCAN Canonical Splice Site Mutation is Associated With Vitreoretinal Degeneration and Disrupts an MMP Proteolytic Site INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE Tang, P. H., Velez, G., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2019; 60 (1): 282–93
View details for DOI 10.1167/iovs.18-25624

View details for Web of Science ID 000456092300026
Proteomic insight into the pathogenesis of CAPN5-vitreoretinopathy. Scientific reports Velez, G., Yang, J., Li, A. S., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2019; 9 (1): 7608
Abstract

CAPN5 Neovascular Inflammatory Vitreoretinopathy (CAPN5-NIV; OMIM 193235) is a poorly-understood rare, progressive inflammatory intraocular disease with limited therapeutic options. To profile disease effector proteins in CAPN5-NIV patient vitreous, liquid vitreous biopsies were collected from two groups: eyes from control subjects (n = 4) with idiopathic macular holes (IMH) and eyes from test subjects (n = 12) with different stages of CAPN5-NIV. Samples were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Protein expression changes were evaluated by principal component analysis, 1-way ANOVA (significant p-value < 0.05), hierarchical clustering, gene ontology, and pathway representation. There were 216 differentially-expressed proteins (between CAPN5-NIV and control vitreous), including those unique to and abundant in each clinical stage. Gene ontology analysis revealed decreased synaptic signaling proteins in CAPN5-NIV vitreous compared to controls. Pathway analysis revealed that inflammatory mediators of the acute phase response and the complement cascade were highly-represented. The CAPN5-NIV vitreous proteome displayed characteristic enrichment of proteins and pathways previously-associated with non-infectious posterior uveitis, rhegmatogenous retinal detachment (RRD), age-related macular degeneration (AMD), proliferative diabetic retinopathy (PDR), and proliferative vitreoretinopathy (PVR). This study expands our knowledge of affected molecular pathways in CAPN5-NIV using unbiased, shotgun proteomic analysis rather than targeted detection platforms. The high-levels and representation of acute phase response proteins suggests a functional role for the innate immune system in CAPN5-NIV pathogenesis.

View details for DOI 10.1038/s41598-019-44031-7

View details for PubMedID 31110225
Comparison of structural progression between ciliopathy and non-ciliopathy associated with autosomal recessive retinitis pigmentosa. Orphanet journal of rare diseases Takahashi, V. K., Xu, C. L., Takiuti, J. T., Apatoff, M. B., Duong, J. K., Mahajan, V. B., Tsang, S. H. 2019; 14 (1): 187
Abstract

To evaluate and compare the progression of ciliopathy and non-ciliopathy autosomal recessive Retinitis Pigmentosa patients (arRP) by measuring the constriction of hyperautofluorescent rings in fundus autofluorescence (FAF) images and the progressive shortening of the ellipsoid zone line width obtained by spectral-domain optical coherence tomography (SD-OCT).For the ciliopathy group, the estimated mean shortening of the ellipsoid zone line was 259 μm per year and the ring area decreased at a rate of 2.46 mm2 per year. For the non-ciliopathy group, the estimated mean shortening of the ellipsoid zone line was 84 μm per year and the ring area decreased at a rate of 0.7 mm2 per year.Our study was able to quantify and compare the loss of EZ line width and short-wavelength autofluorescence (SW-AF) ring constriction progression over time for ciliopathy and non-ciliopathy arRP genes. These results may serve as a basis for modeling RP disease progression, and furthermore, they could potentially be used as endpoints in clinical trials seeking to promote cone and rod survival in RP patients.

View details for DOI 10.1186/s13023-019-1163-9

View details for PubMedID 31370859
CRISPR Base Editing in Induced Pluripotent Stem Cells. Methods in molecular biology (Clifton, N.J.) Chang, Y. J., Xu, C. L., Cui, X., Bassuk, A. G., Mahajan, V. B., Tsai, Y. T., Tsang, S. H. 2019
Abstract

Induced pluripotent stem cells (iPSCs) have demonstrated tremendous potential in numerous disease modeling and regenerative medicine-based therapies. The development of innovative gene transduction and editing technologies has further augmented the potential of iPSCs. Cas9-cytidine deaminases, for example, have developed as an alternative strategy to integrate single-base mutations (C → T or G → A transitions) at specific genomic loci. In this chapter, we specifically describe CRISPR (clustered regularly interspaced short palindromic repeats) base editing in iPSCs for editing precise locations in the genome. This state-of-the-art approach enables highly efficient and accurate modifications in genes. Thus, this technique not only has the potential to have biotechnology and therapeutic applications but also the ability to reveal underlying mechanisms regarding pathologies caused by specific mutations.

View details for DOI 10.1007/7651_2019_243

View details for PubMedID 31250381
Novel mutations in the 3-box motif of the BACK domain of KLHL7 associated with nonsyndromic autosomal dominant retinitis pigmentosa. Orphanet journal of rare diseases Oh, J. K., Lima de Carvalho, J. R., Sun, Y. J., Ragi, S., Yang, J., Levi, S. R., Ryu, J., Bassuk, A. G., Mahajan, V. B., Tsang, S. H. 2019; 14 (1): 295
Abstract

Mutations in the Kelch-like protein 7 (KLHL7) represent a recently described and, to date, poorly characterized etiology of inherited retinal dystrophy. Dominant mutations in KLHL7 are a cause of isolated, non-syndromic retinitis pigmentosa (RP). In contrast, recessive loss-of-function mutations are known to cause Crisponi or Bohring-Opitz like cold induced sweating syndrome-3 (BOS-3). In this study, the phenotype and progression of five unrelated patients with KLHL7 mediated autosomal dominant RP (adRP) are characterized. Clinical evaluation of these patients involved a complete ophthalmic exam, full-field electroretinography (ffERG), and imaging, including fundus photography, spectral domain optical coherence tomography (SD-OCT), short wavelength fundus autofluorescence (SW-AF), and near-infrared fundus autofluorescence (NIR-AF). Molecular diagnoses were performed using whole-exome sequencing or gene panel testing. Disease progression was monitored in three patients with available data for a mean follow up time of 4.5 ± 2.9 years. Protein modeling was performed for all variants found in this study in addition to those documented in the literature for recessive loss-of-function alleles causing Crisponi or Bohring-Opitz like cold-induced sweating syndrome.Genetic testing in three patients identified two novel variants within the 3-box motif of the BACK domain: c.472 T > C:p.(Cys158Arg) and c.433A > T:p.(Asn145Tyr). Clinical imaging demonstrated hyperautofluorescent ring formation on both SW-AF and NIR-AF in three patients, with diffuse peripheral and peripapillary atrophy seen in all but one case. SD-OCT demonstrated a phenotypic spectrum, from parafoveal atrophy of the outer retina with foveal sparing to widespread retinal thinning and loss of photoreceptors. Incidence of cystoid macular edema was high with four of five patients affected. Protein modeling of dominant alleles versus recessive loss-of-function alleles showed dominant alleles localized to the BTB and BACK domains while recessive alleles were found in the Kelch domain.We report the phenotype in five patients with KLHL7 mediated adRP, two novel coding variants, and imaging biomarkers using SW-AF and NIR-AF. These findings may influence future gene-based therapies for adRP and pave the way for mechanistic studies that elucidate the pathogenesis of KLHL7-mediated RP.

View details for DOI 10.1186/s13023-019-1275-2

View details for PubMedID 31856884
SCAPER-associated nonsyndromic autosomal recessive retinitis pigmentosa. American journal of medical genetics. Part A Jauregui, R., Thomas, A. L., Liechty, B., Velez, G., Mahajan, V. B., Clark, L., Tsang, S. H. 2018
Abstract

Mutations in the gene SCAPER (S-phase CyclinA Associated Protein residing in the Endoplasmic Reticulum) have recently been identified as causing syndromic autosomal recessive retinitis pigmentosa with the extraocular manifestations of intellectual disability and attention-deficit/hyperactivity disorder. We present the case of an 11-year-old boy that presented to our clinic with the complaint of decreased night vision. Clinical presentation, family history, and diagnostic imaging were congruent with the diagnosis of autosomal recessive retinitis pigmentosa. Genetic testing of the patient and both parents via whole-exome sequencing revealed the homozygous mutation c.2023-2A>G in SCAPER. Unique to our patient's presentation is the absence of intellectual disability and attention-deficit/hyperactivity disorder, suggesting that SCAPER-associated retinitis pigmentosa can also present without systemic manifestations.

View details for PubMedID 30561111
Review of Ocular Manifestations of Joubert Syndrome. Genes Wang, S. F., Kowal, T. J., Ning, K., Koo, E. B., Wu, A. Y., Mahajan, V. B., Sun, Y. 2018; 9 (12)
Abstract

Joubert syndrome is a group of rare disorders that stem from defects in a sensory organelle, the primary cilia. Affected patients often present with disorders involving multiple organ systems, including the brain, eyes, and kidneys. Common symptoms include breathing abnormalities, mental developmental delays, loss of voluntary muscle coordination, and abnormal eye movements, with a diagnostic "molar tooth" sign observed by magnetic resonance imaging (MRI) of the midbrain. We reviewed the ocular phenotypes that can be found in patients with Joubert syndrome. Ocular motor apraxia is the most frequent (80% of patients), followed by strabismus (74%) and nystagmus (72%). A minority of patients also present with ptosis (43%), chorioretinal coloboma (30%), and optic nerve atrophy (22%). Although mutations in 34 genes have been found to be associated with Joubert syndrome, retinal degeneration has been reported in only 38% of patients. Mutations in AHI1 and CEP290, genes critical to primary cilia function, have been linked to retinal degeneration. In conclusion, Joubert syndrome is a rare pleiotropic group of disorders with variable ocular presentations.

View details for PubMedID 30518138
Review of Ocular Manifestations of Joubert Syndrome GENES Wang, S. F., Kowal, T. J., Ning, K., Koo, E. B., Wu, A. Y., Mahajan, V. B., Sun, Y. 2018; 9 (12)
View details for DOI 10.3390/genes9120605

View details for Web of Science ID 000454717800039
ProSave: an application for restoring quantitative data to manipulated subsets of protein lists SOURCE CODE FOR BIOLOGY AND MEDICINE Machlab, D. A., Velez, G., Bassuk, A. G., Mahajan, V. B. 2018; 13
View details for DOI 10.1186/s13029-018-0070-0

View details for Web of Science ID 000449817600001
Rates of Bone Spicule Pigment Appearance in Patients With Retinitis Pigmentosa Sine Pigmento AMERICAN JOURNAL OF OPHTHALMOLOGY Takahashi, V. L., Takiuti, J. T., Jauregui, R., Mahajan, V. B., Tsang, S. H. 2018; 195: 176–80
Abstract

To determine rate of bone spicule pigmentation appearance in patients with retinitis pigmentosa (RP).Retrospective, observational case series.A total of 240 patients were analyzed for this study.A retrospective analysis was conducted at the Electrodiagnostic Clinic at Columbia University Medical Center of all patients' medical records with a diagnosis of RP between July 2017 and January 2018. The medical records of these patients were analyzed to determine whether the patients presented with pigment migration on their first and last visit to our clinic. Among those who did not have bone spicule at first visit, we examined the time to appearance of newly formed bone spicule. The survival distribution was then estimated using the Kaplan-Meier estimator, where the event is bone spicule and time starts at first visit.From the 240 patients analyzed, 213 patients presented with intraretinal pigmentation on the first visit to our clinic, and 27 patients presented without intraretinal pigmentation. Of these 27 patients, 10 patients developed pigmentation by their follow-up, with a median time to appearance of bone spicule of 5.4 years from first visit, according to the Kaplan-Meier estimates.The timeline of bone spicule pigment appearance in RP has important implications in the natural history characterization of disease progression and application as a biomarker for interventional trials.

View details for PubMedID 30081015
PROGRESSION OF SCOTOPIC SINGLE-FLASH ELECTRORETINOGRAPHY IN THE STAGES OF CAPN5 VITREORETINOPATHY. Retinal cases & brief reports Tang, P. H., Kinnick, T. R., Folk, J. C., Mahajan, M., Bassuk, A. G., Tsang, S. H., Mahajan, V. B. 2018
Abstract

PURPOSE: To characterize the changes found in the electroretinography (ERG) recordings of patients with autosomal dominant neovascular inflammatory vitreoretinopathy and correlate with clinical stages of the disease.METHODS: Retrospective chart review. Bright- and dim-flash full-field scotopic, photopic, and 30-Hz flicker ERGs were obtained according to international standards. The scotopic ERGs were further processed to analyze the oscillatory potential. The patient described in the case report underwent full ERG testing; five patients composed the archival case series data and included scotopic ERG recordings.RESULTS: Stage I autosomal dominant neovascular inflammatory vitreoretinopathy is characterized by a decrease in the b-wave amplitude on scotopic flash ERG and the disappearance of late OPs; however, the a-wave amplitude is normal. In Stage II, attenuation of early OPs and the c-wave are observed in scotopic ERG recordings, but both a- and b-wave amplitudes are unchanged. For patients in Stage III, there is a continued decline of both a- and b-wave amplitudes in scotopic ERG recordings. There was a loss of recordable scotopic ERG response in patients with Stage IV disease.CONCLUSION: Electroretinography may be valuable in determining optimal timing for therapeutic intervention and response before loss of recordable retinal function in CAPN5 vitreoretinopathy.

View details for PubMedID 30300311
Quantitative progression of retinitis pigmentosa by optical coherence tomography angiography SCIENTIFIC REPORTS Jauregui, R., Park, K., Duong, J. K., Mahajan, V. B., Tsang, S. H. 2018; 8: 13130
Abstract

Optical coherence tomography angiography (OCT-A) is a non-invasive alternative to fluorescein angiography that allows for the study of the retinal and choroidal vasculatures. In this retrospective cohort study of 28 patients with retinitis pigmentosa (RP), we used OCT-A to quantify changes in perfusion density, foveal avascular zone (FAZ) area, and choriocapillaris blood flow over time and correlated these variables with ellipsoid zone (EZ) line width and best-corrected visual acuity (BCVA). Perfusion density decreased by 2.42 ± 0.62% per year at the superior capillary plexus (SCP) (P = 0.001) and 2.41 ± 0.76% per year at the deep capillary plexus (DCP) (P = 0.004). FAZ area increased by 0.078 ± 0.021 mm2 per year (P = 0.001) at the SCP and 0.152 ± 0.039 mm2 per year (P = 0.001) at the DCP. No changes were observed in the choriocapillaris blood flow. EZ line width had the strongest correlation to perfusion density at the SCP (r = 0.660 and 0.635, first and second visit, respectively, P = 0.001), while BCVA most strongly correlated with FAZ area at the SCP (r = 0.679 and 0.548, P = 0.001 and 0.003). Our results suggest that OCT-A is a useful tool for monitoring RP disease progression and may be used to measure retinal vascular parameters as outcomes in clinical trials.

View details for PubMedID 30177829
Personalized Proteomics for Precision Health: Identifying Biomarkers of Vitreoretinal Disease TRANSLATIONAL VISION SCIENCE & TECHNOLOGY Velez, G., Tang, P. H., Cabral, T., Cho, G. Y., Machlab, D. A., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2018; 7 (5)
View details for DOI 10.1167/tvst.7.5.12

View details for Web of Science ID 000445701000004
Personalized Proteomics for Precision Health: Identifying Biomarkers of Vitreoretinal Disease. Translational vision science & technology Velez, G., Tang, P. H., Cabral, T., Cho, G. Y., Machlab, D. A., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2018; 7 (5): 12
Abstract

Proteomic analysis is an attractive and powerful tool for characterizing the molecular profiles of diseased tissues, such as the vitreous. The complexity of data available for analysis ranges from single (e.g., enzyme-linked immunosorbent assay [ELISA]) to thousands (e.g., mass spectrometry) of proteins, and unlike genomic analysis, which is limited to denoting risk, proteomic methods take snapshots of a diseased vitreous to evaluate ongoing molecular processes in real time. The proteome of diseased ocular tissues was recently characterized, uncovering numerous biomarkers for vitreoretinal diseases and identifying protein targets for approved drugs, allowing for drug repositioning. These biomarkers merit more attention regarding their therapeutic potential and prospective validation, as well as their value as reproducible, sensitive, and specific diagnostic markers.Translational Relevance: Personalized proteomics offers many advantages over alternative precision-health platforms for the diagnosis and treatment of vitreoretinal diseases, including identification of molecular constituents in the diseased tissue that can be targeted by available drugs.

View details for PubMedID 30271679
Extracellular superoxide dismutase (SOD3) regulates oxidative stress at the vitreoretinal interface FREE RADICAL BIOLOGY AND MEDICINE Wert, K. J., Velez, G., Cross, M. R., Wagner, B. A., Teoh-Fitzgerald, M. L., Buettner, G. R., McAnany, J., Olivier, A., Tsang, S. H., Harper, M. M., Domann, F. E., Bassuk, A. G., Mahajan, V. B. 2018; 124: 408–19
View details for DOI 10.1016/j.freeradbiomed.2018.06.024

View details for Web of Science ID 000441516300039
Missense mutation in SLIT2 associated with congenital myopia, anisometropia, connective tissue abnormalities, and obesity ORPHANET JOURNAL OF RARE DISEASES Liu, K. Y., Sengillo, J. D., Velez, G., Jauregui, R., Sakai, L. Y., Maumenee, I. H., Bassuk, A. G., Mahajan, V. B., Tsang, S. H. 2018; 13: 138
Abstract

SLIT2 is a protein ligand for the Roundabout (ROBO) receptor and was found to play a major role in repulsive midline axon guidance in central nervous system development. Based on studies utilizing knockout models, it has been postulated that SLIT2 is important for preventing inappropriate axonal routing during mammalian optic chiasm development.Case report.Here, we report a case of congenital myopia, anisometropia, and obesity in a patient with a SLIT2 point mutation. Examination of the patient's skin biopsy revealed abnormalities in elastin and collagen fibrils that suggest an underlying connective tissue disorder. Structural modeling placed the novel mutation (p.D1407G) in the EGF-like domain 8 and was predicted to affect interactions with SLIT2 binding partners.To the authors' knowledge, this is the first report of a SLIT2 variant in the context of these ocular findings.

View details for PubMedID 30111362
Deferoxamine-induced electronegative ERG responses DOCUMENTA OPHTHALMOLOGICA Jauregui, R., Park, K., Bassuk, A. G., Mahajan, V. B., Tsang, S. H. 2018; 137 (1): 15–23
Abstract

To report a case of deferoxamine-induced retinopathy characterized by electroretinography (ERG), optical coherence tomography angiography (OCT-A), and other multimodal imaging.This is an observational case report of one patient. Full-field ERG was performed. OCT-A, spectral-domain optical coherence tomography (SD-OCT), color fundus photography, and fundus autofluorescence were used to characterize the retinopathy induced by deferoxamine use.A 64-year-old man with a history of β-thalassemia intermedia presented with worsening visual acuity, nyctalopia, and electronegative ERG. OCT-A revealed atrophy of the choriocapillaris in areas of hypoautofluorescence, corresponding to regions of retinal atrophy. SD-OCT showed disruption of the ellipsoid zone, granular hyperreflective deposits within the retinal pigment epithelium, thinning of the retinal layers, and extensive choroidal sclerosis and atrophy of the retinal pigment epithelium.Deferoxamine-induced retinopathy can manifest with electronegative maximal ERG responses, and OCT-A can be used to detect deferoxamine toxicity.

View details for PubMedID 29770904
CRISPR GENOME SURGERY IN THE RETINA IN LIGHT OF OFF-TARGETING RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Cho, G. Y., Schaefer, K. A., Bassuk, A. G., Tsang, S. H., Mahajan, V. B. 2018; 38 (8): 1443–55
Abstract

Recent concerns regarding the clinical utilization of clustered regularly interspaced short palindromic repeats (CRISPR) involve uncertainties about the potential detrimental effects that many arise due to unintended genetic changes, as in off-target mutagenesis, during CRISPR genome surgery. This review gives an overview of off-targeting detection methods and CRISPR's place in the clinical setting, specifically in the field of ophthalmology.As CRISPR utilization in the laboratory setting has increased, knowledge regarding CRISPR mechanisms including its off-target effects has also increased. Although a perfect method for achieving 100% specificity is yet to be determined, the past few years have seen many developments in off-targeting detection and in increasing efficacy of CRISPR tools.The CRISPR system has high potential to be an invaluable therapeutic tool as it has the ability to modify and repair pathogenic retinal lesions. Although it is not yet a perfect system, with further efforts to improve its specificity and efficacy along with careful screening of off-target mutations, CRISPR-mediated genome surgery potential can become maximized and applied to patients.

View details for PubMedID 29746416

View details for PubMedCentralID PMC6054556
HTRA1, an age-related macular degeneration protease, processes extracellular matrix proteins EFEMP1 and TSP1 AGING CELL Lin, M. K., Yang, J., Hsu, C., Gore, A., Bassuk, A. G., Brown, L. M., Colligan, R., Sengillo, J. D., Mahajan, V. B., Tsang, S. H. 2018; 17 (4): e12710
Abstract

High-temperature requirement protein A1 (HTRA1) is a serine protease secreted by a number of tissues including retinal pigment epithelium (RPE). A promoter variant of the gene encoding HTRA1 is part of a mutant allele that causes increased HTRA1 expression and contributed to age-related macular degeneration (AMD) in genomewide association studies. AMD is characterized by pathological development of drusen, extracellular deposits of proteins and lipids on the basal side of RPE. The molecular pathogenesis of AMD is not well understood, and understanding dysregulation of the extracellular matrix may be key. We assess the high-risk genotype at 10q26 by proteomic comparison of protein levels of RPE cells with and without the mutation. We show HTRA1 protein level is increased in high-risk RPE cells along with several extracellular matrix proteins, including known HTRA1 cleavage targets LTBP-1 and clusterin. In addition, two novel targets of HTRA1 have been identified: EFEMP1, an extracellular matrix protein mutated in Doyne honeycomb retinal dystrophy, a genetic eye disease similar to AMD, and thrombospondin 1 (TSP1), an inhibitor of angiogenesis. Our data support the role of RPE extracellular deposition with potential effects in compromised barrier to neovascularization in exudative AMD.

View details for PubMedID 29730901
RPGR-associated retinitis pigmentosa display unique outer retinal and choroidal vascular changes on optical coherence tomography angiography Tang, P., Tsang, S., Bassuk, A., Do, D. V., Mahajan, V. B. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2018
View details for Web of Science ID 000442912508143
Gene therapy and genome surgery in the retina JOURNAL OF CLINICAL INVESTIGATION DiCarlo, J. E., Mahajan, V. B., Tsang, S. H. 2018; 128 (6): 2177–88
Abstract

Precision medicine seeks to treat disease with molecular specificity. Advances in genome sequence analysis, gene delivery, and genome surgery have allowed clinician-scientists to treat genetic conditions at the level of their pathology. As a result, progress in treating retinal disease using genetic tools has advanced tremendously over the past several decades. Breakthroughs in gene delivery vectors, both viral and nonviral, have allowed the delivery of genetic payloads in preclinical models of retinal disorders and have paved the way for numerous successful clinical trials. Moreover, the adaptation of CRISPR-Cas systems for genome engineering have enabled the correction of both recessive and dominant pathogenic alleles, expanding the disease-modifying power of gene therapies. Here, we highlight the translational progress of gene therapy and genome editing of several retinal disorders, including RPE65-, CEP290-, and GUY2D-associated Leber congenital amaurosis, as well as choroideremia, achromatopsia, Mer tyrosine kinase- (MERTK-) and RPGR X-linked retinitis pigmentosa, Usher syndrome, neovascular age-related macular degeneration, X-linked retinoschisis, Stargardt disease, and Leber hereditary optic neuropathy.

View details for PubMedID 29856367

View details for PubMedCentralID PMC5983345
A novel de novo CAPN5 mutation in a patient with inflammatory vitreoretinopathy, hearing loss, and developmental delay COLD SPRING HARBOR MOLECULAR CASE STUDIES Velez, G., Bassuk, A. G., Schaefer, K. A., Brooks, B., Gakhar, L., Mahajan, M., Kahn, P., Tsang, S. H., Ferguson, P. J., Mahajan, V. B. 2018; 4 (3)
View details for DOI 10.1101/mcs.a002519

View details for Web of Science ID 000450956600006
A novel de novo CAPN5 mutation in a patient with inflammatory vitreoretinopathy, hearing loss, and developmental delay. Cold Spring Harbor molecular case studies Velez, G., Bassuk, A. G., Schaefer, K. A., Brooks, B., Gakhar, L., Mahajan, M., Kahn, P., Tsang, S. H., Ferguson, P. J., Mahajan, V. B. 2018; 4 (3)
Abstract

Mutations that activate the protease calpain-5 (CAPN5) cause a nonsyndromic adult-onset autoinflammatory eye disease characterized by uveitis, altered synaptic signaling, retinal degeneration, neovascularization, and intraocular fibrosis. We describe a pediatric patient with severe inflammatory vitreoretinopathy accompanied by hearing loss and developmental delay associated with a novel, de novo CAPN5 missense mutation (c.865C>T, p.Arg289Trp) that shows greater hyperactivation of the calpain protease, indicating a genotype-phenotype correlation that links mutation severity to proteolytic activity and the possibility of earlier onset syndromic disease with auditory and neurological abnormalities.

View details for PubMedID 29472286
Translation of CRISPR Genome Surgery to the Bedside for Retinal Diseases FRONTIERS IN CELL AND DEVELOPMENTAL BIOLOGY Xu, C. L., Cho, G. Y., Sengillo, J. D., Park, K. S., Mahajan, V. B., Tsang, S. H. 2018; 6: 46
Abstract

In recent years, there has been accelerated growth of clustered regularly interspaced short palindromic repeats (CRISPR) genome surgery techniques. Genome surgery holds promise for diseases for which a cure currently does not exist. In the field of ophthalmology, CRISPR offers possibilities for treating inherited retinal dystrophies. The retina has little regenerative potential, which makes treatment particularly difficult. For such conditions, CRISPR genome surgery methods have shown great potential for therapeutic applications in animal models of retinal dystrophies. Much anticipation surrounds the potential for CRISPR as a therapeutic, as clinical trials of ophthalmic genome surgery are expected to begin as early as 2018. This mini-review summarizes preclinical CRISPR applications in the retina and current CRISPR clinical trials.

View details for PubMedID 29876348
Caring for Hereditary Childhood Retinal Blindness ASIA-PACIFIC JOURNAL OF OPHTHALMOLOGY Jauregui, R., Cho, G. Y., Takahashi, V. L., Takiuti, J. T., Bassuk, A. G., Mahajan, V. B., Tsang, S. H. 2018; 7 (3): 183–91
Abstract

Inherited retinal diseases (IRDs) are a major cause of incurable familial blindness in the Western world. In the pediatric population, IRDs are a major contributor to the 19 million children worldwide with visual impairment. Unfortunately, the road to the correct diagnosis is often complicated in the pediatric population, as typical diagnostic tools such as fundus examination, electrodiagnostic studies, and other imaging modalities may be difficult to perform in the pediatric patient. In this review, we describe the most significant IRDs with onset during the pediatric years (ie, before the age of 18). We describe the pathogenesis, clinical presentation, and potential treatment of these diseases. In addition, we advocate the use of a pedigree (family medical history), electroretinography, and genetic testing as the 3 most crucial tools for the correct diagnosis of IRDs in the pediatric population.

View details for PubMedID 29536675
Acute vitreoretinal trauma and inflammation after traumatic brain injury in mice ANNALS OF CLINICAL AND TRANSLATIONAL NEUROLOGY Evans, L. P., Newell, E. A., Mahajan, M., Tsang, S. H., Ferguson, P. J., Mahoney, J., Hue, C. D., Vogel, E. W., Morrison, B., Arancio, O., Nichols, R., Bassuk, A. G., Mahajan, V. B. 2018; 5 (3): 240–51
Abstract

Limited attention has been given to ocular injuries associated with traumatic brain injury (TBI). The retina is an extension of the central nervous system and evaluation of ocular damage may offer a less-invasive approach to gauge TBI severity and response to treatment. We aim to characterize acute changes in the mouse eye after exposure to two different models of TBI to assess the utility of eye damage as a surrogate to brain injury.A model of blast TBI (bTBI) using a shock tube was compared to a lateral fluid percussion injury model (LFPI) using fluid pressure applied directly to the brain. Whole eyes were collected from mice 3 days post LFPI and 24 days post bTBI and were evaluated histologically using a hematoxylin and eosin stain.bTBI mice showed evidence of vitreous detachment in the posterior chamber in addition to vitreous hemorrhage with inflammatory cells. Subretinal hemorrhage, photoreceptor degeneration, and decreased cellularity in the retinal ganglion cell layer was also seen in bTBI mice. In contrast, eyes of LFPI mice showed evidence of anterior uveitis and subcapsular cataracts.We demonstrated that variations in the type of TBI can result in drastically different phenotypic changes within the eye. As such, molecular and phenotypic changes in the eye following TBI may provide valuable information regarding the mechanism, severity, and ongoing pathophysiology of brain injury. Because vitreous samples are easily obtained, molecular changes within the eye could be utilized as biomarkers of TBI in human patients.

View details for PubMedID 29560370
Proteomic analysis of the human retina reveals region-specific susceptibilities to metabolic-and oxidative stress-related diseases PLOS ONE Velez, G., Machlab, D. A., Tang, P. H., Sun, Y., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2018; 13 (2): e0193250
Abstract

Differences in regional protein expression within the human retina may explain molecular predisposition of specific regions to ophthalmic diseases like age-related macular degeneration, cystoid macular edema, retinitis pigmentosa, and diabetic retinopathy. To quantify protein levels in the human retina and identify patterns of differentially-expressed proteins, we collected foveomacular, juxta-macular, and peripheral retina punch biopsies from healthy donor eyes and analyzed protein content by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Protein expression was analyzed with 1-way ANOVA, gene ontology, pathway representation, and network analysis. We identified a mean of 1,974 proteins in the foveomacular retina, 1,999 in the juxta-macular retina, and 1,779 in the peripheral retina. Six hundred ninety-seven differentially-expressed proteins included those unique to and abundant in each anatomic region. Proteins with higher expression in each region include: heat-shock protein 90-alpha (HSP90AA1), and pyruvate kinase (PKM) in the foveomacular retina; vimentin (VIM) and fructose-bisphosphate aldolase C (ALDOC); and guanine nucleotide-binding protein subunit beta-1 (GNB1) and guanine nucleotide-binding protein subunit alpha-1 (GNAT1) in the peripheral retina. Pathway analysis identified downstream mediators of the integrin signaling pathway to be highly represented in the foveomacular region (P = 6.48 e-06). Metabolic pathways were differentially expressed among all retinal regions. Gene ontology analysis showed that proteins related to antioxidant activity were higher in the juxta-macular and the peripheral retina, but present in lower amounts in the foveomacular retina. Our proteomic analysis suggests that certain retinal regions are susceptible to different forms of metabolic and oxidative stress. The findings give mechanistic insight into retina function, reveal important molecular processes, and prioritize new pathways for therapeutic targeting.

View details for PubMedID 29466423
CRISPR Repair Reveals Causative Mutation in a Preclinical Model of Retinitis Pigmentosa: A Brief Methodology. Methods in molecular biology (Clifton, N.J.) Wu, W. H., Tsai, Y. T., Justus, S., Cho, G. Y., Sengillo, J. D., Xu, Y., Cabral, T., Lin, C. S., Bassuk, A. G., Mahajan, V. B., Tsang, S. H. 2018; 1715: 191–205
Abstract

CRISPR/Cas9 genome engineering is currently the leading genome surgery technology in most genetics laboratories. Combined with other complementary techniques, it serves as a powerful tool for uncovering genotype-phenotype correlations. Here, we describe a simplified protocol that was used in our publication, CRISPR Repair Reveals Causative Mutation in a Preclinical Model of Retinitis Pigmentosa, providing an overview of each section of the experimental process.

View details for PubMedID 29188514
Bevacizumab Injection in Patients with Neovascular Age-Related Macular Degeneration Increases Angiogenic Biomarkers. Ophthalmology. Retina Cabral, T., Lima, L. H., Mello, L. G., Polido, J., Correa, É. P., Oshima, A., Duong, J., Serracarbassa, P., Regatieri, C. V., Mahajan, V. B., Belfort, R. 2018; 2 (1): 31–37
Abstract

To evaluate the expression of 19 angiogenic biomarkers in the aqueous humor before and after intravitreal bevacizumab injection (IVB) in eyes with neovascular age-related macular degeneration (AMD).Prospective, noncomparative, interventional case series.Twenty-three eyes of 23 treatment-naïve patients with choroidal neovascularization (CNV) secondary to neovascular AMD.Eyes were diagnosed with CNV secondary to neovascular AMD and were treated with 3 monthly IVBs. Aqueous humor samples were obtained by anterior chamber paracentesis at baseline and immediately before each intravitreal bevacizumab injection.Aqueous humor levels of 19 angiogenic biomarkers (angiopoietin 2, bone morphogenetic protein 9 [BMP-9], epidermal growth factor [EGF], endoglin, endothelin 1, fibroblast growth factor [FGF]-1 and FGF-2, follistatin, granulocyte colony-stimulating factor [GCSF], heparin-binding EGF-like growth factor [HB-EGF], hepatocyte growth factor [HGF], interleukin 8, leptin, placental growth factor [PLGF], vascular endothelial growth factor [VEGF]-A, VEGF-C, VEGF-D, and tissue inhibitor of metalloproteinases [TIMP]-1 and TIMP-2) were measured. Best-corrected visual acuity (BCVA), spectral-domain OCT parameters, and intraocular pressure also were evaluated.Baseline aqueous VEGF-A expression was elevated in all study eyes before treatment initiation. A statistically significant decrease of VEGF-A was observed at the 1- and 2-month follow-ups. A statistically significant increased concentration was observed in 7 biomarkers: VEGF-C, angiopoietin 2, endothelin 1, follistatin, HB-EGF, HGF, and interleukin 8. The other 11 study biomarker levels (VEGF-D, BMP-9, EGF, endoglin, FGF-1, FGF-2, GCSF, leptin, PLGF, TIMP-1, and TIMP-2) did not show any significant difference during follow-up. The BCVA statistically improved significantly at 2 months. Spectral-domain OCT parameters improved significantly at all follow-ups. Mean intraocular pressure values were not statistically different during the study period.Despite a decrease in VEGF-A, the aqueous levels of VEGF-C, angiopoietin 2, endothelin 1, follistatin, HB-EGF, HGF, and interleukin 8 increased significantly after intravitreal injection of bevacizumab. These upregulated angiogenic biomarkers may represent new therapeutic targets in exudative AMD.

View details for PubMedID 29376143

View details for PubMedCentralID PMC5783314
ProSave: an application for restoring quantitative data to manipulated subsets of protein lists. Source code for biology and medicine Machlab, D. A., Velez, G., Bassuk, A. G., Mahajan, V. B. 2018; 13: 3
Abstract

In proteomics studies, liquid chromatography tandem mass spectrometry data (LC-MS/MS) is quantified by spectral counts or by some measure of ion abundance. Downstream comparative analysis of protein content (e.g. Venn diagrams and network analysis) typically does not include this quantitative data and critical information is often lost. To avoid loss of spectral count data in comparative proteomic analyses, it is critical to implement a tool that can rapidly retrieve this information.We developed ProSave, a free and user-friendly Java-based program that retrieves spectral count data from a curated list of proteins in a large proteomics dataset. ProSave allows for the management of LC-MS/MS datasets and rapidly retrieves spectral count information for a desired list of proteins.ProSave is open source and freely available at https://github.com/MahajanLab/ProSave. The user manual, implementation notes, and description of methodology and examples are available on the site.

View details for PubMedID 30459825

View details for PubMedCentralID PMC6233572
Fibrin Glue and Internal Limiting Membrane Abrasion for Optic Disc Pit Maculopathy. Ophthalmic surgery, lasers & imaging retina Almeida, D. R., Chin, E. K., Arjmand, P., Velez, G., Evans, L. P., Mahajan, V. B. 2018; 49 (12): e271–e277
Abstract

To describe a novel surgical technique using pars plana vitrectomy (PPV), internal limiting membrane (ILM) abrasion, and intravitreal fibrin glue for the treatment of optic disc pit maculopathy.Surgical case series technique with scanning electron microscopy (SEM) of human post-mortem eyes.Using SEM, the authors demonstrate the persistent adherence of vitreous fragments to the optic disc following induction of posterior vitreous detachment in human postmortem eyes. The authors describe a surgical technique using PPV, Tano Diamond Dusted Membrane Scraper for an ILM abrasion, intravitreal fibrin glue (Tisseel), and gas-air exchange to seal optic disc pits. The authors report successful long-term visual and anatomical outcomes in three patients.Intravitreal fibrin glue, when combined with ILM abrasion, may be a viable treatment option for optic disc pit maculopathy with good short- and long-term visual acuity outcomes. SEM shows that ILM abrasion removes vitreous fragments, which are persistently adherent and may lead to failure with other interventional techniques. [Ophthalmic Surg Lasers Imaging Retina. 2018;49:e271-e277.].

View details for DOI 10.3928/23258160-20181203-18

View details for PubMedID 30566713
Autologous stem cell therapy for inherited and acquired retinal disease REGENERATIVE MEDICINE Apatoff, M. L., Sengillo, J. D., White, E. C., Bakhoum, M. F., Bassuk, A. G., Mahajan, V. B., Tsang, S. H. 2018; 13 (1): 89–96
Abstract

The mammalian retina, derived from neural ectoderm, has little regenerative potential. For conditions where irreversible retinal pigment epithelium or photoreceptor cell loss occurs, advanced techniques are required to restore vision. Inherited retinal dystrophies and some acquired conditions, such as age-related macular degeneration, have a similar end result of photoreceptor cell death leading to debilitating vision loss. These diseases stand to benefit from future regenerative medicine as dietary recommendations and current pharmacologic therapy only seek to prevent further disease progression. Cell-based strategies, such as autologously derived induced pluripotent stem cells, have come a long way in overcoming previous technical and ethical concerns. Clinical trials for such techniques are already underway. These trials and the preceding preclinical studies will be discussed in the context of retinal disease.

View details for PubMedID 29360008
Extracellular superoxide dismutase 3 (SOD3) regulates oxidative stress at the vitreoretinal interface. Free radical biology & medicine Wert, K. J., Velez, G., Cross, M. R., Wagner, B. A., Teoh-Fitzgerald, M. L., Buettner, G. R., McAnany, J., Olivier, A., Tsang, S. H., Harper, M. M., Domann, F. E., Bassuk, A. G., Mahajan, V. B. 2018
Abstract

Oxidative stress is a pathogenic feature in vitreoretinal disease. However, the ability of the inner retina to manage metabolic waste and oxidative stress is unknown. Proteomic analysis of antioxidants in the human vitreous, the extracellular matrix opposing the inner retina, identified superoxide dismutase-3 (SOD3) that localized to a unique matrix structure in the vitreous base and cortex. To determine the role of SOD3, Sod3-/- mice underwent histological and clinical phenotyping. Although the eyes were structurally normal, at the vitreoretinal interface Sod3-/- mice demonstrated higher levels of 3-nitrotyrosine, a key marker of oxidative stress. Pattern electroretinography also showed physiological signaling abnormalities within the inner retina. Vitreous biopsies and epiretinal membranes collected from patients with diabetic vitreoretinopathy (DVR) and a mouse model of DVR showed significantly higher levels of nitrates and/or 3-nitrotyrosine oxidative stress biomarkers suggestive of SOD3 dysfunction. This study analyzes the molecular pathways that regulate oxidative stress in human vitreous substructures. The absence or dysregulation of the SOD3 antioxidant at the vitreous base and cortex results in increased oxidative stress and tissue damage to the inner retina, which may underlie DVR pathogenesis and other vitreoretinal diseases.

View details for PubMedID 29940351
Therapeutic drug repositioning using personalized proteomics of liquid biopsies. JCI insight Velez, G., Bassuk, A. G., Colgan, D., Tsang, S. H., Mahajan, V. B. 2017; 2 (24)
Abstract

BACKGROUND: In patients with limited response to conventional therapeutics, repositioning of already approved drugs can bring new, more effective options. Current drug repositioning methods, however, frequently rely on retrospective computational analyses and genetic testing - time consuming methods that delay application of repositioned drugs. Here, we show how proteomic analysis of liquid biopsies successfully guided treatment of neovascular inflammatory vitreoretinopathy (NIV), an inherited autoinflammatory disease with otherwise poor clinical outcomes.METHODS: Vitreous biopsies from NIV patients were profiled by an antibody array for expression of 200 cytokine-signaling proteins. Non-NIV controls were compared with NIV samples from various stages of disease progression. Patterns were identified by 1-way ANOVA, hierarchical clustering, and pathway analysis. Subjects treated with repositioned therapies were followed longitudinally.RESULTS: Proteomic profiles revealed molecular pathways in NIV pathologies and implicated superior and inferior targets for therapy. Anti-VEGF injections resolved vitreous hemorrhages without the need for vitrectomy surgery. Methotrexate injections reversed inflammatory cell reactions without the side effects of corticosteroids. Anti-IL-6 therapy prevented recurrent fibrosis and retinal detachment where all prior antiinflammatory interventions had failed. The cytokine array also showed that TNF-alpha levels were normal and that corticosteroid-sensitive pathways were absent in fibrotic NIV, helping explain prior failure of these conventional therapeutic approaches.CONCLUSIONS: Personalized proteomics can uncover highly personalized therapies for autoinflammatory disease that can be timed with specific pathologic activities. This precision medicine strategy can also help prevent delivery of ineffective drugs. Importantly, proteomic profiling of liquid biopsies offers an endpoint analysis that can directly guide treatment using available drugs.

View details for PubMedID 29263305
Therapeutic drug repositioning using personalized proteomics of liquid biopsies JCI INSIGHT Velez, G., Bassuk, A. G., Colgan, D., Tsang, S. H., Mahajan, V. B. 2017; 2 (24)
View details for DOI 10.1172/jci.insight.97818

View details for Web of Science ID 000418431100018
Genome Surgery and Gene Therapy in Retinal Disorders YALE JOURNAL OF BIOLOGY AND MEDICINE Chan, L., Mahajan, V. B., Tsang, S. H. 2017; 90 (4): 523–32
View details for Web of Science ID 000419234700002
Calpain-5 gene expression in the mouse eye and brain. BMC research notes Schaefer, K., Mahajan, M., Gore, A., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2017; 10 (1): 602
Abstract

OBJECTIVE: Our objective was to characterize CAPN5 gene expression in the mouse central nervous system. Mouse brain and eye sections were probed with two high-affinity RNA oligonucleotide analogs designed to bind CAPN5 RNA and one scramble, control oligonucleotide. Images were captured in brightfield.RESULTS: CAPN5 RNA probes were validated on mouse breast cancer tumor tissue. In the eye, CAPN5 was expressed in the ganglion cell, inner nuclear and outer nuclear layers of the retina. Signal could not be detected in the ciliary body or the iris because of the high density of melanin. In the brain, CAPN5 was expressed in the granule cell layers of the hippocampus and cerebellum. There was scattered expression in pons. The visual cortex showed faint signal. Most signal in the brain was in a punctate pattern.

View details for PubMedID 29157313
Electroretinography Reveals Difference in Cone Function between Syndromic and Nonsyndromic USH2A Patients SCIENTIFIC REPORTS Sengillo, J. D., Cabral, T., Schuerch, K., Duong, J., Lee, W., Boudreault, K., Xu, Y., Justus, S., Sparrow, J. R., Mahajan, V. B., Tsang, S. H. 2017; 7: 11170
Abstract

Usher syndrome is an inherited and irreversible disease that manifests as retinitis pigmentosa (RP) and bilateral neurosensory hearing loss. Mutations in Usherin 2A (USH2A) are not only a frequent cause of Usher syndrome, but also nonsyndromic RP. Although gene- and cell-based therapies are on the horizon for RP and Usher syndrome, studies characterizing natural disease are lacking. In this retrospective analysis, retinal function of USH2A patients was quantified with electroretinography. Both groups had markedly reduced rod and cone responses, but nonsyndromic USH2A patients had 30 Hz-flicker electroretinogram amplitudes that were significantly higher than syndromic patients, suggesting superior residual cone function. There was a tendency for Usher syndrome patients to have a higher distribution of severe mutations, and alleles in this group had a higher odds of containing nonsense or frame-shift mutations. These data suggest that the previously reported severe visual phenotype seen in syndromic USH2A patients could relate to a greater extent of cone dysfunction. Additionally, a genetic threshold may exist where mutation burden relates to visual phenotype and the presence of hearing deficits. The auditory phenotype and allelic hierarchy observed among patients should be considered in prospective studies of disease progression and during enrollment for future clinical trials.

View details for PubMedID 28894305
CRISPR-Cas Genome Surgery in Ophthalmology. Translational vision science & technology DiCarlo, J. E., Sengillo, J. D., Justus, S., Cabral, T., Tsang, S. H., Mahajan, V. B. 2017; 6 (3): 13-?
Abstract

Genetic disease affecting vision can significantly impact patient quality of life. Gene therapy seeks to slow the progression of these diseases by treating the underlying etiology at the level of the genome. Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated systems (Cas) represent powerful tools for studying diseases through the creation of model organisms generated by targeted modification and by the correction of disease mutations for therapeutic purposes. CRISPR-Cas systems have been applied successfully to the visual sciences and study of ophthalmic disease - from the modification of zebrafish and mammalian models of eye development and disease, to the correction of pathogenic mutations in patient-derived stem cells. Recent advances in CRISPR-Cas delivery and optimization boast improved functionality that continues to enhance genome-engineering applications in the eye. This review provides a synopsis of the recent implementations of CRISPR-Cas tools in the field of ophthalmology.

View details for DOI 10.1167/tvst.6.3.13

View details for PubMedID 28573077
ACANTHAMOEBA ENDOPHTHALMITIS AFTER RECURRENT KERATITIS AND NODULAR SCLERITIS. Retinal cases & brief reports Mammo, Z., Almeida, D. R., Cunningham, M. A., Chin, E. K., Mahajan, V. B. 2017; 11 (2): 180-182
Abstract

To describe the clinical course of a patient with Acanthamoeba keratitis, who despite prompt treatment progressed to histopathology-confirmed Acanthamoeba retinitis and endophthalmitis.Case report.A healthy 30-year-old male wearing soft contact lens was diagnosed with Acanthamoeba keratitis and treated medically and surgically over the course of 1 year with presumed resolution of the infection. Yet, his infection recurred with documented spread to sclerokeratitis, and overwhelming endophthalmitis. Concerns about extra-ocular spread prompted a therapeutic enucleation with histopathologic evidence of Acanthamoeba organisms throughout the globe.This is a case of a severe recurrent Acanthamoeba infection presenting initially as keratitis, followed by sclerokeratitis and histolopathology-confirmed endophthalmitis. This case demonstrates that despite persistent medical and surgical intervention, eradication of organisms may not be possible.

View details for DOI 10.1097/ICB.0000000000000323

View details for PubMedID 27152698
Recessive coding and regulatory mutations in FBLIM1 underlie the pathogenesis of chronic recurrent multifocal osteomyelitis (CRMO) PLOS ONE Cox, A. J., Darbro, B. W., Laxer, R. M., Velez, G., Bing, X., Finer, A. L., Erives, A., Mahajan, V. B., Bassuk, A. G., Ferguson, P. J. 2017; 12 (3)
Abstract

Chronic recurrent multifocal osteomyelitis (CRMO) is a rare, pediatric, autoinflammatory disease characterized by bone pain due to sterile osteomyelitis, and is often accompanied by psoriasis or inflammatory bowel disease. There are two syndromic forms of CRMO, Majeed syndrome and DIRA, for which the genetic cause is known. However, for the majority of cases of CRMO, the genetic basis is unknown. Via whole-exome sequencing, we detected a homozygous mutation in the filamin-binding domain of FBLIM1 in an affected child with consanguineous parents. Microarray analysis of bone marrow macrophages from the CRMO murine model (cmo) determined that the Fblim1 ortholog is the most differentially expressed gene, downregulated over 20-fold in the cmo mouse. We sequenced FBLIM1 in 96 CRMO subjects and found a second proband with a novel frameshift mutation in exon 6 and a rare regulatory variant. In SaOS2 cells, overexpressing the regulatory mutation showed the flanking region acts as an enhancer, and the mutation ablates enhancer activity. Our data implicate FBLIM1 in the pathogenesis of sterile bone inflammation and our findings suggest CRMO is a disorder of chronic inflammation and imbalanced bone remodeling.

View details for DOI 10.1371/journal.pone.0169687

View details for Web of Science ID 000396318300002

View details for PubMedID 28301468

View details for PubMedCentralID PMC5354242
Efficacy and safety of voretigene neparvovec (AAV2-hRPE65v2) in patients with RPE65-mediated inherited retinal dystrophy: a randomised, controlled, open-label, phase 3 trial. Lancet (London, England) Russell, S., Bennett, J., Wellman, J. A., Chung, D. C., Yu, Z. F., Tillman, A., Wittes, J., Pappas, J., Elci, O., McCague, S., Cross, D., Marshall, K. A., Walshire, J., Kehoe, T. L., Reichert, H., Davis, M., Raffini, L., George, L. A., Hudson, F. P., Dingfield, L., Zhu, X., Haller, J. A., Sohn, E. H., Mahajan, V. B., Pfeifer, W., Weckmann, M., Johnson, C., Gewaily, D., Drack, A., Stone, E., Wachtel, K., Simonelli, F., Leroy, B. P., Wright, J. F., High, K. A., Maguire, A. M. 2017
Abstract

Phase 1 studies have shown potential benefit of gene replacement in RPE65-mediated inherited retinal dystrophy. This phase 3 study assessed the efficacy and safety of voretigene neparvovec in participants whose inherited retinal dystrophy would otherwise progress to complete blindness.In this open-label, randomised, controlled phase 3 trial done at two sites in the USA, individuals aged 3 years or older with, in each eye, best corrected visual acuity of 20/60 or worse, or visual field less than 20 degrees in any meridian, or both, with confirmed genetic diagnosis of biallelic RPE65 mutations, sufficient viable retina, and ability to perform standardised multi-luminance mobility testing (MLMT) within the luminance range evaluated, were eligible. Participants were randomly assigned (2:1) to intervention or control using a permuted block design, stratified by age (<10 years and ≥10 years) and baseline mobility testing passing level (pass at ≥125 lux vs <125 lux). Graders assessing primary outcome were masked to treatment group. Intervention was bilateral, subretinal injection of 1·5 × 10(11) vector genomes of voretigene neparvovec in 0·3 mL total volume. The primary efficacy endpoint was 1-year change in MLMT performance, measuring functional vision at specified light levels. The intention-to-treat (ITT) and modified ITT populations were included in primary and safety analyses. This trial is registered with ClinicalTrials.gov, number NCT00999609, and enrolment is complete.Between Nov 15, 2012, and Nov 21, 2013, 31 individuals were enrolled and randomly assigned to intervention (n=21) or control (n=10). One participant from each group withdrew after consent, before intervention, leaving an mITT population of 20 intervention and nine control participants. At 1 year, mean bilateral MLMT change score was 1·8 (SD 1·1) light levels in the intervention group versus 0·2 (1·0) in the control group (difference of 1·6, 95% CI 0·72-2·41, p=0·0013). 13 (65%) of 20 intervention participants, but no control participants, passed MLMT at the lowest luminance level tested (1 lux), demonstrating maximum possible improvement. No product-related serious adverse events or deleterious immune responses occurred. Two intervention participants, one with a pre-existing complex seizure disorder and another who experienced oral surgery complications, had serious adverse events unrelated to study participation. Most ocular events were mild in severity.Voretigene neparvovec gene replacement improved functional vision in RPE65-mediated inherited retinal dystrophy previously medically untreatable.Spark Therapeutics.

View details for DOI 10.1016/S0140-6736(17)31868-8

View details for PubMedID 28712537
Retrospective Analysis of Structural Disease Progression in Retinitis Pigmentosa Utilizing Multimodal Imaging. Scientific reports Cabral, T., Sengillo, J. D., Duong, J. K., Justus, S., Boudreault, K., Schuerch, K., Belfort, R., Mahajan, V. B., Sparrow, J. R., Tsang, S. H. 2017; 7 (1): 10347
Abstract

In this report, we assess the natural progression rate of retinitis pigmentosa (RP) over an average of three years using spectral-domain optical coherence tomography (SD-OCT) and short wavelength fundus autofluorescence (SW-AF). Measurement of the ellipsoid zone (EZ) line width and hyperautofluorescent ring diameters was performed in 81 patients with RP in a retrospective, longitudinal fashion. Rate of structural disease progression, symmetry between eyes, and test-retest variability were quantified. We observed on average, EZ-line widths decreased by 140 µm (5.2%, p < 0.001) per year, and average horizontal and vertical hyperautofluorescent ring diameters decreased by 149 µm (3.6%, p < 0.001) and 120 µm (3.9%, p < 0.001) per year, respectively. The 95th percentile of this cohort had differences in progression slopes between eyes that were less than 154 µm, 118 µm, and 132 µm for EZ-line width and horizontal and vertical ring diameters, respectively. For all measures except horizontal ring diameter, progression rates were significantly slower at end-stage disease. From our data, we observed a statistically significant progression rate in EZ line width and SW-AF ring diameters over time, verifying the utility of these measurements for disease monitoring purposes. Additionally, calculated differences in progression slopes between eyes may prove useful for investigators evaluating the efficacy of unilateral treatments for RP in clinical trials.

View details for PubMedID 28871101

View details for PubMedCentralID PMC5583352
Genome Surgery and Gene Therapy in Retinal Disorders. The Yale journal of biology and medicine Chan, L., Mahajan, V. B., Tsang, S. H. 2017; 90 (4): 523–32
Abstract

The emergence of genome surgery techniques like the clustered regularly interspaced short palindromic repeats (CRISPR) editing technology has given researchers a powerful tool for precisely introducing targeted changes within the genome. New modifications to the CRISPR-Cas system have been made since its recent discovery, such as high-fidelity Cas9 variants to reduce off-target effects and transcriptional activation/silencing with CRISPRa/CRISPRi. The applications of CRISPR-Cas and gene therapy in ophthalmic diseases have been necessary and fruitful, especially given the impact of blinding diseases on society and the large number of monogenic disorders of the eye. This review discusses the impact that CRISPR-Cas has had on furthering our understanding of disease mechanisms and potential therapies for inherited eye diseases. Furthermore, we explore a brief overview of recent and ongoing gene therapy clinical trials in retinal diseases, and conclude with the implications of genome surgery on the outlook of future therapeutic interventions.

View details for PubMedID 29259518
Personalized Proteomics in Proliferative Vitreoretinopathy Implicate Hematopoietic Cell Recruitment and mTOR as a Therapeutic Target American Journal of Ophthalmology Roybal, C. N., Velez, G., Toral, M. A., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2017: 30521-4
Abstract

To profile vitreous cytokine expression of proliferative vitreoretinopathy (PVR) patients DESIGN: Case-control study.Liquid biopsies were collected from two groups: control subjects (n=3) undergoing pars plana vitrectomy to remove an epiretinal membrane (ERM), and test subjects (n=7) with varying degrees of PVR. A high-throughput cytokine screen measured expression of 200 cytokines. Cytokine expression patterns were prospectively validated in separate cohorts of control patients and those with PVR-A, PVR-B, and PVR-C (n=10 for each group). Expression changes were evaluated by ANOVA (significant p-value <0.05), hierarchical cluster algorithm, and pathway analysis, to identify candidate pathways for prospective studies.In PVR vitreous, 29 cytokines were upregulated compared to controls. Early-PVR vitreous showed upregulation of T-cell markers, pro-fibrotic cytokines, and cytokines downstream of mTOR activation (IL-2, IL-6, and IL-13), whereas late PVR vitreous, cytokines driving monocyte responses and stem-cell recruitment (SDF-1) prevailed. Prospective validation confirmed the differential-expression of specific cytokines from PVR-A to C.Early PVR is characterized by activation of T-cells and mTOR signaling, whereas advanced-PVR is characterized by a chronic monocyte response. PVR might be treated by rational repositioning of existing drugs that target mTOR and IL-6. Our analysis demonstrates that successful therapeutic intervention will be highly dependent on the specific therapeutic target and the stage of PVR. This study provides insights into cytokines that will serve as biomarkers and therapeutic targets. These biomarkers will help design clinical trials that intervene at appropriate times.

View details for DOI 10.1016/j.ajo.2017.11.025
Retinal and choroidal angiogenesis: a review of new targets. International journal of retina and vitreous Cabral, T., Mello, L. G., Lima, L. H., Polido, J., Regatieri, C. V., Belfort, R., Mahajan, V. B. 2017; 3: 31
Abstract

Retinal and choroidal neovascularization are a major cause of significant visual impairment, worldwide. Understanding the various factors involved in the accompanying physiopathology is vital for development of novel treatments, and most important, for preserving patient vision. The intraocular use of anti-vascular endothelial growth factor therapeutics has improved management of the retinal and choroidal neovascularization but some patients do not respond, suggesting other vascular mediators may also contribute to ocular angiogenesis. Several recent studies examined possible new targets for future anti-angiogenic therapies. Potential targets of retinal and choroidal neovascularization therapy include members of the platelet-derived growth factor family, vascular endothelial growth factor sub-family, epidermal growth factor family, fibroblast growth factor family, transforming growth factor-β superfamily (TGF-β1, activins, follistatin and bone morphogenetic proteins), angiopoietin-like family, galectins family, integrin superfamily, as well as pigment epithelium derived factor, hepatocyte growth factor, angiopoietins, endothelins, hypoxia-inducible factors, insulin-like growth factors, cytokines, matrix metalloproteinases and their inhibitors and glycosylation proteins. This review highlights current antiangiogenic therapies under development, and discusses future retinal and choroidal pro- and anti-angiogenic targets as wells as the importance of developing of new drugs.

View details for PubMedID 28835854

View details for PubMedCentralID PMC5563895
Dissection of Human Retina and RPE-Choroid for Proteomic Analysis. Journal of visualized experiments : JoVE Cabral, T., Toral, M. A., Velez, G., DiCarlo, J. E., Gore, A. M., Mahajan, M., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2017
Abstract

The human retina is composed of the sensory neuroretina and the underlying retinal pigmented epithelium (RPE), which is firmly complexed to the vascular choroid layer. Different regions of the retina are anatomically and molecularly distinct, facilitating unique functions and demonstrating differential susceptibility to disease. Proteomic analysis of each of these regions and layers can provide vital insights into the molecular process of many diseases, including Age-Related Macular Degeneration (AMD), diabetes mellitus, and glaucoma. However, separation of retinal regions and layers is essential before quantitative proteomic analysis can be accomplished. Here, we describe a method for dissection and collection of the foveal, macular, and peripheral retinal regions and underlying RPE-choroid complex, involving regional punch biopsies and manual removal of tissue layers from a human eye.One-dimensional SDS-PAGE as well as downstream proteomic analysis, such as liquid chromatography-tandem mass spectrometry (LC-MS/MS), can be used to identify proteins in each dissected retinal layer, revealing molecular biomarkers for retinal disease.

View details for PubMedID 29155757
CRISPR-mediated Ophthalmic Genome Surgery. Current ophthalmology reports Cho, G. Y., Abdulla, Y., Sengillo, J. D., Justus, S., Schaefer, K. A., Bassuk, A. G., Tsang, S. H., Mahajan, V. B. 2017; 5 (3): 199–206
Abstract

Clustered regularly interspaced short palindromic repeats (CRISPR) is a genome engineering system with great potential for clinical applications due to its versatility and programmability. This review highlights the development and use of CRISPR-mediated ophthalmic genome surgery in recent years.Diverse CRISPR techniques are in development to target a wide array of ophthalmic conditions, including inherited and acquired conditions. Preclinical disease modeling and recent successes in gene editing suggest potential efficacy of CRISPR as a therapeutic for inherited conditions. In particular, the treatment of Leber congenital amaurosis with CRISPR-mediated genome surgery is expected to reach clinical trials in the near future.Treatment options for inherited retinal dystrophies are currently limited. CRISPR-mediated genome surgery methods may be able to address this unmet need in the future.

View details for PubMedID 28966884

View details for PubMedCentralID PMC5613978
Gene Therapy Restores Mfrp and Corrects Axial Eye Length. Scientific reports Velez, G., Tsang, S. H., Tsai, Y. T., Hsu, C. W., Gore, A., Abdelhakim, A. H., Mahajan, M., Silverman, R. H., Sparrow, J. R., Bassuk, A. G., Mahajan, V. B. 2017; 7 (1): 16151
Abstract

Hyperopia (farsightedness) is a common and significant cause of visual impairment, and extreme hyperopia (nanophthalmos) is a consequence of loss-of-function MFRP mutations. MFRP deficiency causes abnormal eye growth along the visual axis and significant visual comorbidities, such as angle closure glaucoma, cystic macular edema, and exudative retinal detachment. The Mfrp rd6 /Mfrp rd6 mouse is used as a pre-clinical animal model of retinal degeneration, and we found it was also hyperopic. To test the effect of restoring Mfrp expression, we delivered a wild-type Mfrp to the retinal pigmented epithelium (RPE) of Mfrp rd6 /Mfrp rd6 mice via adeno-associated viral (AAV) gene therapy. Phenotypic rescue was evaluated using non-invasive, human clinical testing, including fundus auto-fluorescence, optical coherence tomography, electroretinography, and ultrasound. These analyses showed gene therapy restored retinal function and normalized axial length. Proteomic analysis of RPE tissue revealed rescue of specific proteins associated with eye growth and normal retinal and RPE function. The favorable response to gene therapy in Mfrp rd6 /Mfrp rd6 mice suggests hyperopia and associated refractive errors may be amenable to AAV gene therapy.

View details for PubMedID 29170418

View details for PubMedCentralID PMC5701072
Personalized proteomics in proliferative vitreoretinopathy implicate hematopoietic cell recruitment and mTOR as a therapeutic target. American journal of ophthalmology Roybal, C. N., Velez, G., Toral, M., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2017
Abstract

To profile vitreous cytokine expression of proliferative vitreoretinopathy (PVR) patients DESIGN: Case-control study.Liquid biopsies were collected from two groups: control subjects (n=3) undergoing pars plana vitrectomy to remove an epiretinal membrane (ERM), and test subjects (n=7) with varying degrees of PVR. A high-throughput cytokine screen measured expression of 200 cytokines. Cytokine expression patterns were prospectively validated in separate cohorts of control patients and those with PVR-A, PVR-B, and PVR-C (n=10 for each group). Expression changes were evaluated by ANOVA (significant p-value <0.05), hierarchical cluster algorithm, and pathway analysis, to identify candidate pathways for prospective studies.In PVR vitreous, 29 cytokines were upregulated compared to controls. Early-PVR vitreous showed upregulation of T-cell markers, pro-fibrotic cytokines, and cytokines downstream of mTOR activation (IL-2, IL-6, and IL-13), whereas late PVR vitreous, cytokines driving monocyte responses and stem-cell recruitment (SDF-1) prevailed. Prospective validation confirmed the differential-expression of specific cytokines from PVR-A to C.Early PVR is characterized by activation of T-cells and mTOR signaling, whereas advanced-PVR is characterized by a chronic monocyte response. PVR might be treated by rational repositioning of existing drugs that target mTOR and IL-6. Our analysis demonstrates that successful therapeutic intervention will be highly dependent on the specific therapeutic target and the stage of PVR. This study provides insights into cytokines that will serve as biomarkers and therapeutic targets. These biomarkers will help design clinical trials that intervene at appropriate times.

View details for PubMedID 29246578
Limbal Trocar-Cannulas for Complex Vitrectomy Surgery. Retina (Philadelphia, Pa.) Mears, K. A., Mahajan, V. B. 2017
View details for PubMedID 28737532
Small-angle X-ray scattering of calpain-5 reveals a highly open conformation among calpains JOURNAL OF STRUCTURAL BIOLOGY Gakhar, L., Bassuk, A. G., Velez, G., Khan, S., Yang, J., Tsang, S. H., Mahajan, V. B. 2016; 196 (3): 309-318
Abstract

Calpain-5 is a calcium-activated protease expressed in the retina. Mutations in calpain-5 cause autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV, OMIM#193235). The structure of calpain-5 has not been determined, thus hindering the investigation of its proteolytic targets and pathological role in ADNIV. Herein, we report models of the proteolytic core of calpain-5 (mini-calpain-5) containing two globular domains (termed DIIa-IIb) connected by a short, flexible linker, consistent with small-angle X-ray scattering (SAXS) data. Structural modeling in the absence of calcium suggests that mini-calpain-5 adopts a more open conformation when compared to previously determined structures of other calpain cores. This open conformation, achieved by a rotation of DIIa and DIIb with respect to each other, prevents formation of the active site and constrains the enzyme in an inactivated form. The relative domain rotation of 60-100° we found for mini-calpain-5 (a non-classical calpain) is significantly greater than the largest rotation previously observed for a classical calpain (i.e., 55.0° for mini-calpain-9). Together with our prediction that, in the full-length form, a long loop in DIIb (loop C1), a few residues downstream of the inter-domain linker, likely interacts with the shorter, acidic, inactivating loop on domain-III (DIII), these structural insights illuminate the complexity of calpain regulation. Moreover, our studies argue that pursuing higher resolution structural studies are necessary to understand the complex activity regulation prevalent in the calpain family and for the design of specific calpain inhibitors.

View details for DOI 10.1016/j.jsb.2016.07.017

View details for Web of Science ID 000389295000003

View details for PubMedID 27474374

View details for PubMedCentralID PMC5118095
Reprogramming metabolism by targeting sirtuin 6 attenuates retinal degeneration JOURNAL OF CLINICAL INVESTIGATION Zhang, L., Du, J., Justus, S., Hsu, C., Bonet-Ponce, L., Wu, W., Tsai, Y., Wu, W., Jia, Y., Duong, J. K., Mahajan, V. B., Lin, C., Wang, S., Hurley, J. B., Tsang, S. H. 2016; 126 (12): 4659-4673
Abstract

Retinitis pigmentosa (RP) encompasses a diverse group of Mendelian disorders leading to progressive degeneration of rods and then cones. For reasons that remain unclear, diseased RP photoreceptors begin to deteriorate, eventually leading to cell death and, consequently, loss of vision. Here, we have hypothesized that RP associated with mutations in phosphodiesterase-6 (PDE6) provokes a metabolic aberration in rod cells that promotes the pathological consequences of elevated cGMP and Ca2+, which are induced by the Pde6 mutation. Inhibition of sirtuin 6 (SIRT6), a histone deacetylase repressor of glycolytic flux, reprogrammed rods into perpetual glycolysis, thereby driving the accumulation of biosynthetic intermediates, improving outer segment (OS) length, enhancing photoreceptor survival, and preserving vision. In mouse retinae lacking Sirt6, effectors of glycolytic flux were dramatically increased, leading to upregulation of key intermediates in glycolysis, TCA cycle, and glutaminolysis. Both transgenic and AAV2/8 gene therapy-mediated ablation of Sirt6 in rods provided electrophysiological and anatomic rescue of both rod and cone photoreceptors in a preclinical model of RP. Due to the extensive network of downstream effectors of Sirt6, this study motivates further research into the role that these pathways play in retinal degeneration. Because reprogramming metabolism by enhancing glycolysis is not gene specific, this strategy may be applicable to a wide range of neurodegenerative disorders.

View details for DOI 10.1172/JCI86905

View details for Web of Science ID 000390131900025

View details for PubMedID 27841758

View details for PubMedCentralID PMC5127684
Management of Pediatric Aphakic Glaucoma With Vitrectomy and Tube Shunts JOURNAL OF PEDIATRIC OPHTHALMOLOGY & STRABISMUS Elshatory, Y. M., Gauger, E. H., Kwon, Y. H., Alward, W. L., Boldt, H. C., Russell, S. R., Mahajan, V. B. 2016; 53 (6): 339-343
Abstract

To review the impact of vitrectomy and tube shunts on mean intraocular pressure (IOP) and number of glaucoma medications in pediatric aphakic glaucoma.A retrospective review of pediatric patients who underwent combined vitrectomy and glaucoma tube shunt surgery for aphakic glaucoma was conducted. Inclusion criteria were: age 18 years or younger, diagnosis of aphakic glaucoma, preoperative IOP data, and postoperative IOP data for at least 6 months. Mean IOP lowering at 1 year, number of glaucoma medications at 1 year, and surgical complications, including tube occlusion in the postoperative period, were noted.The mean ± standard deviation preoperative IOP was 33.9 ± 10.6 mm Hg (range: 18 to 57 mm Hg) with a mean of three topical IOP-lowering medications. A total of 5 (36%) Ahmed and 9 (64%) Baerveldt tube shunts were placed. One of the Baerveldt tube shunt procedures was combined with revision of a traumatically dislocated tube. The mean IOP at 12 months postoperatively was 16.6 ± 5.8 mm Hg (range: 6 to 28 mm Hg; P < .01, t = 3.74, df = 13) with a mean of 2.3 glaucoma medications. There were no cases of tube occlusion, corneal decompensation, endophthalmitis, or retinal detachment over the 12 months of follow-up.Combined vitrectomy and placement of a glaucoma tube shunt can be safe and effective in lowering IOP based on mean IOP values and number of glaucoma medications at 1 year. [J Pediatr Ophthalmol Strabismus. 2016;53(6):339-343.].

View details for DOI 10.3928/01913913-20160818-01

View details for Web of Science ID 000393050000005

View details for PubMedID 27668871
COMBINED VITRECTOMY AND INTRAVITREAL DEXAMETHASONE (OZURDEX) SUSTAINED-RELEASE IMPLANT RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Zheng, A., Chin, E. K., Almeida, D. R., Tsang, S. H., Mahajan, V. B. 2016; 36 (11): 2087-2092
Abstract

To evaluate the safety and efficacy of combining intravitreal dexamethasone implantation (Ozurdex) with pars plana vitrectomy (PPV).A retrospective review was conducted on cases where Ozurdex injection was performed in the operating room in conjunction with pars plana vitrectomy. Our primary outcome measure was the presence of surgical complications in the perioperative and 3-month postoperative window. We also measured visual acuity, intraocular pressure (IOP), and macular edema at baseline, one, and 3 months after surgery.Fifteen eyes in 14 cases were reviewed. There were no complications intraoperatively or at 1-month postoperatively. Two patients (2 eyes) with prior retinal detachment developed proliferative vitreoretinopathy and redetachment at 3 months. Visual acuity improved in 7 of 15 eyes, and an average improvement of 2 lines was achieved for the entire cohort. There was no overall change in intraocular pressure although 1 patient developed an increase in intraocular pressure >5 mmHg. Five of 9 patients with baseline macular edema experienced improvement or resolution at 3 months.Intraoperative Ozurdex in combination with PPV may be safe and effective in treating macular edema caused by many different underlying diseases.

View details for Web of Science ID 000387079800014

View details for PubMedID 27148836

View details for PubMedCentralID PMC5077653
Catenin delta-1 (CTNND1) phosphorylation controls the mesenchymal to epithelial transition in astrocytic tumors HUMAN MOLECULAR GENETICS Yang, J., Bassuk, A. G., Merl-Pham, J., Hsu, C., Colgan, D. F., Li, X., Au, K. S., Zhang, L., Smemo, S., Justus, S., Nagahama, Y., Grossbach, A. J., Howard, M. A., Kawasaki, H., Feldstein, N. A., Dobyns, W. B., Northrup, H., Hauck, S. M., Ueffing, M., Mahajan, V. B., Tsang, S. H. 2016; 25 (19): 4201-4210
Abstract

Inactivating mutations of the TSC1/TSC2 complex (TSC1/2) cause tuberous sclerosis (TSC), a hereditary syndrome with neurological symptoms and benign hamartoma tumours in the brain. Since TSC effectors are largely unknown in the human brain, TSC patient cortical tubers were used to uncover hyperphosphorylation unique to TSC primary astrocytes, the cell type affected in the brain. We found abnormal hyperphosphorylation of catenin delta-1 S268, which was reversible by mTOR-specific inhibitors. In contrast, in three metastatic astrocytoma cell lines, S268 was under phosphorylated, suggesting S268 phosphorylation controls metastasis. TSC astrocytes appeared epithelial (i.e. tightly adherent, less motile, and epithelial (E)-cadherin positive), whereas wild-type astrocytes were mesenchymal (i.e. E-cadherin negative and highly motile). Despite their epithelial phenotype, TSC astrocytes outgrew contact inhibition, and monolayers sporadically generated tuberous foci, a phenotype blocked by the mTOR inhibitor, Torin1. Also, mTOR-regulated phosphokinase C epsilon (PKCe) activity induced phosphorylation of catenin delta-1 S268, which in turn mediated cell-cell adhesion in astrocytes. The mTOR-dependent, epithelial phenotype of TSC astrocytes suggests TSC1/2 and mTOR tune the phosphorylation level of catenin delta-1 by controlling PKCe activity, thereby regulating the mesenchymal-epithelial-transition (MET). Thus, some forms of TSC could be treated with PKCe inhibitors, while metastasis of astrocytomas might be blocked by PKCe stimulators.

View details for DOI 10.1093/hmg/ddw253

View details for PubMedID 27516388
ELEVATED INTRAOCULAR PRESSURE FOLLOWING PARS PLANA VITRECTOMY DUE TO TRAPPED GAS IN THE POSTERIOR CHAMBER. Retinal cases & brief reports Chin, E. K., Almeida, D. R., Strohbehn, A. L., Mahajan, V. B., Russell, S. R., Folk, J. C. 2016; 10 (4): 334-337
Abstract

Elevated intraocular pressure is relatively common following pars plana vitrectomy and intraocular gas tamponade. We discuss a series of patients who experienced elevated intraocular pressure from pupillary block and angle closure secondary to trapped gas in the posterior chamber.Case series.Case 1 is a patient who underwent pars plana vitrectomy for retinal detachment repair. The intraocular pressure was elevated on postoperative Day 3 because of trapped gas in the posterior chamber, and it did not lower with prone positioning, maximal medical therapy, and laser peripheral iridotomies. Aspiration of the trapped gas was done with the patient sitting upright using a 27-gauge needle at the limbus, which was curative. Case 2 provides anterior-segment optical coherence tomography images that confirmed the location of the trapped gas resulting in angle closure. Case 3 demonstrates the unfortunate sequelae of a central retinal artery occlusion following delayed recognition of this entity. Case 4 highlights the challenges encountered when migratory gas is also seen elsewhere in the eye.Clinicians should be aware of elevated intraocular pressure secondary to trapped gas in the posterior chamber, which may be recalcitrant to medical therapy. Aspiration of the trapped gas can alleviate both pupillary block and angle closure without compromising the gas tamponade.

View details for DOI 10.1097/ICB.0000000000000256

View details for PubMedID 26630244
SURGICAL EMBOLECTOMY FOR FOVEA-THREATENING ACUTE RETINAL ARTERY OCCLUSION. Retinal cases & brief reports Almeida, D. R., Mammo, Z., Chin, E. K., Mahajan, V. B. 2016; 10 (4): 331-333
Abstract

To describe a technique of surgical intraocular embolectomy in patients with acute fovea-threatening branch retinal artery occlusion.Pars plana vitrectomy with embolectomy involving embolus isolation, dissection, and removal in patients with an acute fovea-threatening arterial occlusion without a patent cilioretinal artery.The surgical technique involves a core vitrectomy. The blocked artery is incised using a microvitreoretinal blade, and microsurgical forceps are used to retrieve the embolus. No significant complications were noted. The study technique offers an excellent safety profile and minimizes the risk of vitreous hemorrhage by carefully dissecting the vascular adventitial sheath and isolating the embolus.Surgical embolectomy is a viable technique for patients with acute fovea-threatening arterial occlusions without patent cilioretinal artery. Careful dissection and retrieval of the embolus minimizes the risk of vitreous hemorrhage, which is an important improvement in previous techniques for management of acute retinal arterial occlusions.

View details for DOI 10.1097/ICB.0000000000000257

View details for PubMedID 26674278

View details for PubMedCentralID PMC5051520
Reprogramming towards anabolism impedes degeneration in a preclinical model of retinitis pigmentosa HUMAN MOLECULAR GENETICS Zhang, L., Justus, S., Xu, Y., Pluchenik, T., Hsu, C., Yang, J., Duong, J. K., Lin, C., Jia, Y., Bassuk, A. G., Mahajan, V. B., Tsang, S. H. 2016; 25 (19): 4244-4255
Abstract

Retinitis pigmentosa (RP) is an incurable neurodegenerative condition featuring photoreceptor death that leads to blindness. Currently, there is no approved therapeutic for photoreceptor degenerative conditions like RP and atrophic age-related macular degeneration (AMD). Although there are promising results in human gene therapy, RP is a genetically diverse disorder, such that gene-specific therapies would be practical in a small fraction of patients with RP. Here, we explore a non-gene-specific strategy that entails reprogramming photoreceptors towards anabolism by upregulating the mechanistic target of rapamycin (mTOR) pathway. We conditionally ablated the tuberous sclerosis complex 1 (Tsc1) gene, an mTOR inhibitor, in the rods of the Pde6bH620Q/H620Qpreclinical RP mouse model and observed, functionally and morphologically, an improvement in the survival of rods and cones at early and late disease stages. These results elucidate the ability of reprogramming the metabolome to slow photoreceptor degeneration. This strategy may also be applicable to a wider range of neurodegenerative diseases, as enhancement of nutrient uptake is not gene-specific and is implicated in multiple pathologies. Enhancing anabolism promoted neuronal survival and function and could potentially benefit a number of photoreceptor and other degenerative conditions.

View details for DOI 10.1093/hmg/ddw256

View details for Web of Science ID 000395807800008

View details for PubMedID 27516389

View details for PubMedCentralID PMC5291198
CRISPR Repair Reveals Causative Mutation in a Preclinical Model of Retinitis Pigmentosa MOLECULAR THERAPY Wu, W., Tsai, Y., Justus, S., Lee, T., Zhang, L., Lin, C., Bassuk, A. G., Mahajan, V. B., Tsang, S. H. 2016; 24 (8): 1388-1394
Abstract

Massive parallel sequencing enables identification of numerous genetic variants in mutant organisms, but determining pathogenicity of any one mutation can be daunting. The most commonly studied preclinical model of retinitis pigmentosa called the "rodless" (rd1) mouse is homozygous for two mutations: a nonsense point mutation (Y347X) and an intronic insertion of a leukemia virus (Xmv-28). Distinguishing which mutation causes retinal degeneration is still under debate nearly a century after the discovery of this model organism. Here, we performed gene editing using the CRISPR/Cas9 system and demonstrated that the Y347X mutation is the causative variant of disease. Genome editing in the first generation produced animals that were mosaic for the corrected allele but still showed neurofunction preservation despite low repair frequencies. Furthermore, second-generation CRISPR-repaired mice showed an even more robust rescue and amelioration of the disease. This predicts excellent outcomes for gene editing in diseased human tissue, as Pde6b, the mutated gene in rd1 mice, has an orthologous intron-exon relationship comparable with the human PDE6B gene. Not only do these findings resolve the debate surrounding the source of neurodegeneration in the rd1 model, but they also provide the first example of homology-directed recombination-mediated gene correction in the visual system.

View details for DOI 10.1038/mt.2016.107

View details for Web of Science ID 000383280900010

View details for PubMedID 27203441

View details for PubMedCentralID PMC5023380
BESTROPHIN1 mutations cause defective chloride conductance in patient stem cell-derived RPE HUMAN MOLECULAR GENETICS Moshfegh, Y., Velez, G., Li, Y., Bassuk, A. G., Mahajan, V. B., Tsang, S. H. 2016; 25 (13): 2672-2680
Abstract

Bestrophin1 (BEST1) is expressed in human retinal pigment epithelium (RPE) and mutations in the BEST1 gene commonly cause retinal dysfunction and macular degeneration. BEST1 is presumed to assemble into a calcium-activated chloride channel and be involved in chloride transport but there is no direct evidence in live human RPE cells to support this idea. To test whether BEST1 functions as a chloride channel in living tissue, BEST1-mutant RPE (R218H, L234P, A243T) were generated from patient-derived induced pluripotent stem cells and compared with wild-type RPE in a retinal environment, using a biosensor that visualizes calcium-induced chloride ion flux in the cell. Calcium stimulation elicited chloride ion export in normal RPE but not in RPE derived from three patients with BEST1 mutations. These data, along with three-dimensional modeling, provide evidence that BEST1 assembles into a key calcium-sensing chloride channel in human RPE.

View details for DOI 10.1093/hmg/ddw126

View details for Web of Science ID 000393064400005

View details for PubMedID 27193166

View details for PubMedCentralID PMC5181636
Complication of Autologous Stem Cell Transplantation in Retinitis Pigmentosa JAMA OPHTHALMOLOGY Boudreault, K., Justus, S., Lee, W., Mahajan, V. B., Tsang, S. H. 2016; 134 (6): 711-712
View details for DOI 10.1001/jamaophthalmol.2016.0803

View details for Web of Science ID 000379596300021

View details for PubMedID 27149677
Calpain-5 Expression in the Retina Localizes to Photoreceptor Synapses INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE Schaefer, K. A., Toral, M. A., Velez, G., Cox, A. J., Baker, S. A., Borcherding, N. C., Colgan, D. F., Bondada, V., Mashburn, C. B., Yu, C., Geddes, J. W., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2016; 57 (6): 2509-2521
Abstract

We characterize calpain-5 (CAPN5) expression in retinal and neuronal subcellular compartments.CAPN5 gene variants were classified using the exome variant server, and RNA-sequencing was used to compare expression of CAPN5 mRNA in the mouse and human retina and in retinoblastoma cells. Expression of CAPN5 protein was ascertained in humans and mice in silico, in mouse retina by immunohistochemistry, and in neuronal cancer cell lines and fractionated central nervous system tissue extracts by Western analysis with eight antibodies targeting different CAPN5 regions.Most CAPN5 genetic variation occurs outside its protease core; and searches of cancer and epilepsy/autism genetic databases found no variants similar to hyperactivating retinal disease alleles. The mouse retina expressed one transcript for CAPN5 plus those of nine other calpains, similar to the human retina. In Y79 retinoblastoma cells, the level of CAPN5 transcript was very low. Immunohistochemistry detected CAPN5 expression in the inner and outer nuclear layers and at synapses in the outer plexiform layer. Western analysis of fractionated retinal extracts confirmed CAPN5 synapse localization. Western blots of fractionated brain neuronal extracts revealed distinct subcellular patterns and the potential presence of autoproteolytic CAPN5 domains.CAPN5 is moderately expressed in the retina and, despite higher expression in other tissues, hyperactive disease mutants of CAPN5 only manifest as eye disease. At the cellular level, CAPN5 is expressed in several different functional compartments. CAPN5 localization at the photoreceptor synapse and with mitochondria explains the neural circuitry phenotype in human CAPN5 disease alleles.

View details for DOI 10.1167/iovs.15-18680

View details for Web of Science ID 000378041700018

View details for PubMedID 27152965

View details for PubMedCentralID PMC4868102
Chronic Recurrent Pseudophakic Endophthalmitis JAMA OPHTHALMOLOGY Chin, E. K., Almeida, D. R., Mahajan, V. B. 2016; 134 (4): 455-456
View details for DOI 10.1001/jamaophthalmol.2015.3638

View details for Web of Science ID 000373988900025

View details for PubMedID 26868359
Precision Medicine Personalized Proteomics for the Diagnosis and Treatment of Idiopathic Inflammatory Disease JAMA OPHTHALMOLOGY Velez, G., Roybal, C. N., Colgan, D., Tsang, S. H., Bassuk, A. G., Mahajan, V. B. 2016; 134 (4): 444-448
Abstract

To better characterize posterior uveitis, vitreous samples from 15 patients were subjected to antibody arrays, and the expression levels of 200 human cytokines were evaluated. Expression was analyzed by 1-way analysis of variance (significance at P < .01), unsupervised cluster algorithm, and pathway analysis.Unbiased clustering of patients, based on their cytokine expression profile, suggested that particular protein networks and molecular pathways are altered in various forms of uveitis. Expression of interleukin 23 (IL-23), IL-1 receptor I (IL-1RI), IL-17R, tissue inhibitors of metalloproteinase 1 and 2 (TIMP-1 and TIMP-2), insulinlike growth factor-binding protein 2 (IGFBP-2), nerve growth factor (b-NGF), platelet-derived growth factor receptor β polypeptide (PDGFRb), bone morphogenic protein 4 (BMP-4), and stem cell factor (SCF) constituted a common cytokine signature in the vitreous of patients with uveitis. In 1 patient with progressive, idiopathic visual loss, this last-line analysis implicated retinal autoimmunity, a diagnosis that was validated when her serum sample was found to contain antibodies to S-arrestin, a retinal protein and potent cause of autoimmune retinal degeneration.The analysis identifies a common cytokine signature for posterior uveitis and guides the diagnosis of a patient with idiopathic uveitis. Personalized treatment reversed the visual loss, illustrating how proteomic tools may individualize therapy.

View details for DOI 10.1001/jamaophthalmol.2015.5934

View details for Web of Science ID 000373988900022

View details for PubMedID 26848019

View details for PubMedCentralID PMC4833518
Response to Sandford et al.: PRICKLE2 Variants in Epilepsy: A Call for Precision Medicine AMERICAN JOURNAL OF HUMAN GENETICS Mahajan, V. B., Bassuk, A. G. 2016; 98 (3): 590-591
View details for DOI 10.1016/j.ajhg.2016.02.002

View details for Web of Science ID 000372383100022

View details for PubMedID 26942292

View details for PubMedCentralID PMC4800048
Autism Linked to Increased Oncogene Mutations but Decreased Cancer Rate PLOS ONE Darbro, B. W., Singh, R., Zimmerman, M. B., Mahajan, V. B., Bassuk, A. G. 2016; 11 (3)
Abstract

Autism spectrum disorder (ASD) is one phenotypic aspect of many monogenic, hereditary cancer syndromes. Pleiotropic effects of cancer genes on the autism phenotype could lead to repurposing of oncology medications to treat this increasingly prevalent neurodevelopmental condition for which there is currently no treatment. To explore this hypothesis we sought to discover whether autistic patients more often have rare coding, single-nucleotide variants within tumor suppressor and oncogenes and whether autistic patients are more often diagnosed with neoplasms. Exome-sequencing data from the ARRA Autism Sequencing Collaboration was compared to that of a control cohort from the Exome Variant Server database revealing that rare, coding variants within oncogenes were enriched for in the ARRA ASD cohort (p<1.0 x 10(-8)). In contrast, variants were not significantly enriched in tumor suppressor genes. Phenotypically, children and adults with ASD exhibited a protective effect against cancer, with a frequency of 1.3% vs. 3.9% (p<0.001), but the protective effect decreased with age. The odds ratio of neoplasm for those with ASD relative to controls was 0.06 (95% CI: 0.02, 0.19; p<0.0001) in the 0 to 14 age group; 0.35 (95% CI: 0.14, 0.87; p = 0.024) in the 15 to 29 age group; 0.41 (95% CI: 0.15, 1.17; p = 0.095) in the 30 to 54 age group; and 0.49 (95% CI: 0.14, 1.74; p = 0.267) in those 55 and older. Both males and females demonstrated the protective effect. These findings suggest that defects in cellular proliferation, and potentially senescence, might influence both autism and neoplasm, and already approved drugs targeting oncogenic pathways might also have therapeutic value for treating autism.

View details for DOI 10.1371/journal.pone.0149041

View details for Web of Science ID 000371724200016

View details for PubMedID 26934580

View details for PubMedCentralID PMC4774916
Precision Medicine: Genetic Repair of Retinitis Pigmentosa in Patient-Derived Stem Cells SCIENTIFIC REPORTS Bassuk, A. G., Zheng, A., Li, Y., Tsang, S. H., Mahajan, V. B. 2016; 6
Abstract

Induced pluripotent stem cells (iPSCs) generated from patient fibroblasts could potentially be used as a source of autologous cells for transplantation in retinal disease. Patient-derived iPSCs, however, would still harbor disease-causing mutations. To generate healthy patient-derived cells, mutations might be repaired with new gene-editing technology based on the bacterial system of clustered regularly interspersed short palindromic repeats (CRISPR)/Cas9, thereby yielding grafts that require no patient immunosuppression. We tested whether CRISPR/Cas9 could be used in patient-specific iPSCs to precisely repair an RPGR point mutation that causes X-linked retinitis pigmentosa (XLRP). Fibroblasts cultured from a skin-punch biopsy of an XLRP patient were transduced to produce iPSCs carrying the patient's c.3070G > T mutation. The iPSCs were transduced with CRISPR guide RNAs, Cas9 endonuclease, and a donor homology template. Despite the gene's repetitive and GC-rich sequences, 13% of RPGR gene copies showed mutation correction and conversion to the wild-type allele. This is the first report using CRISPR to correct a pathogenic mutation in iPSCs derived from a patient with photoreceptor degeneration. This important proof-of-concept finding supports the development of personalized iPSC-based transplantation therapies for retinal disease.

View details for DOI 10.1038/srep19969

View details for Web of Science ID 000368675500001

View details for PubMedID 26814166

View details for PubMedCentralID PMC4728485
Neuroretinal hypoxic signaling in a new preclinical murine model for proliferative diabetic retinopathy. Signal transduction and targeted therapy Wert, K. J., Mahajan, V. B., Zhang, L., Yan, Y., Li, Y., Tosi, J., Hsu, C. W., Nagasaki, T., Janisch, K. M., Grant, M. B., Mahajan, M., Bassuk, A. G., Tsang, S. H. 2016; 1
Abstract

Diabetic retinopathy (DR) affects approximately one-third of diabetic patients and, if left untreated, progresses to proliferative DR (PDR) with associated vitreous hemorrhage, retinal detachment, iris neovascularization, glaucoma and irreversible blindness. In vitreous samples of human patients with PDR, we found elevated levels of hypoxia inducible factor 1 alpha (HIF1α). HIFs are transcription factors that promote hypoxia adaptation and have important functional roles in a wide range of ischemic and inflammatory diseases. To recreate the human PDR phenotype for a preclinical animal model, we generated a mouse with neuroretinal-specific loss of the von Hippel Lindau tumor suppressor protein, a protein that targets HIF1α for ubiquitination. We found that the neuroretinal cells in these mice overexpressed HIF1α and developed severe, irreversible ischemic retinopathy that has features of human PDR. Rapid progression of retinopathy in these mutant mice should facilitate the evaluation of therapeutic agents for ischemic and inflammatory blinding disorders. In addition, this model system can be used to manipulate the modulation of the hypoxia signaling pathways, for the treatment of non-ocular ischemic and inflammatory disorders.

View details for PubMedID 27195131
Comparison of microbiology and visual outcomes of patients undergoing small-gauge and 20-gauge vitrectomy for endophthalmitis. Clinical ophthalmology (Auckland, N.Z.) Almeida, D. R., Chin, E. K., Shah, S. S., Bakall, B., Gehrs, K. M., Boldt, H. C., Russell, S. R., Folk, J. C., Mahajan, V. B. 2016; 10: 167-172
Abstract

The role of pars plana vitrectomy (PPV) for endophthalmitis has evolved over recent decades but the literature is lacking on comparisons between small-gauge and 20-gauge vitrectomy.To evaluate evolving etiological and microbiological trends in patients undergoing vitrectomy for endophthalmitis and to compare culture-positive rates and visual outcomes between small-gauge (23- and 25-gauge) and 20-gauge instrumentation during vitrectomy for endophthalmitis.Ten-year retrospective comparative case series and prospective laboratory in vitro testing. Tertiary care academic referral center. Patients who underwent PPV for endophthalmitis between 2003 and 2013. Vitreous biopsies were obtained in all cases. The effect of vitrectomy gauge (20-, 23-, and 25-gauge) and vitreous cutting rate (1,500 and 5,000 cuts per minute) on the viability of bacterial culture was evaluated in an in vitro prospective laboratory investigation.Comparison of etiology, microbiology culture-positive rates, and visual outcomes between small-gauge and 20-gauge instrumentation in patients undergoing PPV for infectious endophthalmitis.A total of 61 cases of vitrectomy for endophthalmitis were identified over a 10-year period; of these, 34 were treated with small-gauge (23- and 25-gauge) vitrectomy and 27 were treated with 20-gauge vitrectomy. In the small-gauge group, 12 cases (35.3%) yielded culture-positive results versus 20 cases (74.1%) with culture positivity in the 20-gauge cohort (P=0.002). The most common cause of endophthalmitis was cataract surgery and the most frequently identified organism was coagulase-negative Staphylococci in both groups. There was no significant difference in mean postoperative visual acuities between groups (P=0.33). Etiological trends indicate an increase in endophthalmitis due to intravitreal injection in the small-gauge group (n=9) compared to the 20-gauge group (n=3) (P=0.001). In vitro laboratory testing revealed no significant difference in rates of culture growth for different vitrectomy gauge sizes or vitreous cutting speeds.Small-gauge vitrectomy for endophthalmitis yields final visual outcomes comparable to 20-gauge instrumentation. A significant difference in culture-positive rates was observed between small-gauge and 20-gauge instrumentation for vitrectomy in endophthalmitis; however, laboratory testing indicates this is not related to either vitreous gauge size or cutter speed. Intravitreal injections are emerging as a common etiology of vitrectomy for endophthalmitis.

View details for DOI 10.2147/OPTH.S95906

View details for PubMedID 26858522

View details for PubMedCentralID PMC4730992
Secondary glaucoma in CAPN5-associated neovascular inflammatory vitreoretinopathy. Clinical ophthalmology (Auckland, N.Z.) Cham, A., Bansal, M., Banda, H. K., Kwon, Y., Tlucek, P. S., Bassuk, A. G., Tsang, S. H., Sobol, W. M., Folk, J. C., Yeh, S., Mahajan, V. B. 2016; 10: 1187-1197
Abstract

The objective of this study was to review the treatment outcomes of patients with secondary glaucoma in cases of autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV), a hereditary autoimmune uveitis due to mutations in CAPN5.A retrospective, observational case series was assembled from ADNIV patients with secondary glaucoma. The main outcome measures were intraocular pressure (IOP), visual acuity, use of antiglaucoma medications, ocular surgeries, and adverse outcomes. Perimetry and optic disk optical coherence tomography (OCT) were also analyzed.Nine eyes of five ADNIV patients with secondary glaucoma were reviewed. Each received a fluocinolone acetonide (FA) implant for the management of posterior uveitis. Following implantation, no eyes developed neovascular glaucoma. Five eyes (in patients 1, 2, and 5) required Ahmed glaucoma valve surgery for the management of steroid-responsive glaucoma. Patient 2 also developed angle closure with iris bombe and underwent laser peripheral iridotomy. Patient 4 had both hypotony and elevated IOP that required periodic antiglaucoma medication in the FA-implanted eye. Patient 3 did not develop steroid-response glaucoma in either eye. Optic disk examinations were obscured by fibrosis and better assessed with OCT.ADNIV patients show combined mechanism secondary glaucoma best assessed by OCT of the optic disk. The FA implants have reduced uveitic and neovascular glaucoma. Nevertheless, IOP management remains complex due to steroid-response glaucoma, angle closure glaucoma, and hypotony.

View details for DOI 10.2147/OPTH.S103324

View details for PubMedID 27390515

View details for PubMedCentralID PMC4930228
Intravitreal Anti-VEGF Injections in Pregnancy: Case Series and Review of Literature JOURNAL OF OCULAR PHARMACOLOGY AND THERAPEUTICS Polizzi, S., Mahajan, V. B. 2015; 31 (10): 605-610
Abstract

The use of intravitreal antivascular endothelial growth factor (anti-VEGF) injection is gaining wide acceptance as an off-label therapy for diseases that may affect pregnant women. However, these drugs may cause systemic side effects in the mother and fetal harm. This could lead specialists to not administer the drug or women to abort the fetus or to refuse treatment during pregnancy. We report the course of pregnancy in 3 women treated with intravitreal bevacizumab and provide a review of the literature on the use of intravitreal anti-VEGF in pregnancy. Our patients did not have any drug-related adverse event and delivered healthy full-term infants, although one of the women had risk factors for miscarriage. Infants reached all developmental milestones appropriately during infancy. A literature search on the use of intravitreal anti-VEGF injection in pregnancy was undertaken. Data for this review were identified by searches of PubMed and references from relevant articles using the search terms "pegaptanib," "bevacizumab," "ranibizumab," "aflibercept," "anti-VEGF," "intravitreal injection," "pregnant," "pregnancy," "abortion," "miscarriage," "preeclampsia," "embryo-fetal toxicity," "fetal malformations," "teratogenesis," "adverse events," and "maternofetal complications" in multiple combinations. We believe that intravitreal anti-VEGF can be given during pregnancy only when potential benefit to the woman justifies the potential risks to the fetus. When making a decision about whether to give drugs during pregnancy, it is important to consider the timing of exposure and its relationship to windows of developmental sensitivity. We believe that this review will be useful to specialists to inform and possibly treat their pregnant patients.

View details for DOI 10.1089/jop.2015.0056

View details for Web of Science ID 000365694500003

View details for PubMedID 26302032

View details for PubMedCentralID PMC4677108
Management of Choroidal Granulomas Involving the Macula in Corticosteroid-Intolerant Patients JAMA OPHTHALMOLOGY Chin, E. K., Almeida, D. R., Mahajan, V. B. 2015; 133 (11): 1351-1352
View details for DOI 10.1001/jamaophthalmol.2015.1951

View details for Web of Science ID 000365034000027

View details for PubMedID 26356326
CAPN5 mutation in hereditary uveitis: the R243L mutation increases calpain catalytic activity and triggers intraocular inflammation in a mouse model HUMAN MOLECULAR GENETICS Wert, K. J., Bassuk, A. G., Wu, W., Gakhar, L., Coglan, D., Mahajan, M., Wu, S., Yang, J., Lin, C., Tsang, S. H., Mahajan, V. B. 2015; 24 (16): 4584-4598
Abstract

A single amino acid mutation near the active site of the CAPN5 protease was linked to the inherited blinding disorder, autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV, OMIM #193235). In homology modeling with other calpains, this R243L CAPN5 mutation was situated in a mobile loop that gates substrate access to the calcium-regulated active site. In in vitro activity assays, the mutation increased calpain protease activity and made it far more active at low concentrations of calcium. To test whether the disease allele could yield an animal model of ADNIV, we created transgenic mice expressing human (h) CAPN5(R243L) only in the retina. The resulting hCAPN5(R243L) transgenic mice developed a phenotype consistent with human uveitis and ADNIV, at the clinical, histological and molecular levels. The fundus of hCAPN5(R243L) mice showed enhanced autofluorescence (AF) and pigment changes indicative of reactive retinal pigment epithelial cells and photoreceptor degeneration. Electroretinography showed mutant mouse eyes had a selective loss of the b-wave indicating an inner-retina signaling defect. Histological analysis of mutant mouse eyes showed protein extravasation from dilated vessels into the anterior chamber and vitreous, vitreous inflammation, vitreous and retinal fibrosis and retinal degeneration. Analysis of gene expression changes in the hCAPN5(R243L) mouse retina showed upregulation of several markers, including members of the Toll-like receptor pathway, chemokines and cytokines, indicative of both an innate and adaptive immune response. Since many forms of uveitis share phenotypic characteristics of ADNIV, this mouse offers a model with therapeutic testing utility for ADNIV and uveitis patients.

View details for DOI 10.1093/hmg/ddv189

View details for Web of Science ID 000361315400009

View details for PubMedID 25994508

View details for PubMedCentralID PMC4512628
Macular Hole Closure With Internal Limiting Membrane Abrasion Technique JAMA OPHTHALMOLOGY Mahajan, V. B., Chin, E. K., Tarantola, R. M., Almeida, D. R., Somani, R., Boldt, H. C., Folk, J. C., Gehrs, K. M., Russell, S. R. 2015; 133 (6): 635-641
Abstract

Internal limiting membrane (ILM) abrasion is an alternative surgical technique for successful full-thickness macular hole (MH) repair.To study the effects of ILM abrasion as an alternative method of MH repair.Retrospective consecutive case series from January 2006 to December 2008. Demographic data and preoperative, intraoperative, and postoperative examination records of all patients were reviewed for patients who underwent ILM abrasion with a diamond-dusted membrane scraper during vitrectomy for MH repair. A total of 100 eyes underwent ILM abrasion as an alternative to traditional ILM peeling.Rate of MH closure and visual acuity (VA) outcomes at 3 months after surgery.Macular hole closure was achieved with a single surgical procedure in 94 of 100 eyes (94.0%; 95% CI, 87.4%-97.8%). Among all patients, the median preoperative VA was 20/100 (range, 20/30 to hand motions; 25th quartile, 20/60; and 75th quartile, 20/160), and the median postoperative VA at 3 months after surgery was 20/60 (range, 20/20 to hand motions; 25th quartile, 20/40; and 75th quartile, 20/100). Among all patients with stage 2 MHs, 30 of 38 patients (78.9%) had at least 2 lines of VA gain: 15 of 23 (65.2%) were phakic, and 15 of 15 (100%) were pseudophakic. Four of 38 patients (10.5%) with stage 2 MHs had at least 2 lines of VA loss, and all were phakic. Among all patients with stage 3 or 4 MHs, 42 of 62 (67.7%) had at least 2 lines of VA gain, of which 30 of 38 (78.9%) were phakic and 22 of 24 (91.7%) were pseudophakic. Six of 62 patients (9.7%) with stage 3 or 4 MHs had at least 2 lines of VA loss: 4 were phakic, and 2 were pseudophakic. In total, 35.0% (95% CI, 25.7%-44.3%) of patients achieved 20/40 vision or better, and 52.0% (95% CI, 42.2%-61.8%) of patients achieved 20/50 vision or better.Abrasion of the ILM with a diamond-dusted membrane scraper at the time of vitrectomy achieves high rates of MH closure. This technique avoids complete removal of the retinal ILM basement membrane and subjacent tissues and appears to provide MH closure rates similar to those of traditional ILM peeling.

View details for DOI 10.1001/jamaophthalmol.2015.204

View details for Web of Science ID 000356044400012

View details for PubMedID 25764352
Proteomic Insight into the Molecular Function of the Vitreous PLOS ONE Skeie, J. M., Roybal, C. N., Mahajan, V. B. 2015; 10 (5)
Abstract

The human vitreous contains primarily water, but also contains proteins which have yet to be fully characterized. To gain insight into the four vitreous substructures and their potential functions, we isolated and analyzed the vitreous protein profiles of three non-diseased human eyes. The four analyzed substructures were the anterior hyaloid, the vitreous cortex, the vitreous core, and the vitreous base. Proteins were separated by multidimensional liquid chromatography and identified by tandem mass spectrometry. Bioinformatics tools then extracted the expression profiles, signaling pathways, and interactomes unique to each tissue. From each substructure, a mean of 2,062 unique proteins were identified, with many being differentially expressed in a specific substructure: 278 proteins were unique to the anterior hyaloid, 322 to the vitreous cortex, 128 to the vitreous base, and 136 to the vitreous core. When the identified proteins were organized according to relevant functional pathways and networks, key patterns appeared. The blood coagulation pathway and extracellular matrix turnover networks were highly represented. Oxidative stress regulation and energy metabolism proteins were distributed throughout the vitreous. Immune functions were represented by high levels of immunoglobulin, the complement pathway, damage-associated molecular patterns (DAMPs), and evolutionarily conserved antimicrobial proteins. The majority of vitreous proteins detected were intracellular proteins, some of which originate from the retina, including rhodopsin (RHO), phosphodiesterase 6 (PDE6), and glial fibrillary acidic protein (GFAP). This comprehensive analysis uncovers a picture of the vitreous as a biologically active tissue, where proteins localize to distinct substructures to protect the intraocular tissues from infection, oxidative stress, and energy disequilibrium. It also reveals the retina as a potential source of inflammatory mediators. The vitreous proteome catalogues the dynamic interactions between the vitreous and surrounding tissues. It therefore could be an indirect and effective method for surveying vitreoretinal disease for specific biomarkers.

View details for DOI 10.1371/journal.pone.0127567

View details for Web of Science ID 000355187300045

View details for PubMedID 26020955

View details for PubMedCentralID PMC4447289
Effect of Internal Limiting Membrane Abrasion on Retinal Tissues in Macular Holes INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE Almeida, D. R., Chin, E. K., Tarantola, R. M., Folk, J. C., Boldt, H. C., Skeie, J. M., Mullins, R. F., Russell, S. R., Mahajan, V. B. 2015; 56 (5): 2783-2789
Abstract

The purpose of this study was to identify the structural and histological effects of a Tano diamond-dusted membrane scraper (DDMS) on the retinal surface after internal limiting membrane (ILM) abrasion in macular hole surgery.Institutional experimental study was performed in 11 eyes. All eyes underwent ILM abrasion in the operating room with a DDMS for macular hole repair as an alternative to traditional ILM peeling. Three human donor eyes underwent an identical procedure in the laboratory. Retinal tissues were removed by ILM abrasion with a DDMS during vitrectomy for macular hole repair and retinal tissues remaining in human donor eyes. Main outcome measures were microscopic and immunohistological characteristics of instrument tip tissues and retinal structure after ILM abrasion.The tips of the Tano DDMS showed evidence of cellular membranes and ILM removal. The retinas showed distinct areas of lamellar ILM removal without penetration of the retinal nerve fiber layer (RNFL).Application of the Tano DDMS instrument is sufficient to remove membranes from the surface of the ILM and layers of the ILM without disruption of the underlying RNFL. Internal limiting membrane abrasion can be a useful and effective alternative to complete ILM removal for macular surgery.

View details for DOI 10.1167/iovs.14-16355

View details for Web of Science ID 000356439200001

View details for PubMedID 26024069

View details for PubMedCentralID PMC4416746
Structural Modeling of a Novel CAPN5 Mutation that Causes Uveitis and Neovascular Retinal Detachment PLOS ONE Bassuk, A. G., Yeh, S., Wu, S., Martin, D. F., Tsang, S. H., Gakhar, L., Mahajan, V. B. 2015; 10 (4)
Abstract

CAPN5 mutations have been linked to autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV), a blinding autoimmune eye disease. Here, we link a new CAPN5 mutation to ADNIV and model the three-dimensional structure of the resulting mutant protein. In our study, a kindred with inflammatory vitreoretinopathy was evaluated by clinical eye examinations, DNA sequencing, and protein structural modeling to investigate the disease-causing mutation. Two daughters of an affected mother demonstrated symptoms of stage III ADNIV, with posterior uveitis, cystoid macular edema, intraocular fibrosis, retinal neovascularization, retinal degeneration, and cataract. The women also harbored a novel guanine to thymine (c.750G>T, p.Lys250Asn) missense mutation in exon 6 of CAPN5, a gene that encodes a calcium-activated cysteine protease, calpain-5. Modeling based on the structures of all known calpains revealed the mutation falls within a calcium-sensitive flexible gating loop that controls access to the catalytic groove. Three-dimensional modeling placed the new mutation in a region adjacent to two previously identified disease-causing mutations, all three of which likely disrupt hydrogen bonding within the gating loop, yielding a CAPN5 with altered enzymatic activity. This is the third case of a CAPN5 mutation leading to inherited uveitis and neovascular vitreoretinopathy, suggesting patients with ADNIV features should be tested for CAPN5 mutations. Structural modeling of novel variants can be used to support mechanistic consequences of the disease-causing variants.

View details for DOI 10.1371/journal.pone.0122352

View details for Web of Science ID 000352588500037

View details for PubMedID 25856303

View details for PubMedCentralID PMC4391918
Spontaneous dislocation of a fluocinolone acetonide implant (Retisert) into the anterior chamber and its successful extraction in sympathetic ophthalmia. Retinal cases & brief reports Almeida, D. R., Chin, E. K., Mears, K., Russell, S. R., Mahajan, V. B. 2015; 9 (2): 142-144
Abstract

To report a case of spontaneous fluocinolone acetonide (Retisert) implant dislocation and migration into the anterior chamber.Retrospective case report.One patient with sympathetic ophthalmia uveitis was well controlled by fluocinolone acetonide implants.The fluocinolone acetonide implant spontaneously migrated into the anterior chamber causing corneal edema and anterior chamber reaction. The implant was subsequently successfully removed through a temporal corneal wound.Clinicians should be aware of the spontaneous dislocation and migration of an intact fluocinolone implant as a potential surgical complication, particularly in pseudophakic eyes with iridectomy.

View details for DOI 10.1097/ICB.0000000000000119

View details for PubMedID 25411930
Seizures Are Regulated by Ubiquitin-specific Peptidase 9 X-linked (USP9X), a De-Ubiquitinase PLOS GENETICS Paemka, L., Mahajan, V. B., Ehaideb, S. N., Skeie, J. M., Tan, M. C., Wu, S., Cox, A. J., Sowers, L. P., Gecz, J., Jolly, L., Ferguson, P. J., Darbro, B., Schneider, A., Scheffer, I. E., Carvill, G. L., Mefford, H. C., El-Shanti, H., Wood, S. A., Manak, J. R., Bassuk, A. G. 2015; 11 (3)
Abstract

Epilepsy is a common disabling disease with complex, multifactorial genetic and environmental etiology. The small fraction of epilepsies subject to Mendelian inheritance offers key insight into epilepsy disease mechanisms; and pathologies brought on by mutations in a single gene can point the way to generalizable therapeutic strategies. Mutations in the PRICKLE genes can cause seizures in humans, zebrafish, mice, and flies, suggesting the seizure-suppression pathway is evolutionarily conserved. This pathway has never been targeted for novel anti-seizure treatments. Here, the mammalian PRICKLE-interactome was defined, identifying prickle-interacting proteins that localize to synapses and a novel interacting partner, USP9X, a substrate-specific de-ubiquitinase. PRICKLE and USP9X interact through their carboxy-termini; and USP9X de-ubiquitinates PRICKLE, protecting it from proteasomal degradation. In forebrain neurons of mice, USP9X deficiency reduced levels of Prickle2 protein. Genetic analysis suggests the same pathway regulates Prickle-mediated seizures. The seizure phenotype was suppressed in prickle mutant flies by the small-molecule USP9X inhibitor, Degrasyn/WP1130, or by reducing the dose of fat facets a USP9X orthologue. USP9X mutations were identified by resequencing a cohort of patients with epileptic encephalopathy, one patient harbored a de novo missense mutation and another a novel coding mutation. Both USP9X variants were outside the PRICKLE-interacting domain. These findings demonstrate that USP9X inhibition can suppress prickle-mediated seizure activity, and that USP9X variants may predispose to seizures. These studies point to a new target for anti-seizure therapy and illustrate the translational power of studying diseases in species across the evolutionary spectrum.

View details for DOI 10.1371/journal.pgen.1005022

View details for Web of Science ID 000352197100018

View details for PubMedID 25763846

View details for PubMedCentralID PMC4357451
Quantitative Autofluorescence as a Clinical Tool for Expedited Differential Diagnosis of Retinal Degeneration JAMA OPHTHALMOLOGY Marsiglia, M., Lee, W., Mahajan, V. B., Zernant, J., Delori, F. C., Tsang, S. H., Sparrow, J. R. 2015; 133 (2): 219-220
View details for DOI 10.1001/jamaophthalmol.2014.4507

View details for Web of Science ID 000349673100016

View details for PubMedID 25375877

View details for PubMedCentralID PMC4370211
Scleral buckle hemorrhagic cyst masquerading as an orbital tumor. Clinical ophthalmology (Auckland, N.Z.) Almeida, D. R., Chin, E. K., Boldt, H. C., Mahajan, V. B. 2015; 9: 343-345
Abstract

We present a case of a 41-year-old man who was referred for evaluation of a choroidal tumor with a remote history of scleral buckle placement for traumatic retinal detachment. Ocular imaging, echography, and magnetic resonance imaging could not rule out a neoplastic process so the patient was taken for surgical exploration where a hemorrhagic cyst was discovered. This is the first case in the literature of a silicone scleral buckle-associated hemorrhagic cyst presenting as orbital mass.

View details for DOI 10.2147/OPTH.S76351

View details for PubMedID 25709397

View details for PubMedCentralID PMC4335608
The metabolic syndrome and severity of diabetic retinopathy. Clinical ophthalmology (Auckland, N.Z.) Chen, J. J., Wendel, L. J., Birkholz, E. S., Vallone, J. G., Coleman, A. L., Yu, F., Mahajan, V. B. 2015; 9: 757-764
Abstract

While metabolic syndrome has been strongly implicated as a risk factor for macrovascular diseases, such as stroke and cardiovascular disease, its relationship with microvascular diseases, including diabetic retinopathy, has been less defined. The purpose of this pilot study was to investigate the association between metabolic syndrome and the presence and severity of diabetic retinopathy.A retrospective case-control chart review at the University of Iowa ophthalmology and primary care clinics included 100 patients with proliferative diabetic retinopathy (PDR), 100 patients with nonproliferative diabetic retinopathy (NPDR), 100 diabetic patients without diabetic retinopathy, and 100 nondiabetic patients who were randomly selected. Using the International Diabetes Foundation definition, the prevalence of metabolic syndrome and the number of components of metabolic syndrome were compared among these groups.The prevalence of metabolic syndrome in patients with diabetes was 69.3%, which was significantly higher than that in patients without diabetes (27%; P<0.0001) (odds ratio [OR] =6.28; 95% confidence interval [CI]: 3.76-10.49; P=0.0004). However, there was no significant difference in the prevalence of metabolic syndrome between diabetics with and without diabetic retinopathy, with rates of 67.5% and 73%, respectively (P=0.36) (OR =0.77; 95% CI: 0.45-1.32; P=0.34). In addition, there was no significant difference between the PDR and NPDR groups, with rates of 63% and 72%, respectively (P=0.23) (OR =0.70; 95% CI: 0.38-1.30; P=0.26).The metabolic syndrome was highly prevalent in patients with diabetes, but it was not associated with the presence or severity of retinopathy.

View details for DOI 10.2147/OPTH.S80355

View details for PubMedID 25995613

View details for PubMedCentralID PMC4425343
Proteomic Landscape of the Human Choroid-Retinal Pigment Epithelial Complex JAMA OPHTHALMOLOGY Skeie, J. M., Mahajan, V. B. 2014; 132 (11): 1271-1281
Abstract

Differences in geographical protein expression in the human choroid-retinal pigment epithelial (RPE) complex may explain molecular predisposition of regions to ophthalmic diseases such as age-related macular degeneration.To characterize the proteome of the human choroid-RPE complex and to identify differentially expressed proteins in specific anatomic regions.Experimental study of choroid-RPE tissue from 3 nondiseased eyes. The choroid-RPE complex underwent biopsy from beneath the foveal, macular, and peripheral retina. Protein fractions were isolated and subjected to multidimensional liquid chromatography and tandem mass spectrometry. A bioinformatic pipeline matched peptide spectra to the human proteome, assigned gene ontology classification, and identified protein signaling pathways unique to each of the choroid-RPE regions.Mean number of mass spectra, statistically significant differentially expressed proteins, gene ontology classification, and pathway representation.We identified a mean of 4403 unique proteins in each of the foveal, macular, and peripheral choroid-RPE tissues. Six hundred seventy-one differentially expressed proteins included previously known risk factors for retinal diseases related to oxidative stress, inflammation, and the complement cascade. Gene ontology analysis showed that unique categories in the foveal and macular regions included immune process proteins as well as protein complexes and plasma membrane proteins. The peripheral region contained unique antioxidant activity proteins. Many proteins had the highest expression in the foveal or macular regions, including inflammation-related proteins HLA-A, HLA-B, and HLA-C antigens; intercellular adhesion molecule 1 (ICAM-1); S100; transcription factor ERG; antioxidant superoxide dismutase 1 (SOD1); chloride intracellular channel 6 ion (CLIC6); activators of the complement cascade C1q, C6, and C8; and complement factor H. Proteins with higher expression in the periphery included bestrophin 1 (BEST1), transcription factor RNA binding motif protein 39 (RBM39), inflammatory mediator macrophage migration inhibitory factor, antioxidant SOD3, ion channel voltage-dependent anion-selective channel protein 3 (VDAC3), and complement inhibitor CD55. The complement activation was among the highest represented pathways (P < 7.5e-13).This proteomic data set identifies novel molecular signatures in anatomically sensitive regions of the choroid-RPE complex. The findings give mechanistic insight into choroid-RPE function, reveal important choroid-RPE processes, and prioritize new pathways for therapeutic targeting.

View details for DOI 10.1001/jamaophthalmol.2014.2065

View details for Web of Science ID 000345210100002

View details for PubMedID 25058583
Spinster Homolog 2 (Spns2) Deficiency Causes Early Onset Progressive Hearing Loss PLOS GENETICS Chen, J., Ingham, N., Kelly, J., Jadeja, S., Goulding, D., Pass, J., Mahajan, V. B., Tsang, S. H., Nijnik, A., Jackson, I. J., White, J. K., Forge, A., Jagger, D., Steel, K. P. 2014; 10 (10)
Abstract

Spinster homolog 2 (Spns2) acts as a Sphingosine-1-phosphate (S1P) transporter in zebrafish and mice, regulating heart development and lymphocyte trafficking respectively. S1P is a biologically active lysophospholipid with multiple roles in signalling. The mechanism of action of Spns2 is still elusive in mammals. Here, we report that Spns2-deficient mice rapidly lost auditory sensitivity and endocochlear potential (EP) from 2 to 3 weeks old. We found progressive degeneration of sensory hair cells in the organ of Corti, but the earliest defect was a decline in the EP, suggesting that dysfunction of the lateral wall was the primary lesion. In the lateral wall of adult mutants, we observed structural changes of marginal cell boundaries and of strial capillaries, and reduced expression of several key proteins involved in the generation of the EP (Kcnj10, Kcnq1, Gjb2 and Gjb6), but these changes were likely to be secondary. Permeability of the boundaries of the stria vascularis and of the strial capillaries appeared normal. We also found focal retinal degeneration and anomalies of retinal capillaries together with anterior eye defects in Spns2 mutant mice. Targeted inactivation of Spns2 in red blood cells, platelets, or lymphatic or vascular endothelial cells did not affect hearing, but targeted ablation of Spns2 in the cochlea using a Sox10-Cre allele produced a similar auditory phenotype to the original mutation, suggesting that local Spns2 expression is critical for hearing in mammals. These findings indicate that Spns2 is required for normal maintenance of the EP and hence for normal auditory function, and support a role for S1P signalling in hearing.

View details for DOI 10.1371/journal.pgen.1004688

View details for Web of Science ID 000344650700052

View details for PubMedID 25356849

View details for PubMedCentralID PMC4214598
CAPN5 gene silencing by short hairpin RNA interference. BMC research notes Nelson, N. G., Skeie, J. M., Muradov, H., Rowell, H. A., Seo, S., Mahajan, V. B. 2014; 7: 642-?
Abstract

The purpose of this project was to identify short hairpin RNA (shRNA) sequences that can suppress expression of human CAPN5 in which gain-of-function mutants cause autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV). We created HEK293T cells that stably express an ADNIV disease allele, CAPN5-p.R243L. Transfection protocols were optimized for neuroblastoma SHSY5Y cells. The gene silencing effect of four different shRNA plasmids that target CAPN5 was tested. RNA and protein expression was determined using quantitative RT-PCR and immunoblot analysis.Two of four shRNA plasmids reduced mutant CAPN5 RNA in a stable cell line. Similar knockdown was observed in SH-SY5Y cells that natively express CAPN5. Lactose dehydrogenase assays showed that down-regulation of CAPN5 was not cytotoxic.CAPN5 expression can be suppressed by shRNA-based RNA interference. Further testing in ADNIV models will determine the potential of gene silencing as a strategy to treat, delay, or prevent blindness in ADNIV patients.

View details for DOI 10.1186/1756-0500-7-642

View details for PubMedID 25216694

View details for PubMedCentralID PMC4169796
Incomplete vitreomacular traction release using intravitreal ocriplasmin. Case reports in ophthalmology Chin, E. K., Almeida, D. R., Sohn, E. H., Boldt, H. C., Mahajan, V. B., Gehrs, K. M., Russell, S. R., Folk, J. C. 2014; 5 (3): 455-462
Abstract

To report the clinical course of our first 7 consecutive patients treated with intravitreal ocriplasmin (Jetrea(®)).Retrospective case series of the first 7 patients treated with ocriplasmin between January and December 2013 at an academic tertiary care center.The average age was 78.4 years (range: 63-92). Five patients were pseudophakic and 2 patients were phakic in the injected eye. The median baseline visual acuity (VA) was 20/60 (range: 20/25 to 20/200). The median 1-month postinjection VA was 20/70, with a mean loss of 2 lines of VA among all patients. None of the patients had complete resolution of their vitreomacular traction or macular hole at 1 month of follow-up. Three patients had subsequent pars plana vitrectomy and membrane peeling surgery. The mean follow-up period for those who did not undergo vitrectomy was 9 months (range: 1-13). One patient with known ocular hypertension had an increase in intraocular pressure requiring topical pressure-lowering eyedrops. There were no cases of postinjection uveitis, endophthalmitis, retinal tears, or retinal detachment.While ocriplasmin may be a viable pharmacological agent for vitreolysis, we present a series of patients that all had incomplete resolution of vitreomacular traction with and without full-thickness macular hole. There was an associated reduction in VA after ocriplasmin treatment at 1 month of follow-up. Careful analysis of the vitreoretinal interface and comorbid eye conditions is required to optimize outcome success with ocriplasmin.

View details for DOI 10.1159/000370024

View details for PubMedID 25606039

View details for PubMedCentralID PMC4296250
DECREASED MACULAR THICKNESS IN NONPROLIFERATIVE MACULAR TELANGIECTASIA TYPE 2 WITH ORAL CARBONIC ANHYDRASE INHIBITORS RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Chen, J. J., Sohn, E. H., Folk, J. C., Mahajan, V. B., Kay, C. N., Boldt, H. C., Russell, S. R. 2014; 34 (7): 1400-1406
Abstract

To evaluate whether carbonic anhydrase inhibitors reduce the macular thickness and/or cystic spaces in patients with macular telangiectasia (MacTel) Type 2.Retrospective review of patients with nonproliferative cystoid changes associated with MacTel seen at the University of Iowa between 2009 and 2012. Carbonic anhydrase inhibitors were used in 8 patients with MacTel Type 2. Five patients with MacTel Type 2 were observed during this period. Initial and final visual acuities were documented. The presence of cystic spaces and the retinal thickness were measured with spectral-domain optical coherence tomography.Patients treated with oral carbonic anhydrase inhibitors showed significant reduction in both the cystoid cavities and central macular thickness when compared with the patients who were observed (-12.2 μm; P = 0.020). The reduction in retinal thickness was more pronounced in patients receiving acetazolamide (-20.13 μm; P = 0.007) compared with methazolamide (-6.25 μm; P = 0.177). There was no significant change in visual acuity in patients receiving carbonic anhydrase inhibitors. Five patients with MacTel Type 2 did not receive treatment and demonstrated no change in visual acuity, cystoid cavities, or central macular thickness.Oral carbonic anhydrase inhibitors, particularly acetazolamide, may decrease macular cystic cavities and reduce central macular thickness but does not appear to improve visual acuity. These findings have yet to be confirmed with a prospective treatment trial.

View details for Web of Science ID 000338772600019

View details for PubMedID 24451922
Silencing of tuberin enhances photoreceptor survival and function in a preclinical model of retinitis pigmentosa (an american ophthalmological society thesis). Transactions of the American Ophthalmological Society Tsang, S. H., Chan, L., Tsai, Y., Wu, W., Hsu, C., Yang, J., Tosi, J., Wert, K. J., Davis, R. J., Mahajan, V. B. 2014; 112: 103-115
Abstract

To assess the functional consequences of silencing of tuberin, an inhibitor of the mTOR signaling pathway, in a preclinical model of retinitis pigmentosa (RP) in order to test the hypothesis that insufficient induction of the protein kinase B (PKB)-regulated tuberin/mTOR self-survival pathway initiates apoptosis.In an unbiased genome-scale approach, kinase peptide substrate arrays were used to analyze self-survival pathways at the onset of photoreceptor degeneration. The mutant Pde6b(H620Q)/Pde6b(H620Q) at P14 and P18 photoreceptor outer segment (OS) lysates were labeled with P-ATP and hybridized to an array of 1,164 different synthetic peptide substrates. At this stage, OS of Pde6b(H620Q)/Pde6b(H620Q) rods are morphologically normal. In vitro kinase assays and immunohistochemistry were used to validate phosphorylation. Short hairpin RNA (shRNA) gene silencing was used to validate tuberin's role in regulating survival.At the onset of degeneration, 162 peptides were differentially phosphorylated. Protein kinases A, G, C (AGC kinases), and B exhibited increased activity in both peptide array and in vitro kinase assays. Immunohistochemical data confirmed altered phosphorylation patterns for phosphoinositide-dependent kinase-1 (PDK1), ribosomal protein S6 (RPS6), and tuberin. Tuberin gene silencing rescued photoreceptors from degeneration.Phosphorylation of tuberin and RPS6 is due to the upregulated activity of PKB. PKB/tuberin cell growth/survival signaling is activated before the onset of degeneration. Substrates of the AGC kinases in the PKB/tuberin pathway are phosphorylated to promote cell survival. Knockdown of tuberin, the inhibitor of the mTOR pathway, increased photoreceptor survival and function in a preclinical model of RP.

View details for PubMedID 25646031
A novel RPGR mutation masquerading as Stargardt disease BRITISH JOURNAL OF OPHTHALMOLOGY Bassuk, A. G., Sujirakul, T., Tsang, S. H., Mahajan, V. B. 2014; 98 (5): 709-?
View details for DOI 10.1136/bjophthalmol-2013-304822

View details for Web of Science ID 000334397400027

View details for PubMedID 24489377

View details for PubMedCentralID PMC4170590
Translational vitreous proteomics PROTEOMICS CLINICAL APPLICATIONS Mahajan, V. B., Skeie, J. M. 2014; 8 (3-4): 204-208
Abstract

The vitreous is an extracellular matrix that is still poorly understood. Although many constituents and characteristics have been previously determined, there are many attributes still being discovered. Currently, using protein arrays, MS, and bioinformatics, the vitreous provides a wealth of knowledge regarding ocular diseases and potential targets for personalized therapeutics.

View details for DOI 10.1002/prca.201300062

View details for Web of Science ID 000334251600009

View details for PubMedID 24115652

View details for PubMedCentralID PMC3964148
Defective Motile Cilia in Prickle2-Deficient Mice JOURNAL OF NEUROGENETICS Sowers, L. P., Yin, T., Mahajan, V. B., Bassuk, A. G. 2014; 28 (1-2): 146-152
Abstract

Motile cilia play diverse roles across phyla and cell types, and abnormalities in motile cilia lead to numerous disease states, including hydrocephalus. Although motile ciliary abnormalities in Prickle2 mutants have not yet been described, the planar cell polarity genes, including Prickle2, are implicated in the development and function of motile cilia. This report evaluates Prickle2-deficient mice for dysfunction in processes known to depend on functioning motile cilia. Prickle2-deficient mice do not develop hydrocephalus, but do display abnormal morphology and motility in the motile cilia of the ependyma. The morphology of tracheal motile cilia is also abnormal. Taken together, these results demonstrate that Prickle2 is required for normal ependymal motile cilia development and function.

View details for DOI 10.3109/01677063.2014.885966

View details for Web of Science ID 000336409000012

View details for PubMedID 24708399
PRICKLE1 Interaction with SYNAPSIN I Reveals a Role in Autism Spectrum Disorders PLOS ONE Paemka, L., Mahajan, V. B., Skeie, J. M., Sowers, L. P., Ehaideb, S. N., Gonzalez-Alegre, P., Sasaoka, T., Tao, H., Miyagi, A., Ueno, N., Takao, K., Miyakawa, T., Wu, S., Darbro, B. W., Ferguson, P. J., Pieper, A. A., Britt, J. K., Wemmie, J. A., Rudd, D. S., Wassink, T., El-Shanti, H., Mefford, H. C., Carvill, G. L., Manak, J. R., Bassuk, A. G. 2013; 8 (12)
Abstract

The frequent comorbidity of Autism Spectrum Disorders (ASDs) with epilepsy suggests a shared underlying genetic susceptibility; several genes, when mutated, can contribute to both disorders. Recently, PRICKLE1 missense mutations were found to segregate with ASD. However, the mechanism by which mutations in this gene might contribute to ASD is unknown. To elucidate the role of PRICKLE1 in ASDs, we carried out studies in Prickle1(+/-) mice and Drosophila, yeast, and neuronal cell lines. We show that mice with Prickle1 mutations exhibit ASD-like behaviors. To find proteins that interact with PRICKLE1 in the central nervous system, we performed a yeast two-hybrid screen with a human brain cDNA library and isolated a peptide with homology to SYNAPSIN I (SYN1), a protein involved in synaptogenesis, synaptic vesicle formation, and regulation of neurotransmitter release. Endogenous Prickle1 and Syn1 co-localize in neurons and physically interact via the SYN1 region mutated in ASD and epilepsy. Finally, a mutation in PRICKLE1 disrupts its ability to increase the size of dense-core vesicles in PC12 cells. Taken together, these findings suggest PRICKLE1 mutations contribute to ASD by disrupting the interaction with SYN1 and regulation of synaptic vesicles.

View details for DOI 10.1371/journal.pone.0080737

View details for Web of Science ID 000327947800020

View details for PubMedID 24312498

View details for PubMedCentralID PMC3849077
Proteomic Interactions in the Mouse Vitreous-Retina Complex PLOS ONE Skeie, J. M., Mahajan, V. B. 2013; 8 (11)
Abstract

Human vitreoretinal diseases are due to presumed abnormal mechanical interactions between the vitreous and retina, and translational models are limited. This study determined whether nonstructural proteins and potential retinal biomarkers were expressed by the normal mouse vitreous and retina.Vitreous and retina samples from mice were collected by evisceration and analyzed by liquid chromatography-tandem mass spectrometry. Identified proteins were further analyzed for differential expression and functional interactions using bioinformatic software.We identified 1,680 unique proteins in the retina and 675 unique proteins in the vitreous. Unbiased clustering identified protein pathways that distinguish retina from vitreous including oxidative phosphorylation and neurofilament cytoskeletal remodeling, whereas the vitreous expressed oxidative stress and innate immunology pathways. Some intracellular protein pathways were found in both retina and vitreous, such as glycolysis and gluconeogenesis and neuronal signaling, suggesting proteins might be shuttled between the retina and vitreous. We also identified human disease biomarkers represented in the mouse vitreous and retina, including carbonic anhydrase-2 and 3, crystallins, macrophage inhibitory factor, glutathione peroxidase, peroxiredoxins, S100 precursors, and von Willebrand factor.Our analysis suggests the vitreous expresses nonstructural proteins that functionally interact with the retina to manage oxidative stress, immune reactions, and intracellular proteins may be exchanged between the retina and vitreous. This novel proteomic dataset can be used for investigating human vitreoretinopathies in mouse models. Validation of vitreoretinal biomarkers for human ocular diseases will provide a critical tool for diagnostics and an avenue for therapeutics.

View details for DOI 10.1371/journal.pone.0082140

View details for Web of Science ID 000327670300074

View details for PubMedID 24312404

View details for PubMedCentralID PMC3843729
Combination Therapy for Neovascular Age-related Macular Degeneration Refractory to Anti-Vascular Endothelial Growth Factor Agents OPHTHALMOLOGY Tozer, K., Roller, A. B., Chong, L. P., Sadda, S., Folk, J. C., Mahajan, V. B., Russell, S. R., Boldt, H. C., Sohn, E. H. 2013; 120 (10): 2029-2034
Abstract

To examine the outcomes of combination anti-vascular endothelial growth factor (VEGF) and photodynamic therapy (PDT) for the treatment of neovascular age-related macular degeneration (AMD) refractory to anti-VEGF monotherapy.Retrospective, interventional case series.Twenty-six eyes of 26 patients treated with anti-VEGF monotherapy for neovascular AMD with persistent subretinal or intraretinal fluid after at least 3 anti-VEGF injections in the 7 months before combination treatment.Combination anti-VEGF treatment and PDT.Visual acuity at 1 or 2, 3, and 6 months and central retinal thickness at 1 or 2, 3, and 6 months. Secondary outcome measures were change in number of fluid-free visits and interval between treatments in the 7 months before and 6 months after combination therapy.Statistically significant improvements in logarithm of the minimum angle of resolution visual acuities were present at 1 month (P = 0.01) and 3 months (P = 0.01). Significant decreases in central subfield retinal thickness on optic coherence tomography (OCT) were seen at 1 month (P = 4×10(-5)), 3 months (P = 3×10(-4)), and 6 months (P = 4×10(-5)) as compared with precombination treatment OCT scans. The percentage of patient visits with no subretinal fluid increased from 0.5% to 41% after the initiation of combination therapy (P = 1×10(-5)). The interval between treatments increased from once every 1.6 months in the 7 months before combination treatment to once every 2.7 months in the 6 months after combination treatment (P = 0.002). No ocular complications attributable to PDT were seen.Rescue therapy with the combination of anti-VEGF and PDT in eyes that have failed anti-VEGF monotherapy resulted in a mean improvement in vision, a decreased central subfield retinal thickness, and an increase in fluid-free intervals.The author(s) have no proprietary or commercial interest in any materials discussed in this article.

View details for DOI 10.1016/j.ophtha.2013.03.016

View details for Web of Science ID 000325086400019

View details for PubMedID 23714319
Mutations in Extracellular Matrix Genes NID1 and LAMC1 Cause Autosomal Dominant Dandy-Walker Malformation and Occipital Cephaloceles HUMAN MUTATION Darbro, B. W., Mahajan, V. B., Gakhar, L., Skeie, J. M., Campbell, E., Wu, S., Bing, X., Millen, K. J., Dobyns, W. B., Kessler, J. A., Jalali, A., Cremer, J., Segre, A., Manak, J. R., Aldinger, K. A., Suzuki, S., Natsume, N., Ono, M., Huynh Dai Hai, H. D., Le Thi Viet, L. T., Loddo, S., Valente, E. M., Bernardini, L., Ghonge, N., Ferguson, P. J., Bassuk, A. G. 2013; 34 (8): 1075-1079
Abstract

We performed whole-exome sequencing of a family with autosomal dominant Dandy-Walker malformation and occipital cephaloceles and detected a mutation in the extracellular matrix (ECM) protein-encoding gene NID1. In a second family, protein interaction network analysis identified a mutation in LAMC1, which encodes a NID1-binding partner. Structural modeling of the NID1-LAMC1 complex demonstrated that each mutation disrupts the interaction. These findings implicate the ECM in the pathogenesis of Dandy-Walker spectrum disorders.

View details for DOI 10.1002/humu.22351

View details for Web of Science ID 000321759900005

View details for PubMedID 23674478

View details for PubMedCentralID PMC3714376
Genome-wide Generation and Systematic Phenotyping of Knockout Mice Reveals New Roles for Many Genes CELL White, J. K., Gerdin, A., Karp, N. A., Ryder, E., Buljan, M., Bussell, J. N., Salisbury, J., Clare, S., Ingham, N. J., Podrini, C., Houghton, R., Estabel, J., Bottomley, J. R., Melvin, D. G., Sunter, D., Adams, N. C., Tannahill, D., Logan, D. W., MacArthur, D. G., Flint, J., Mahajan, V. B., Tsang, S. H., Smyth, I., Watt, F. M., Skarnes, W. C., Dougan, G., Adams, D. J., Ramirez-Solis, R., Bradley, A., Steel, K. P. 2013; 154 (2): 452-464
Abstract

Mutations in whole organisms are powerful ways of interrogating gene function in a realistic context. We describe a program, the Sanger Institute Mouse Genetics Project, that provides a step toward the aim of knocking out all genes and screening each line for a broad range of traits. We found that hitherto unpublished genes were as likely to reveal phenotypes as known genes, suggesting that novel genes represent a rich resource for investigating the molecular basis of disease. We found many unexpected phenotypes detected only because we screened for them, emphasizing the value of screening all mutants for a wide range of traits. Haploinsufficiency and pleiotropy were both surprisingly common. Forty-two percent of genes were essential for viability, and these were less likely to have a paralog and more likely to contribute to a protein complex than other genes. Phenotypic data and more than 900 mutants are openly available for further analysis. PAPERCLIP:

View details for DOI 10.1016/j.cell.2013.06.022

View details for Web of Science ID 000321950700020

View details for PubMedID 23870131

View details for PubMedCentralID PMC3717207
Aflibercept Therapy for Exudative Age-related Macular Degeneration Resistant to Bevacizumab and Ranibizumab AMERICAN JOURNAL OF OPHTHALMOLOGY Bakall, B., Folk, J. C., Boldt, H. C., Sohn, E. H., Stone, E. M., Russell, S. R., Mahajan, V. B. 2013; 156 (1): 15-22
Abstract

To evaluate the outcome of intravitreal injection of aflibercept in cases with exudative age-related macular degeneration, (AMD) resistant to injections of bevacizumab or ranibizumab.Retrospective observational case series.A retrospective chart review at a single institution was conducted to identify patients with exudative AMD and choroidal neovascularization (CNV) in 1 or both eyes resistant to treatment with ranibizumab or bevacizumab who were switched to treatment with at least 3 monthly injections of aflibercept. In total, 36 eyes from 31 patients were included. The demographic data, visual acuities, central macular thickness on optical coherence tomography (OCT), complications, and number of injections were reviewed.The mean patient age was 79 years (range 60-88). There were 13 male and 18 female patients. The number of prior injections with either bevacizumab or ranibizumab ranged from 6-74. After 3 monthly injections of aflibercept, there was a reduction of either subretinal or intraretinal fluid in 18 of 36 (50.0%) of the treated eyes; the amount of fluid remained stable in 15 eyes (41.7%) and worsened in 3 eyes (8.3%). A significant average decrease was observed for the central macular thickness after 3 injections of 65 μm (P = 2.9 × 10(-6)), with no significant change in visual acuity.Aflibercept therapy appears to be beneficial in a subset of patients with neovascular age-related macular degeneration who exhibit recurrent or resistant intraretinal or subretinal fluid following multiple injections with either bevacizumab or ranibizumab.

View details for DOI 10.1016/j.ajo.2013.02.017

View details for Web of Science ID 000321531900004

View details for PubMedID 23706500
ELIMINATION OF INFUSION BUBBLES AND UNCONTROLLED REFLUX IN THE ALCON CONSTELLATION VITRECTOMY VISION SYSTEM RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Russell, S. R., Sohn, E. H., Boldt, H. C., Folk, J. C., Tarantola, R. M., Kay, C. N., Mahajan, V. B. 2013; 33 (4): 803-806
Abstract

To identify the sources and management of 2 problems associated with the Alcon Constellation Vitrectomy (Alcon Laboratories, Inc) System: 1) infusion bubbles and 2) uncontrolled reflux.Surgical and analytical videos were evaluated to identify the source of intraoperative bubbles, which localized to the duckbill valve (DV). Intraoperatively, the authors modified the infusion tubing and its control by removing the DV. The DV was repurposed as a one-way valve to block reflux originating from the vitrectomy console.Twenty consecutive 23-gauge vitrectomies in 20 eyes of 20 subjects from 2 surgeons (S.R.R. and E.H.S.) were reviewed. Infusion bubbles at the DV developed with each transitory tubing pressure drop upon opening of the infusion clamp. Removal of the DV from the infusion line eliminated infusion bubbles in 20 consecutive 23-gauge cases. Adding a one-way valve, which was fashioned from the DV, to the aspiration tubing, resulted in elimination of infusion bubbles and console-originated reflux in 20 eyes. Placement of the DV to block reflux eliminated both uncontrolled and purposeful console-originated reflux.Intraoperative modification of Constellation tubing may eliminate two potentially harmful problems until manufacturer correction is instituted. Because the authors' modified connections represent off-label connectivity, the manufacturer cannot contact potentially affected surgeons or suggest temporary alternative connectivity improvements.

View details for DOI 10.1097/IAE.0b013e31826c20c6

View details for Web of Science ID 000316801900017

View details for PubMedID 23222392
Mcph1-Deficient Mice Reveal a Role for MCPH1 in Otitis Media PLOS ONE Chen, J., Ingham, N., Clare, S., Raisen, C., Vancollie, V. E., Ismail, O., McIntyre, R. E., Tsang, S. H., Mahajan, V. B., Dougan, G., Adams, D. J., White, J. K., Steel, K. P. 2013; 8 (3)
Abstract

Otitis media is a common reason for hearing loss, especially in children. Otitis media is a multifactorial disease and environmental factors, anatomic dysmorphology and genetic predisposition can all contribute to its pathogenesis. However, the reasons for the variable susceptibility to otitis media are elusive. MCPH1 mutations cause primary microcephaly in humans. So far, no hearing impairment has been reported either in the MCPH1 patients or mouse models with Mcph1 deficiency. In this study, Mcph1-deficient (Mcph1(tm1a) (/tm1a) ) mice were produced using embryonic stem cells with a targeted mutation by the Sanger Institute's Mouse Genetics Project. Auditory brainstem response measurements revealed that Mcph1(tm1a) (/tm1a) mice had mild to moderate hearing impairment with around 70% penetrance. We found otitis media with effusion in the hearing-impaired Mcph1(tm1a) (/tm1a) mice by anatomic and histological examinations. Expression of Mcph1 in the epithelial cells of middle ear cavities supported its involvement in the development of otitis media. Other defects of Mcph1(tm1a) (/tm1a) mice included small skull sizes, increased micronuclei in red blood cells, increased B cells and ocular abnormalities. These findings not only recapitulated the defects found in other Mcph1-deficient mice or MCPH1 patients, but also revealed an unexpected phenotype, otitis media with hearing impairment, which suggests Mcph1 is a new gene underlying genetic predisposition to otitis media.

View details for DOI 10.1371/journal.pone.0058156

View details for Web of Science ID 000316849200039

View details for PubMedID 23516444

View details for PubMedCentralID PMC3596415
PROLIFERATIVE VITREORETINOPATHY MAY BE A RISK FACTOR IN COMBINED MACULAR HOLE RETINAL DETACHMENT CASES RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Cunningham, M. A., Tarantola, R. M., Folk, J. C., Sohn, E. H., Boldt, H. C., Graff, J. A., Elkins, K., Russell, S. R., Mahajan, V. B. 2013; 33 (3): 579-585
Abstract

To review the incidence and closure rate of full-thickness macular holes (MH) in cases associated with concomitant rhegmatogenous retinal detachment (RRD).A retrospective consecutive case series was performed from patients undergoing surgical repair of RRD and simultaneous closure of MH. The presence of proliferative vitreoretinopathy (PVR), rates of hole closure and reattachment, and visual acuity outcomes were evaluated.There were a total of 607 RRDs during the study period. The incidence of concomitant MH in RRD cases was 2.3% (14 of 607), and the overall incidence of PVR was 15.8% (96 of 607). All eyes with a MH had a primary break that was distinct from the MH. Five patients did not meet the inclusion criteria for review of the postoperative outcomes. In the remaining 9 patients, the retinal reattachment rate was 100%, and MH closure was achieved in 8 of 9 (89%) eyes after a single surgery. At the time of primary repair, PVR was present in 6 of these 9 cases (66.7%). There was a significant association between the presence of PVR and a concomitant MH (P = 0.0027). The mean preoperative visual acuity was 2.59 ± 0.649 logarithm of the minimum angle of resolution units and significantly improved to 1.23 ± 1.01 logarithm of the minimum angle of resolution units (P = 0.00124). Overall, 88.8% of patients showed an improvement in visual acuity at the final postoperative visit, and a visual acuity of 20/125 or better was achieved in 66.7% of cases.Macular holes combined with a RRD are infrequent, and good anatomical results can be achieved after a simultaneous repair. Also, PVR may be more frequently encountered in this particular subset of RRDs.

View details for DOI 10.1097/IAE.0b013e31826b0c41

View details for Web of Science ID 000315455200016

View details for PubMedID 23222494
INTRAOPERATIVE SCLEROTOMY-RELATED RETINAL BREAKS DURING 23-GAUGE PARS PLANA VITRECTOMY RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Tarantola, R. M., Tsui, J. Y., Graff, J. M., Russell, S. R., Boldt, H. C., Folk, J. C., Mahajan, V. B. 2013; 33 (1): 136-142
Abstract

To study the incidence and characteristics of intraoperative sclerotomy-related retinal breaks encountered during 23-gauge pars plana vitrectomy.A retrospective consecutive case series was assembled from the surgical logs and charts of patients who underwent 23-gauge pars plana vitrectomy. Demographic data and preoperative, intraoperative, and postoperative records were examined.A total 548 eyes met the inclusion criteria. Of them, 145 eyes underwent pars plana vitrectomy for repair of a rhegmatogenous retinal detachment (RRD) and 403 eyes for other indications. Sclerotomy-related retinal breaks were found in 8 of 548 (1.45%) eyes. No breaks were found in the 145 RRD eyes. In non-RRD cases, 8 of 403 (1.98%) eyes had sclerotomy-related breaks. All breaks were adjacent to the superior sclerotomies. The incidence of postoperative retinal detachment was 0% (0 of 403) in the non-RRD group. In eyes with breaks, the primary surgical indication was vitreomacular traction in six of eight eyes and epiretinal membrane in two of eight eyes. Posterior vitreous detachment was absent in six of eight eyes, and six of eight eyes were phakic. Eyes with vitreomacular traction had a significantly higher incidence of breaks (P < 0.0001). Eyes with a surgical indication other than RRD had a higher incidence of breaks, but this was not statistically significant when compared with eyes with RRD (P = 0.087).Pars plana vitrectomy (23-gauge) is associated with a low incidence of sclerotomy-related retinal breaks and postoperative retinal detachments. Eyes with breaks are more likely to be phakic and without a preoperative posterior vitreous detachment. The presence of vitreomacular traction may be a risk factor for the development of intraoperative sclerotomy-related breaks.

View details for DOI 10.1097/IAE.0b013e31825e1d62

View details for Web of Science ID 000313422500017

View details for PubMedID 22791174
Surgical management of fibrotic encapsulation of the fluocinolone acetonide implant in CAPN5-associated proliferative vitreoretinopathy. Clinical ophthalmology (Auckland, N.Z.) Tlucek, P. S., Folk, J. C., Sobol, W. M., Mahajan, V. B. 2013; 7: 1093-1098
Abstract

To review fibrosis of fluocinolone acetonide (FA) implants in subjects with CAPN5 autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV).A retrospective case series was assembled from ADNIV patients in which there was fibrotic encapsulation of a fluocinolone acetonide implant. CAPN5 genotypes and surgical repair techniques were reviewed.Two eyes of two ADNIV patients developed a fibrotic capsule over the fluocinolone acetonide implant. Both patients had Stage IV disease. Patient A had a c.731T > C mutation in the CAPN5 gene and patient B had a c.728G > T mutation. The fibrotic membrane was surgically excised and the implant function was restored.The exuberant fibrotic response in later stages of ADNIV may be resistant to local immunosuppression with steroids. Surgical excision of fibrotic membranes over FA implants can reestablish local steroid delivery in cases of severe proliferative vitreoretinopathy.

View details for DOI 10.2147/OPTH.S43939

View details for PubMedID 23785231

View details for PubMedCentralID PMC3682853
Subretinal injection of gene therapy vectors and stem cells in the perinatal mouse eye. Journal of visualized experiments : JoVE Wert, K. J., Skeie, J. M., Davis, R. J., Tsang, S. H., Mahajan, V. B. 2012
Abstract

The loss of sight affects approximately 3.4 million people in the United States and is expected to increase in the upcoming years.(1) Recently, gene therapy and stem cell transplantations have become key therapeutic tools for treating blindness resulting from retinal degenerative diseases. Several forms of autologous transplantation for age-related macular degeneration (AMD), such as iris pigment epithelial cell transplantation, have generated encouraging results, and human clinical trials have begun for other forms of gene and stem cell therapies.(2) These include RPE65 gene replacement therapy in patients with Leber's congenital amaurosis and an RPE cell transplantation using human embryonic stem (ES) cells in Stargardt's disease.(3-4) Now that there are gene therapy vectors and stem cells available for treating patients with retinal diseases, it is important to verify these potential therapies in animal models before applying them in human studies. The mouse has become an important scientific model for testing the therapeutic efficacy of gene therapy vectors and stem cell transplantation in the eye.(5-8) In this video article, we present a technique to inject gene therapy vectors or stem cells into the subretinal space of the mouse eye while minimizing damage to the surrounding tissue.

View details for DOI 10.3791/4286

View details for PubMedID 23207897
Disruption of Mouse Cenpj, a Regulator of Centriole Biogenesis, Phenocopies Seckel Syndrome PLOS GENETICS McIntyre, R. E., Chavali, P. L., Ismail, O., Carragher, D. M., Sanchez-Andrade, G., Forment, J. V., Fu, B., Velasco-Herrera, M. D., Edwards, A., van der Weyden, L., Yang, F., Ramirez-Solis, R., Estabel, J., Gallagher, F. A., Logan, D. W., Arends, M. J., Tsang, S. H., Mahajan, V. B., Scudamore, C. L., White, J. K., Jackson, S. P., Gergely, F., Adams, D. J. 2012; 8 (11)
Abstract

Disruption of the centromere protein J gene, CENPJ (CPAP, MCPH6, SCKL4), which is a highly conserved and ubiquitiously expressed centrosomal protein, has been associated with primary microcephaly and the microcephalic primordial dwarfism disorder Seckel syndrome. The mechanism by which disruption of CENPJ causes the proportionate, primordial growth failure that is characteristic of Seckel syndrome is unknown. By generating a hypomorphic allele of Cenpj, we have developed a mouse (Cenpj(tm/tm)) that recapitulates many of the clinical features of Seckel syndrome, including intrauterine dwarfism, microcephaly with memory impairment, ossification defects, and ocular and skeletal abnormalities, thus providing clear confirmation that specific mutations of CENPJ can cause Seckel syndrome. Immunohistochemistry revealed increased levels of DNA damage and apoptosis throughout Cenpj(tm/tm) embryos and adult mice showed an elevated frequency of micronucleus induction, suggesting that Cenpj-deficiency results in genomic instability. Notably, however, genomic instability was not the result of defective ATR-dependent DNA damage signaling, as is the case for the majority of genes associated with Seckel syndrome. Instead, Cenpj(tm/tm) embryonic fibroblasts exhibited irregular centriole and centrosome numbers and mono- and multipolar spindles, and many were near-tetraploid with numerical and structural chromosomal abnormalities when compared to passage-matched wild-type cells. Increased cell death due to mitotic failure during embryonic development is likely to contribute to the proportionate dwarfism that is associated with CENPJ-Seckel syndrome.

View details for DOI 10.1371/journal.pgen.1003022

View details for Web of Science ID 000311891600013

View details for PubMedID 23166506

View details for PubMedCentralID PMC3499256
PROTEOMIC ANALYSIS OF VITREOUS BIOPSY TECHNIQUES RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Skeie, J. M., Brown, E. N., Martinez, H. D., Russell, S. R., Birkholz, E. S., Folk, J. C., Boldt, H. C., Gehrs, K. M., Stone, E. M., Wright, M. E., Mahajan, V. B. 2012; 32 (10): 2141-2149
Abstract

To compare vitreous biopsy methods using analysis platforms used in proteomics biomarker discovery.Vitreous biopsies from 10 eyes were collected sequentially using a 23-gauge needle and a 23-gauge vitreous cutter instrument. Paired specimens were evaluated by UV absorbance spectroscopy, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and liquid chromatography tandem mass spectrometry (LC-MS/MS).The total protein concentration obtained with a needle and vitrectomy instrument biopsy averaged 1.10 mg/mL (standard error of the mean = 0.35) and 1.13 mg/mL (standard error of the mean = 0.25), respectively. In eight eyes with low or medium viscidity, there was a very high correlation (R = 0.934) between the biopsy methods. When data from 2 eyes with high viscidity vitreous were included, the correlation was reduced (R = 0.704). The molecular weight protein sodium dodecyl sulfate-polyacrylamide gel electrophoresis profiles of paired needle and vitreous cutter samples were similar, except for a minority of pairs with single band intensity variance. Using LC-MS/MS, equivalent peptides were identified with similar frequencies (R ≥ 0.90) in paired samples.Proteins and peptides collected from vitreous needle biopsies are nearly equivalent to those obtained from a vitreous cutter instrument. This study suggests both techniques may be used for most proteomic and biomarker discovery studies of vitreoretinal diseases, although a minority of proteins and peptides may differ in concentration.

View details for DOI 10.1097/IAE.0b013e3182562017

View details for Web of Science ID 000310520400021

View details for PubMedID 23095728

View details for PubMedCentralID PMC3637028
Inhibition of Neovascularization but Not Fibrosis With the Fluocinolone Acetonide Implant in Autosomal Dominant Neovascular Inflammatory Vitreoretinopathy ARCHIVES OF OPHTHALMOLOGY Tlucek, P. S., Folk, J. C., Orien, J. A., Stone, E. M., Mahajan, V. B. 2012; 130 (11): 1395-1401
Abstract

OBJECTIVE To review the effect of the fluocinolone acetonide implant in subjects with autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV), an inherited autoimmune uveitis. METHODS A retrospective case series was assembled from patients with ADNIV who received fluocinolone acetonide implants. Visual acuity and features of ADNIV, including inflammatory cells, neovascularization, fibrosis, and cystoid macular edema, were reviewed. RESULTS Nine eyes of 5 related patients with ADNIV with uncontrolled inflammation were reviewed. Follow-up ranged from 21.7 to 56.7 months. Visual acuity at implantation ranged from 20/40 to hand motion. Preoperatively, 8 eyes had vitreous cells (a ninth had diffuse vitreous hemorrhage). Eight eyes had cystoid macular edema, 7 had an epiretinal membrane, and 3 had retinal neovascularization. Following implantation, vitreous cells resolved in all eyes and neovascularization regressed or failed to develop. Central macular thickness improved in 4 eyes. During the postoperative course, however, visual acuity continued to deteriorate, with visual acuity at the most recent examination ranging from 20/60 to no light perception. There was also progressive intraocular fibrosis and phthisis in 1 case. Four eyes underwent cataract surgery. Six of the 7 eyes without previous glaucoma surgery had elevated intraocular pressure at some point, and 3 of these required glaucoma surgery. CONCLUSIONS The fluocinolone acetonide implant may inhibit specific features of ADNIV such as inflammatory cells and neovascularization but does not stabilize long-term vision, retinal thickening, or fibrosis. All eyes in this series required cataract extraction, and more than half required surgical intervention for glaucoma. Further studies may identify additional therapies and any benefit of earlier implantation.

View details for DOI 10.1001/archophthalmol.2012.1971

View details for Web of Science ID 000310986700004

View details for PubMedID 22777573

View details for PubMedCentralID PMC3885610
Calpain-5 Mutations Cause Autoimmune Uveitis, Retinal Neovascularization, and Photoreceptor Degeneration PLOS GENETICS Mahajan, V. B., Skeie, J. M., Bassuk, A. G., Fingert, J. H., Braun, T. A., Daggett, H. T., Folk, J. C., Sheffield, V. C., Stone, E. M. 2012; 8 (10)
Abstract

Autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV) is an autoimmune condition of the eye that sequentially mimics uveitis, retinitis pigmentosa, and proliferative diabetic retinopathy as it progresses to complete blindness. We identified two different missense mutations in the CAPN5 gene in three ADNIV kindreds. CAPN5 encodes calpain-5, a calcium-activated cysteine protease that is expressed in retinal photoreceptor cells. Both mutations cause mislocalization from the cell membrane to the cytosol, and structural modeling reveals that both mutations lie within a calcium-sensitive domain near the active site. CAPN5 is only the second member of the large calpain gene family to cause a human Mendelian disorder, and this is the first report of a specific molecular cause for autoimmune eye disease. Further investigation of these mutations is likely to provide insight into the pathophysiologic mechanisms of common diseases ranging from autoimmune disorders to diabetic retinopathy.

View details for DOI 10.1371/journal.pgen.1003001

View details for Web of Science ID 000310528400026

View details for PubMedID 23055945

View details for PubMedCentralID PMC3464205
TEMPORAL APPROACH FOR SMALL-GAUGE PARS PLANA VITRECTOMY COMBINED WITH ANTERIOR SEGMENT SURGERY RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Tarantola, R. M., Graff, J. M., Somani, R., Mahajan, V. B. 2012; 32 (8): 1614-1623
Abstract

To describe our preliminary experience with temporal small-gauge pars plana vitrectomy (PPV) techniques used to treat anterior and posterior segment pathology.A retrospective consecutive case review of patients who underwent temporal PPV was performed. Patients underwent combined temporal small-gauge PPV and anterior segment intervention. Pre- and postoperative visual acuity, intraocular pressure, surgical indications, intraoperative techniques, postoperative course, and a survey to determine how the change in position affected surgery were examined.Temporal PPV was performed on 23 eyes with various posterior segment indications and anterior segment pathologies including cataract, pupillary membrane, endophthalmitis, superior filtering blebs, and anterior vitreous membranes. In 20 eyes, 23-gauge instruments were used, and in 3 eyes, 25-gauge instruments were used. Mean postoperative follow-up duration was 7.6 ± 5.0 months (range, 3-22 months). Surgical objectives were achieved in all cases, and no complications occurred in any study eye. Preoperative logarithm of the minimum angle of resolution mean visual acuity was 1.89 ± 0.76 and improved significantly on postoperative Week 1 (1.45 ± 0.81, P = 0.0003), Month 1 (1.13 ± 0.86, P = 0.0001), and at final follow-up (0.88 ± 0.79, P = 0.0001). There was no significant difference in preoperative and postoperative intraocular pressures. Surgeon surveys indicated significant advantages with a temporal approach for each anterior segment indication, no significant differences in performing the basic surgical steps of PPV, and relative ease of adopting this technique.Performing PPV from the temporal position seems to be advantageous in cases combining posterior and anterior segment surgery such as cataract extraction, pupillary membrane dissection, preservation of superior conjunctival blebs, and trimanual vitrectomy.

View details for DOI 10.1097/IAE.0b013e318244536f

View details for Web of Science ID 000308672300023

View details for PubMedID 22511071
Infrared Imaging and Optical Coherence Tomography Reveal Early-Stage Astrocytic Hamartomas Not Detectable by Fundoscopy AMERICAN JOURNAL OF OPHTHALMOLOGY Xu, L., Burke, T. R., Greenberg, J. P., Mahajan, V. B., Tsang, S. H. 2012; 153 (5): 883-889
Abstract

To describe and correlate the features of astrocytic hamartomas using multimodal imaging.Prospective, noncomparative, observational case series.This was a single-center study of 4 patients (8 eyes) with tuberous sclerosis complex. A complete ophthalmologic examination, fundus photography, fundus autofluorescence (FAF), infrared imaging, and spectral-domain optical coherence tomography (SD-OCT) were performed for each patient. Images from each modality were analyzed and compared.In 2 patients, infrared imaging and SD-OCT detected occult retinal astrocytic hamartomas that were not observed on clinical examination or color fundus photography. FAF demonstrated the greatest contrast between lesions and surrounding retina but failed to identify 1 occult lesion that was detected with infrared imaging and SD-OCT. SD-OCT revealed lesions arising from the retinal nerve fiber layer with overlying vitreous adhesions, hyperreflective dots, and optically empty spaces at all depths of the tumor. Hamartomas were hyporeflective on infrared imaging and hypoautofluorescent on FAF. FAF of some lesions demonstrated hyperautofluorescent spots.Infrared imaging and SD-OCT aid in the detection of astrocytic hamartomas that are not visible on clinical examination or color fundus photography. SD-OCT enhances visualization of structural details. FAF is a useful adjunctive test to obtain greater contrast between lesions and surrounding retina. The ability to monitor structural changes over time in astrocytic hamartomas using SD-OCT may be beneficial for monitoring the success of systemic chemotherapy in the treatment of various tuberous sclerosis tumors.

View details for DOI 10.1016/j.ajo.2011.10.033

View details for Web of Science ID 000303964800013

View details for PubMedID 22310082

View details for PubMedCentralID PMC3331884
23-Gauge Pediatric Vitrectomy Using Limbus-Based Trocar-Cannulas RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Kay, C. N., Quiram, P. H., Mahajan, V. B. 2012; 32 (5): 1023-1027
View details for DOI 10.1097/IAE.0b013e3182533d85

View details for Web of Science ID 000303502200023

View details for PubMedID 22466464
Re: Intraoperative choroidal detachment during 23-gauge vitrectomy. Retina (Philadelphia, Pa.) Tarantola, R. M., Mahajan, V. B. 2012; 32 (2): 411-?
View details for DOI 10.1097/IAE.0b013e3182398122

View details for PubMedID 22222759
Monozygotic twins with CAPN5 autosomal dominant neovascular inflammatory vitreoretinopathy. Clinical ophthalmology (Auckland, N.Z.) Rowell, H. A., Bassuk, A. G., Mahajan, V. B. 2012; 6: 2037-2044
Abstract

The purpose of this study was to describe the clinical findings in a set of monozygotic twins with autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV) over a 23-year period.A pair of female twins were examined between 26 and 49 years of age. The concordance and discordance of their clinical features were determined. The CAPN5 gene was sequenced using genomic DNA.Both twins of an affected father demonstrated Stage I ADNIV with mild vitreous cells and a negative b-wave on electroretinography. Genetic analysis confirmed a guanine to thymine nucleotide (c.728G>T, pArg243Leu) mutation in the CAPN5 gene. Over the course of 23 years, each twin progressed to stage III disease, showing posterior uveitis, cystoid macular edema, intraocular fibrosis, early retinal neovascularization, retinal degeneration, and cataract. Disease progression varied moderately between each twin and was asymmetrical between eyes. Twin A had 20/70 and 20/125 in the right and left eye, respectively, and underwent vitrectomy surgery and intravitreal injections with bevacizumab for recurrent cystoid macular edema. Twin B maintained 20/20 and 20/40 in the right and left eye, respectively without intervention.There was asymmetry between the eyes and some discordance in the rate of disease progression in these monozygotic twins with ADNIV. The overall high disease concordance suggests genetic factors play a major role in clinical manifestations in CAPN5 vitreoretinopathy.

View details for DOI 10.2147/OPTH.S40086

View details for PubMedID 23271883

View details for PubMedCentralID PMC3526908
Mouse eye enucleation for remote high-throughput phenotyping. Journal of visualized experiments : JoVE Mahajan, V. B., Skeie, J. M., Assefnia, A. H., Mahajan, M., Tsang, S. H. 2011
Abstract

The mouse eye is an important genetic model for the translational study of human ophthalmic disease. Blinding diseases in humans, such as macular degeneration, photoreceptor degeneration, cataract, glaucoma, retinoblastoma, and diabetic retinopathy have been recapitulated in transgenic mice.(1-5) Most transgenic and knockout mice have been generated by laboratories to study non-ophthalmic diseases, but genetic conservation between organ systems suggests that many of the same genes may also play a role in ocular development and disease. Hence, these mice represent an important resource for discovering new genotype-phenotype correlations in the eye. Because these mice are scattered across the globe, it is difficult to acquire, maintain, and phenotype them in an efficient, cost-effective manner. Thus, most high-throughput ophthalmic phenotyping screens are restricted to a few locations that require on-site, ophthalmic expertise to examine eyes in live mice. (6-9) An alternative approach developed by our laboratory is a method for remote tissue-acquisition that can be used in large or small-scale surveys of transgenic mouse eyes. Standardized procedures for video-based surgical skill transfer, tissue fixation, and shipping allow any lab to collect whole eyes from mutant animals and send them for molecular and morphological phenotyping. In this video article, we present techniques to enucleate and transfer both unfixed and perfusion fixed mouse eyes for remote phenotyping analyses.

View details for DOI 10.3791/3184

View details for PubMedID 22126835

View details for PubMedCentralID PMC3308585
Automated Discovery and Quantification of Image-Based Complex Phenotypes: A Twin Study of Drusen Phenotypes in Age-Related Macular Degeneration INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE Quellec, G., Russell, S. R., Seddon, J. M., Reynolds, R., Scheetz, T., Mahajan, V. B., Stone, E. M., Abramoff, M. D. 2011; 52 (12): 9195-9206
Abstract

Determining the relationships between phenotype and genotype of many disorders can improve clinical diagnoses, identify disease mechanisms, and enhance therapy. Most genetic disorders result from interaction of many genes that obscure the discovery of such relationships. The hypothesis for this study was that image analysis has the potential to enable formalized discovery of new visible phenotypes. It was tested in twins affected with age-related macular degeneration (AMD).Fundus images from 43 monozygotic (MZ) and 32 dizygotic (DZ) twin pairs with AMD were examined. First, soft and hard drusen were segmented. Then newly defined phenotypes were identified by using drusen distribution statistics that significantly separate MZ from DZ twins. The ACE model was used to identify the contributions of additive genetic (A), common environmental (C), and nonshared environmental (E) effects on drusen distribution phenotypes.Four drusen distribution characteristics significantly separated MZ from DZ twin pairs. One encoded the quantity, and the remaining three encoded the spatial distribution of drusen, achieving a zygosity prediction accuracy of 76%, 74%, 68%, and 68%. Three of the four phenotypes had a 55% to 77% genetic effect in an AE model, and the fourth phenotype showed a nonshared environmental effect (E model).Computational discovery of genetically determined features can reveal quantifiable AMD phenotypes that are genetically determined without explicitly linking them to specific genes. In addition, it can identify phenotypes that appear to result predominantly from environmental exposure. The approach is rapid and unbiased, suitable for large datasets, and can be used to reveal unknown phenotype-genotype relationships.

View details for DOI 10.1167/iovs.10-6793

View details for Web of Science ID 000297631400055

View details for PubMedID 22039249

View details for PubMedCentralID PMC3302481
INTRAVITREAL BEVACIZUMAB FOR TREATMENT OF PROLIFERATIVE AND NONPROLIFERATIVE TYPE 2 IDIOPATHIC MACULAR TELANGIECTASIA RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Roller, A. B., Folk, J. C., Patel, N. M., Boldt, H. C., Russell, S. R., Abramoff, M. D., Mahajan, V. B. 2011; 31 (9): 1848-1855
Abstract

To determine the effect of treatment with intravitreal bevacizumab on retinal thickness and visual acuity in the nonproliferative and proliferative forms of Type 2 idiopathic macular telangiectasia.Retrospective chart review of clinic patients treated with bevacizumab for macular telangiectasia Type 2. Treatment was performed until no further changes were seen after repeated bevacizumab injections. All patients had Snellen visual acuity testing, fundus fluorescein angiography, and measurement of central macular thickness by optical coherence tomography at baseline. Visual acuity and central macular thickness were recorded at follow-up visits.Fourteen eyes of 10 patients were included. In 5 eyes with nonproliferative macular telangiectasia Type 2, average follow-up was 17 months (± 7 months), and no eye demonstrated improvement in visual acuity or decrease in central macular thickness at final follow-up compared with baseline. In 9 eyes with proliferative disease, follow-up averaged 17 months (± 9 months). At 6 weeks, central macular thickness decreased 63 μm (± 58 μm), and acuity improved 1.7 lines (± 2 lines). At final follow-up, central macular thickness decreased 48 μm (± 89 μm) and acuity improved 1.1 lines (± 3 lines). Subretinal neovascularization resolved in eight of nine eyes with proliferative disease after treatment.Bevacizumab did not improve acuity or reduce retinal thickness in nonproliferative macular telangiectasia Type 2 at final follow-up. In proliferative macular telangiectasia Type 2, bevacizumab caused involution of neovascularization and improved visual acuity.

View details for DOI 10.1097/IAE.0b013e31820d3feb

View details for Web of Science ID 000295318800016

View details for PubMedID 21610563
Uveitis Following Intravitreal Bevacizumab: A Non-Infectious Cluster OPHTHALMIC SURGERY LASERS & IMAGING Kay, C. N., Tarantola, R. M., Gehrs, K. M., Folk, J. C., Mahajan, V. B., Boldt, H. C., Syed, N. A., Russell, S. R. 2011; 42 (4): 292-296
Abstract

In this retrospective case series, the authors report seven cases of bevacizumab-related uveitis that occurred within a 4-month period.Seven eyes of six patients developed non-infectious uveitis following bevacizumab intravitreal injections in a cohort of 978 consecutive bevacizumab injections.The mean age of patients was 74.6 years (range: 26 to 92). All patients developed symptom onset within 1 day of injection. Shared signs and symptoms included corneal edema, anterior chamber and vitreous cell, conjunctival injection, ocular pain, and lack of hypopyon. In all patients, visual acuity returned to within one line of baseline acuity. All seven eyes had been previously injected with bevacizumab, with a mean number of antecedent injections of 6.1 (range: 3 to 12).A cluster of sterile bevacizumab-related uveitic reactions was described in this case series. Acute onset of symptoms, absence of hypopyon, a predominant anterior segment reaction, and prompt improvement on topical steroid therapy are useful clinical features distinguishing this uveitic syndrome from infectious endophthalmitis.

View details for DOI 10.3928/15428877-20110603-04

View details for Web of Science ID 000305342000004

View details for PubMedID 21800802
Sutureless Triplanar Sclerotomy for 23-Gauge Vitrectomy ARCHIVES OF OPHTHALMOLOGY Mahajan, V. B., Tarantola, R. M., Graff, J. M., Boldt, H. C., Abramoff, M. D., Russell, S. R., Folk, J. C. 2011; 129 (5): 585-590
Abstract

To describe and test the intraoperative integrity of triplanar sclerotomies.A prospective consecutive case series of 180 sclerotomies in 60 eyes was studied. After conjunctival dissection, triplanar transscleral wounds were created with a 23-gauge trocar using a standardized technique. At the conclusion of surgery, an air-fluid exchange was performed, and cannulas were removed. Then, unsutured scleral wound integrity was tested for permeability to vitreous, gas, and fluid by application of a cellulose sponge, observation of gas escape, and examination by the Seidel method, respectively. Postoperative intraocular pressure was recorded. Laboratory studies with fresh human donor globes were conducted to evaluate the histologic characteristics of triplanar sclerotomies.Unsutured triplanar wounds were closed to vitreous, gas, and fluid in 169 of 180 sclerotomies (93.9%). Eleven sclerotomies were open and showed positive results using only 1 testing method each. In these cases, vitreous was detected in 1 wound, gas escaped from 8 wounds, and Seidel test results were positive in 2 wounds. Complex retinal detachment repairs had a higher rate of wounds requiring suture placement. One patient had transient postoperative day 1 hypotony.It is possible to achieve high rates of unsutured wound closure with triplanar transscleral sclerotomies. No single method of evaluating wound leakage appeared to be the most sensitive. More complex cases requiring longer surgical times, more instrument passes, and increased wound manipulation resulted in higher rates of wound leakage. Leakage can be subtle, and surgeons should use multiple methods to adequately assess sclerotomy closure.

View details for Web of Science ID 000290437100007

View details for PubMedID 21555611
INTRAOPERATIVE CHOROIDAL DETACHMENT DURING 23-GAUGE VITRECTOMY RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Tarantola, R. M., Folk, J. C., Shah, S. S., Boldt, H. C., Abramoff, M. D., Russell, S. R., Mahajan, V. B. 2011; 31 (5): 893-901
Abstract

To review intraoperative choroidal detachments during 23-gauge vitrectomy and examine possible mechanism(s) involved.A retrospective consecutive case review of 23-gauge vitrectomies was performed. Main outcomes included choroidal detachment incidence, location, extent, relation to infusion cannula, and postoperative course. Laboratory study of human donor eyes was conducted by placing 23-gauge cannulas at various angles through the pars plana and injecting viscoelastic material after cannula retraction.Among 338 consecutive 23-gauge vitrectomy cases, 12 (3.55%) intraoperative choroidal detachments occurred. These included 6 (1.77%) serous detachments, 4 (1.18%) limited hemorrhagic detachments, and 1 case each of gas and silicone oil during an exchange. In four of six serous detachments and three of four hemorrhagic detachments, the detachment originated from the infusion cannula site. Intraoperative infusion cannula retraction (5 of 12 cases) and blockage (2 of 12 cases) caused transient hypotony. All cases of serous, hemorrhagic, and gas detachment resolved without intervention. Cannulas were placed at various angles to the sclera in human donor eyes. Choroidal detachments were produced after injecting viscoelastic material through obliquely placed cannulas after 1 mm of retraction.Infusion cannula retraction is an important mechanism and risk factor for the development of intraoperative choroidal detachment during 23-gauge vitrectomy. Precautions to prevent retraction and intraoperative repositioning may help avoid this complication.

View details for DOI 10.1097/IAE.0b013e3181f4429b

View details for Web of Science ID 000289933600011

View details for PubMedID 21273944
Evisceration of mouse vitreous and retina for proteomic analyses. Journal of visualized experiments : JoVE Skeie, J. M., Tsang, S. H., Mahajan, V. B. 2011
Abstract

While the mouse retina has emerged as an important genetic model for inherited retinal disease, the mouse vitreous remains to be explored. The vitreous is a highly aqueous extracellular matrix overlying the retina where intraocular as well as extraocular proteins accumulate during disease.(1-3) Abnormal interactions between vitreous and retina underlie several diseases such as retinal detachment, proliferative diabetic retinopathy, uveitis, and proliferative vitreoretinopathy.(1,4) The relative mouse vitreous volume is significantly smaller than the human vitreous (Figure 1), since the mouse lens occupies nearly 75% of its eye.(5) This has made biochemical studies of mouse vitreous challenging. In this video article, we present a technique to dissect and isolate the mouse vitreous from the retina, which will allow use of transgenic mouse models to more clearly define the role of this extracellular matrix in the development of vitreoretinal diseases.

View details for DOI 10.3791/2795

View details for PubMedID 21490583

View details for PubMedCentralID PMC3169273
Seroreactivity Against Aqueous-Soluble and Detergent-Soluble Retinal Proteins in Posterior Uveitis ARCHIVES OF OPHTHALMOLOGY Ko, A. C., Brinton, J. P., Mahajan, V. B., Zimmerman, B., Brinton, G. S., Stone, E. M., Folk, J. C., Mullins, R. F. 2011; 129 (4): 415-420
Abstract

To characterize the seroreactivity against retinal proteins in patients with posterior uveitis, retinal disease of noninflammatory origin, and healthy controls.Patients with posterior uveitis (n = 47), molecularly confirmed photoreceptor degenerations (n = 11), and healthy controls (n = 33) received dilated fundus examinations at the University of Iowa. Aqueous-soluble and detergent-soluble fractions of human retina were separated by gel electrophoresis and transferred to polyvinylidene fluoride membranes. Membranes were probed with patient serum samples to detect IgG, IgA, and IgM human antibodies that react with retinal antigens. The number of bands detected by Western blot was counted, and their molecular weights were determined.Antibodies recognizing retinal proteins were found in healthy controls, in patients with posterior uveitis, and in patients with molecularly confirmed heritable retinal degenerations. In healthy controls, 42% of individuals had circulating autoantibodies that recognized retinal proteins. Healthy controls had a low odds ratio of serum reactivity to soluble antigens (0.7; 95% confidence interval [CI], 0.4-1.2). Patients with inflammatory retinal diseases and inherited retinal diseases had 4.89 (95% CI, 2.25-10.64; P < .001) and 2.71 (95% CI, 1.19-6.16; P = .02) times more activity against soluble retinal antigens compared with controls.Healthy control patients exhibited a significantly higher level of background autoantibody activity against retinal proteins than previously reported. Antibody activity in healthy controls was primarily directed against membrane-bound retinal proteins, whereas in patients with pathologic retinal conditions, antibodies targeting nonmembrane-bound retinal proteins predominate.

View details for Web of Science ID 000289378900005

View details for PubMedID 21482867
Mutations in Prickle Orthologs Cause Seizures in Flies, Mice, and Humans AMERICAN JOURNAL OF HUMAN GENETICS Tao, H., Manak, J. R., Sowers, L., Mei, X., Kiyonari, H., Abe, T., Dandaleh, N. S., Yang, T., Wu, S., Chen, S., Fox, M. H., Gurnett, C., Montine, T., Bird, T., Shaffer, L. G., Rosenfeld, J. A., McConne, J., Madan-Khetarpal, S., Berry-Kravis, E., Griesbach, H., Saneto, R. P., Scott, M. P., Antic, D., Reed, J., Boland, R., Ehaideb, S. N., El-Shanti, H., Mahajan, V. B., Ferguson, P. J., Axelrod, J. D., Lehesjoki, A., Fritzsch, B., Slusarski, D. C., Wemmie, J., Ueno, N., Bassuk, A. G. 2011; 88 (2): 138-149
Abstract

Epilepsy is heritable, yet few causative gene mutations have been identified, and thus far no human epilepsy gene mutations have been found to produce seizures in invertebrates. Here we show that mutations in prickle genes are associated with seizures in humans, mice, and flies. We identified human epilepsy patients with heterozygous mutations in either PRICKLE1 or PRICKLE2. In overexpression assays in zebrafish, prickle mutations resulted in aberrant prickle function. A seizure phenotype was present in the Prickle1-null mutant mouse, two Prickle1 point mutant (missense and nonsense) mice, and a Prickle2-null mutant mouse. Drosophila with prickle mutations displayed seizures that were responsive to anti-epileptic medication, and homozygous mutant embryos showed neuronal defects. These results suggest that prickle mutations have caused seizures throughout evolution.

View details for DOI 10.1016/j.ajhg.2010.12.012

View details for Web of Science ID 000287684100002

View details for PubMedID 21276947

View details for PubMedCentralID PMC3035715
Dissection of human vitreous body elements for proteomic analysis. Journal of visualized experiments : JoVE Skeie, J. M., Mahajan, V. B. 2011
Abstract

The vitreous is an optically clear, collagenous extracellular matrix that fills the inside of the eye and overlies the retina. (1,2) Abnormal interactions between vitreous substructures and the retina underlie several vitreoretinal diseases, including retinal tear and detachment, macular pucker, macular hole, age-related macular degeneration, vitreomacular traction, proliferative vitreoretinopathy, proliferative diabetic retinopathy, and inherited vitreoretinopathies. (1,2) The molecular composition of the vitreous substructures is not known. Since the vitreous body is transparent with limited surgical access, it has been difficult to study its substructures at the molecular level. We developed a method to separate and preserve these tissues for proteomic and biochemical analysis. The dissection technique in this experimental video shows how to isolate vitreous base, anterior hyaloid, vitreous core, and vitreous cortex from postmortem human eyes. One-dimensional SDS-PAGE analyses of each vitreous component showed that our dissection technique resulted in four unique protein profiles corresponding to each substructure of the human vitreous body. Identification of differentially compartmentalized proteins will reveal candidate molecules underlying various vitreoretinal diseases.

View details for DOI 10.3791/2455

View details for PubMedID 21304469

View details for PubMedCentralID PMC3182653
BILATERAL INTRAVITREAL INJECTION OF ANTIVASCULAR ENDOTHELIAL GROWTH FACTOR THERAPY RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Mahajan, V. B., Elkins, K. A., Russell, S. R., Boldt, H. C., Gehrs, K. M., Weingeist, T. A., Stone, E. M., Abramoff, M. D., Liu, D., Folk, J. C. 2011; 31 (1): 31-35
Abstract

The purpose of this study was to review adverse events and patient preference after bilateral intravitreal injection of antibodies to vascular endothelial growth factor.A retrospective case-control study. Patients with exudative age-related macular degeneration who received intravitreal antivascular endothelial growth factor agent injections in both eyes (bilateral group) on the same day over a 23-month period were compared with patients who received injections in only 1 eye. The occurrence of endophthalmitis, cerebrovascular accident, myocardial infarction, death, patient discomfort, and patient preference was compared between the two groups.One hundred and two patients received an average of 4.43 bilateral injections (range 1-13). A case-control group of 102 patients received an average of 10.2 unilateral injections, (range 2-28). Bevacizumab was injected 45.5%, ranibizumab 45.5%, and a combination of bevacizumab and ranibizumab 9% of the time for bilateral injections. Bevacizumab was used 50.3% and ranubizumab 49.7% of the time in unilateral injections. The follow-up of both groups averaged 18.4 months (range 4.7-36.5 months). There were no cases of endophthalmitis or cerebrovascular accident in either group. There was a single case of myocardial infarction in each group. There were two deaths in the bilateral group and three deaths in the unilateral group. More than 90% strongly preferred bilateral injections to unilateral injections.Bilateral injections of antivascular endothelial growth factor agents on the same day did not increase the rate of adverse events and was preferred by the majority of patients.

View details for DOI 10.1097/IAE.0b013e3181ed8c80

View details for Web of Science ID 000285685100005

View details for PubMedID 21187731
Collagen XVIII Mutation in Knobloch Syndrome With Acute Lymphoblastic Leukemia AMERICAN JOURNAL OF MEDICAL GENETICS PART A Mahajan, V. B., Olney, A. H., Garrett, P., Chary, A., Dragan, E., Lerner, G., Murray, J., Bassuk, A. G. 2010; 152A (11): 2875-2879
Abstract

Knobloch syndrome (KNO) is caused by mutations in the collagen XVIII gene (COL18A1) and patients develop encephalocele and vitreoretinal degeneration. Here, we report an El Salvadorian family where two sisters showed features of KNO. One of the siblings also developed acute lymphoblastic leukemia. DNA sequencing of COL18A1 revealed a homozygous, 2-bp deletion (c3514-3515delCT) in exon 41, which leads to abnormal collagen XVIII and deficiency of its proteolytic cleavage product endostatin. KNO patients with mutations in COL18A1 may be at risk for endostatin-related conditions including malignancy.

View details for DOI 10.1002/ajmg.a.33621

View details for Web of Science ID 000284005700034

View details for PubMedID 20799329

View details for PubMedCentralID PMC2965270
Patients With an Acute Zonal Occult Outer Retinopathy-like Illness Rapidly Improve With Valacyclovir Treatment AMERICAN JOURNAL OF OPHTHALMOLOGY Mahajan, V. B., Stone, E. M. 2010; 150 (4): 511-518
Abstract

To describe 3 cases of an acute zonal occult outer retinopathy-like illness responsive to valacyclovir hydrochloride.Retrospective, interventional case series.Three patients were treated with valacyclovir and monitored by clinical examination, Goldmann visual field testing, and electroretinography.Patients with an acute zonal occult outer retinopathy-like illness presented following progressive vision loss. This course was immediately reversed by treatment with oral valacyclovir, and visual acuity and visual field improved significantly at 1 week and 1 month. Patients remained stable without treatment during a follow-up period ranging from 1 to 3 years.Some conditions with features of acute zonal occult outer retinopathy may be attributable to a subacute herpetic viral infection that is responsive to oral antiviral medication.

View details for DOI 10.1016/j.ajo.2010.05.024

View details for Web of Science ID 000282867500012

View details for PubMedID 20691421

View details for PubMedCentralID PMC4669885
INTRAVITREAL BEVACIZUMAB DURING PREGNANCY RETINA-THE JOURNAL OF RETINAL AND VITREOUS DISEASES Tarantola, R. M., Folk, J. C., Boldt, H. C., Mahajan, V. B. 2010; 30 (9): 1405-1411
Abstract

To report the clinical course of four women treated with intravitreal bevacizumab during pregnancy.Observational case series.Four pregnant women were treated with intravitreal bevacizumab for choroidal neovascularization (CNV) because of presumed ocular histoplasmosis syndrome punctate inner choroidopathy, or sarcoid uveitis. Patients received a mean of 2.6 ± 2.3 injections (range, 1-6 injections) while pregnant. One patient was treated with five additional injections while breastfeeding. The mean follow-up duration after the most recent injection was 14 ± 2.9 months (range, 11-18 months). Snellen visual acuity improved in all 4 patients with a mean of 5.75 ± 2.2 lines (range, 3-8 lines). At the most recent follow-up, all patients had involuted CNV that did not require additional treatment. All patients had an uneventful prenatal course and delivered healthy full-term infants. All children have remained healthy, exhibiting normal development and growth during infancy.Offering pregnant patients intravitreal bevacizumab therapy during pregnancy for off-label ocular indications can result in significant visual improvement. No adverse events related to treatment occurred in any patient included in this study. Additional studies with more patients and longer follow-up duration are required to identify any risks associated with treatment.

View details for DOI 10.1097/IAE.0b013e3181f57d58

View details for Web of Science ID 000282561800008

View details for PubMedID 20924262
Effects of Vitrectomy on Age-Related Macular Degeneration OPHTHALMOLOGY Roller, A. B., Mahajan, V. B., Boldt, H. C., Abramoff, M. D., Russell, S. R., Folk, J. C. 2010; 117 (7): 1381-1386
Abstract

To determine whether vitrectomy alters the long-term progression of age-related macular degeneration (AMD).Retrospective case-control study.Forty-four eyes of 22 patients with AMD who underwent vitrectomy in 1 eye were included in the study. The progression of AMD at follow-up in the 22 eyes that underwent vitrectomy was compared with the 22 fellow, nonvitrectomized eyes.The charts and photographs of subjects with Age-Related Eye Disease Study category 3 AMD in both eyes who previously underwent vitrectomy surgery for an epiretinal membrane or macular hole were reviewed. Subjects were excluded if they had had a vitrectomy in both eyes, had <2 years of follow-up, had previous choroidal neovascularization (CNV), retinal detachment, diabetic retinopathy, angioid streaks, high myopia, vascular occlusions, or extensive macular scarring in either eye, or insufficient hospital records or photographs to determine the extent of AMD. Clinical notes throughout the follow-up interval were reviewed. Two vitreoretinal specialists independently graded pre- and postvitrectomy fundus photographs of all eyes in a masked fashion.The development or progression of geographic atrophy of the retinal pigment epithelium and the development of CNV.Twenty-two patients were included. The average follow up interval was 5.5 years (range, 2-15). Choroidal neovascularization developed in 5 control eyes and in 2 vitrectomized eyes, and atrophy developed in 7 control and 4 vitrectomized eyes. The difference between vitrectomized eyes and fellow eyes for the combined end points of RPE geographic atrophy or CNV was significant (P = 0.02).In this pilot study, we did not detect that vitrectomy increased the progression of AMD. In fact, it was associated with a reduced progression to geographic atrophy or CNV. Additional studies are needed to confirm or refute this association.The authors have no proprietary or commercial interest in any of the materials discussed in this article.

View details for DOI 10.1016/j.ophtha.2009.11.007

View details for Web of Science ID 000279436200015

View details for PubMedID 20176401
T-cell infiltration in autosomal dominant neovascular inflammatory vitreoretinopathy MOLECULAR VISION Mahajan, V. B., Vallone, J. G., Lin, J. H., Mullins, R. F., Ko, A. C., Folk, J. C., Stone, E. M. 2010; 16 (114-15): 1034-1040
Abstract

Autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV) is a familial blinding disease of unknown pathophysiology. The eyes and sera from patients with ADNIV were studied to understand the immune response in this condition.The clinical case of an ADNIV patient was reviewed. Eye specimens from two donors with ADNIV were examined with a panel of standard histopathological stains and immunohistochemical markers. These findings were compared to specimens of noninflammatory eye disease. Sera from twelve patients were also tested against retinal protein western blots for the presence of autoretinal antibodies.Each of the ADNIV and control eyes showed degenerative features of phthisis bulbi. Immunohistological stains revealed a supraciliary T-cell infiltrate in ADNIV eyes composed of cluster of differentiation-4 (CD4) positive and cluster of differentiation-8 (CD8)-positive cells. No immunoglobulin or B cells were detected in these eyes. Inflammatory cells were absent from the control eyes. No specific autoretinal antibodies were detected in ADNIV sera.Aberrant T-cell-mediated processes may underlie ADNIV, and therapeutics directed at T cells may better manage inflammation in these patients. Genes related to T-cell function are high priority screening candidates.

View details for Web of Science ID 000279676800001

View details for PubMedID 20596252

View details for PubMedCentralID PMC2893052
Automated Early Detection of Diabetic Retinopathy OPHTHALMOLOGY Abramoff, M. D., Reinhardt, J. M., Russell, S. R., Folk, J. C., Mahajan, V. B., Niemeijer, M., Quellec, G. 2010; 117 (6): 1147-1154
Abstract

To compare the performance of automated diabetic retinopathy (DR) detection, using the algorithm that won the 2009 Retinopathy Online Challenge Competition in 2009, the Challenge2009, against that of the one currently used in EyeCheck, a large computer-aided early DR detection project.Evaluation of diagnostic test or technology.Fundus photographic sets, consisting of 2 fundus images from each eye, were evaluated from 16670 patient visits of 16,670 people with diabetes who had not previously been diagnosed with DR.The fundus photographic set from each visit was analyzed by a single retinal expert; 793 of the 16,670 sets were classified as containing more than minimal DR (threshold for referral). The outcomes of the 2 algorithmic detectors were applied separately to the dataset and were compared by standard statistical measures.The area under the receiver operating characteristic curve (AUC), a measure of the sensitivity and specificity of DR detection.Agreement was high, and examination results indicating more than minimal DR were detected with an AUC of 0.839 by the EyeCheck algorithm and an AUC of 0.821 for the Challenge2009 algorithm, a statistically nonsignificant difference (z-score, 1.91). If either of the algorithms detected DR in combination, the AUC for detection was 0.86, the same as the theoretically expected maximum. At 90% sensitivity, the specificity of the EyeCheck algorithm was 47.7% and that of the Challenge2009 algorithm was 43.6%.Diabetic retinopathy detection algorithms seem to be maturing, and further improvements in detection performance cannot be differentiated from best clinical practices, because the performance of competitive algorithm development now has reached the human intrareader variability limit. Additional validation studies on larger, well-defined, but more diverse populations of patients with diabetes are needed urgently, anticipating cost-effective early detection of DR in millions of people with diabetes to triage those patients who need further care at a time when they have early rather than advanced DR.

View details for DOI 10.1016/j.ophtha.2010.03.046

View details for Web of Science ID 000278224400009

View details for PubMedID 20399502

View details for PubMedCentralID PMC2881172
Erythropoetin receptor expression in the human diabetic retina. BMC research notes Shah, S. S., Tsang, S. H., Mahajan, V. B. 2009; 2: 234-?
Abstract

Recent evidence suggests erythropoietin (EPO) and the erythropoietin receptor (EPOR) may play a direct role in the pathogenesis of diabetic retinopathy. Better characterization of the EPO-EPOR signaling system in the ischemic retina may offer a new therapeutic modality for ischemic ophthalmic diseases. This study was performed to identify EPOR mRNA expression in the human diabetic eye.EPOR antisense RNA probes were validated on human pancreas tissue. In the normal eye, EPOR was expressed in the retinal ganglion cell layer. Minimal expression was observed in the inner and outer nuclear layer. Under conditions of diabetic retinopathy, EPOR expression shifted to photoreceptor cells. Increased expression was also observed in the peripheral retina.EPOR expression may be a biomarker or contribute to disease mechanisms in diabetic retinopathy.

View details for DOI 10.1186/1756-0500-2-234

View details for PubMedID 19930719

View details for PubMedCentralID PMC2785834
A New Macular Dystrophy With Anomalous Vascular Development, Pigment Spots, Cystic Spaces, and Neovascularization ARCHIVES OF OPHTHALMOLOGY Mahajan, V. B., Russell, S. R., Stone, E. M. 2009; 127 (11): 1449-1457
Abstract

To clinically phenotype an inherited macular dystrophy with peculiar intraretinal pigment spots, cysts, and hemorrhage in a 24-year-old female proband and her family.Extended family members of the proband underwent dilated fundus examination, optical coherence tomography, and, in selected cases, fluorescein angiography and electroretinography.Seventeen family members, representing 3 generations and ranging in age from 5 to 64 years, were clinically examined. Visual acuities ranged from 20/20 to 20/200. Amblyopia and strabismus were frequently present in affected individuals. Consistent with an autosomal dominant pattern of inheritance, 7 family members had multiple central macular cystic spaces and flat, round, densely pigmented spots within the retina. There were right-angle vessels and telangiectasis in the central macula. Two subjects showed evidence of active macular neovascularization with leakage on fluorescein angiography at ages 7 and 24 years, which was responsive to either focal laser or a single injection of bevacizumab. In those cases examined, multifocal electroretinography showed a diminished foveal response.This spotted cystic neovascular macular dystrophy appears to represent a new autosomal dominant retinal condition. Because these patients are at risk for choroidal neovascularization, identification of the responsible gene may provide insight into the mechanisms of pathological neovascularization.

View details for Web of Science ID 000271583700006

View details for PubMedID 19901210
Management of Sympathetic Ophthalmia with the Fluocinolone Acetonide Implant OPHTHALMOLOGY Mahajan, V. B., Gehrs, K. M., Goldstein, D. A., Fischer, D. H., Lopez, J. S., Folk, J. C. 2009; 116 (3): 552-557
Abstract

We examined whether implantation of the fluocinolone acetonide (Retisert) implant achieved control of inflammation and a reduced need for oral corticosteroids or immunosuppressives in patients with sympathetic ophthalmia (SO).Retrospective, noncomparative case series.Eight patients with active SO.The results of fluocinolone acetonide implantation in 8 patients with active SO were studied with a follow-up period of 6 months to 2 years.Presence or absence of intraocular inflammation, visual acuity, intraocular pressure, need for further surgery, and the need for additional use of oral or locally injected corticosteroids and/or immunosuppressives.All patients demonstrated a significant reduction in the systemic medication required to maintain control of inflammation. Two patients had recurrent inflammatory episodes requiring the resumption of an oral immunosuppressive. Vision improved or was stabilized in all 8 patients.The fluocinolone acetonide implant provides inflammatory control and reduces the dependence on systemic immunosuppression in patients with SO.

View details for DOI 10.1016/j.ophtha.2008.10.024

View details for Web of Science ID 000264005400028

View details for PubMedID 19147232
Estrogen receptor alpha and matrix metalloproteinase 2 polymorphisms and age-related maculopathy in older women AMERICAN JOURNAL OF EPIDEMIOLOGY Seitzman, R. L., Mahajan, V. B., Mangione, C., Cauley, J. A., Ensrud, K., Stone, K. L., Cummings, S. R., Hochberg, M. C., Hillier, T., Sinsheimer, J. S., Yu, F., Coleman, A. L. 2008; 167 (10): 1217-1225
Abstract

In this study, the authors sought to determine whether single nucleotide polymorphisms in the estrogen receptor alpha (ESR1) and matrix metalloproteinase 2 (MMP2) genes are associated with age-related maculopathy (ARM) in older women. Subjects comprised a random sample of Caucasian women aged > or =74 years participating in the Study of Osteoporotic Fractures year 10 follow-up (n = 906) in 1997-1998. Fundus photographs were graded for ARM using a modification of the Wisconsin Age-Related Maculopathy Grading System. The prevalences of early ARM and late ARM were 46% and 4%, respectively. The MMP2 rs2287074 single nucleotide polymorphism (G-->A) was associated with ARM. The A allele was present in 47%, 43%, and 30% of subjects with no, early, and late ARM, respectively (p = 0.01), and was associated with lower odds of any ARM (for AG vs. GG, odds ratio = 0.80, 95% confidence interval: 0.65, 0.99; for AA vs. GG, odds ratio = 0.64, 95% confidence interval: 0.42, 0.98). An interaction with use of postmenopausal hormone therapy was significant (p = 0.02). The MMP2 rs2287074 A allele may be associated with a lower likelihood of ARM in older Caucasian women, particularly those who have never used hormone therapy. The role of MMP2 rs2287074 in ARM should be further elucidated.

View details for DOI 10.1093/aje/kwn024

View details for Web of Science ID 000255756100010

View details for PubMedID 18359774
Tissue-specific gene expression of head and neck squamous cell carcinoma in vivo by complementary DNA microarray analysis ARCHIVES OF OTOLARYNGOLOGY-HEAD & NECK SURGERY Sok, J. C., Kuriakose, M. A., Mahajan, V. B., Pearlman, A. N., Delacure, M. D., Chen, F. A. 2003; 129 (7): 760-770
Abstract

To identify distinct gene expression profiles of human head and neck squamous cell carcinomas (HNSCCAs) using complementary DNA (cDNA) microarray analysis and to create a preliminary, comprehensive database of HNSCCA gene expression.Nine patients with histologically confirmed HNSCCAs, staged according to the American Joint Committee on Cancer, were enrolled. The HNSCCA tumor tissue and normal mucosal tissue were harvested at the time of surgery. A cDNA library was constructed from the paired fresh-frozen human surgical specimens of HNSCCAs and nonmalignant epithelial tissues. Biotinylated RNA was transcribed from the cDNA library and hybridized to high-density microarrays containing approximately 12 000 human genes. Altered gene expression of HNSCCAs was identified by comparison to corresponding normal mucosal tissues after a bayesian statistical analysis of variance. Results were analyzed using the gene database of the National Institutes of Health. Hierarchical clustering of the genomic data sets was determined by similarity metrics based on Pearson correlation.Hierarchical clustering analysis revealed that the gene expression profiles obtained from the nonselected panel of 12 000 genes could distinguish the tumors from nonmalignant tissues. Gene expression changes were reproducibly observed in 227 genes representing previously identified chemokines, tumor suppressors, differentiation markers, matrix molecules, membrane receptors, and transcription factors that correlated with neoplasia, including 46 previously uncharacterized genes. Moreover, significant expression of the collagen type XI alpha1 gene and a novel gene was reproducibly observed in all 9 tumors, whereas these genes were virtually undetectable in their corresponding, adjacent nonmalignant tissues.Complementary DNA microarray analysis of human HNSCCAs has produced a preliminary, comprehensive database of tumor-specific gene expression profiles and provided important insights into modeling gene expression changes implicated in carcinogenesis. A large-scale analysis of gene expression carries the future potential of identifying sensitive molecular markers for early tumor detection, prognosis, and novel targets for interceptive therapeutics.

View details for Web of Science ID 000184104400014

View details for PubMedID 12874079
Microarray analysis of corneal fibroblast gene expression after interleukin-1 treatment INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE Mahajan, V. B., Wei, C., McDonnell, P. J. 2002; 43 (7): 2143-2151
Abstract

To identify changes in gene expression in human corneal fibroblasts after exposure to interleukin-1alpha.RNA was isolated from cultured human corneal fibroblasts after treatment with interleukin-1alpha and subjected to DNA microarray analysis. Changes in gene expression were determined by comparison with untreated cells in three independent experiments after a Bayesian statistical analysis of variance.Changes in gene expression were reproducibly observed in 165 genes representing previously identified and novel chemokines, matrix molecules, membrane receptors, angiogenic mediators, and transcription factors that correlated with pathophysiological responses to inflammation. Dramatic increases in gene expression were observed with exodus-1 (CCL20), MMP-12, and RhoA.DNA microarray analysis of the corneal fibroblast response to interleukin-1alpha provides important insight into modeling changes in gene expression and suggests novel therapeutic targets for the control of corneal inflammation.

View details for Web of Science ID 000176482300014

View details for PubMedID 12091409
Thrombin receptor signaling to cytoskeleton requires Hsp90 JOURNAL OF BIOLOGICAL CHEMISTRY Pai, K. S., Mahajan, V. B., Lau, A., CUNNINGHAM, D. D. 2001; 276 (35): 32642-32647
Abstract

Thrombin is a serine protease that evokes various cellular responses involved in injury and repair of the nervous system through the activation of protease-activated receptor-1 (PAR-1). Signals that modulate cell morphology precede most PAR-1 effects, but the initial signal transduction molecules are not known. Using the yeast two-hybrid system, we identified Hsp90, a chaperone with known signaling properties, as a binding partner of PAR-1. The interaction was confirmed by glutathione S-transferase pull-down, overlay, and co-immunoprecipitation assays. Morphological assays in mouse astrocytes were carried out to evaluate the importance of Hsp90 during cytoskeletal signaling. Reducing Hsp90 levels by antisense treatment or disruption of the Hsp90.PAR-1 complex by the Hsp90-specific drug geldanamycin attenuated thrombin-mediated astrocyte shape changes. Furthermore, overexpression of the PAR-1 cytoplasmic tail abrogated thrombin-induced cytoskeletal changes in neuronal cells. Treatment with geldanamycin specifically inhibited activation of RhoA without affecting thrombin-mediated intracellular calcium release, revealing the regulation of a distinct signaling pathway by Hsp90. Taken together, these studies demonstrate that Hsp90 may be essential for PAR-1-mediated signaling to the cytoskeleton.

View details for Web of Science ID 000170746000036

View details for PubMedID 11413145
Creatine kinase, an ATP-generating enzyme, is required for thrombin receptor signaling to the cytoskeleton PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Mahajan, V. B., Pai, K. S., Lau, A., CUNNINGHAM, D. D. 2000; 97 (22): 12062-12067
Abstract

Thrombin orchestrates cellular events after injury to the vascular system and extravasation of blood into surrounding tissues. The pathophysiological response to thrombin is mediated by protease-activated receptor-1 (PAR-1), a seven-transmembrane G protein-coupled receptor expressed in the nervous system that is identical to the thrombin receptor in platelets, fibroblasts, and endothelial cells. Once activated by thrombin, PAR-1 induces rapid and dramatic changes in cell morphology, notably the retraction of growth cones, axons, and dendrites in neurons and processes in astrocytes. The signal is conveyed by a series of localized ATP-dependent reactions directed to the actin cytoskeleton. How cells meet the dynamic and localized energy demands during signal transmission is unknown. Using the yeast two-hybrid system, we identified an interaction between PAR-1 cytoplasmic tail and the brain isoform of creatine kinase, a key ATP-generating enzyme that regulates ATP within subcellular compartments. The interaction was confirmed in vitro and in vivo. Reducing creatine kinase levels or its ATP-generating potential inhibited PAR-1-mediated cellular shape changes as well as a PAR-1 signaling pathway involving the activation of RhoA, a small G protein that relays signals to the cytoskeleton. Thrombin-stimulated intracellular calcium release was not affected. Our results suggest that creatine kinase is bound to PAR-1 where it may be poised to provide bursts of site-specific high-energy phosphate necessary for efficient receptor signal transduction during cytoskeletal reorganization.

View details for Web of Science ID 000090071000065

View details for PubMedID 11050237

View details for PubMedCentralID PMC17294
CONNECTIN - A HOMOPHILIC CELL-ADHESION MOLECULE EXPRESSED ON A SUBSET OF MUSCLES AND THE MOTONEURONS THAT INNERVATE THEM IN DROSOPHILA CELL Nose, A., Mahajan, V. B., Goodman, C. S. 1992; 70 (4): 553-567
Abstract

Each abdominal hemisegment in the Drosophila embryo contains a stereotyped array of 30 muscles, each specifically innervated by one or a few motoneurons. We screened 11,000 enhancer trap lines, isolated several expressing beta-galactosidase in small subsets of muscle fibers prior to innervation, and identified two of these as inserts in connectin and Toll, members of the leucine-rich repeat gene family. Connectin contains a signal sequence, ten leucine-rich repeats, and a putative phosphatidylinositol membrane linkage; in S2 cells, connectin can mediate homophilic cell adhesion. Connectin is expressed on the surface of eight muscles, the motoneurons that innervate them, and several glial cells along the pathways leading to them. During synapse formation, the protein localizes to synaptic sites; afterward, it largely disappears. Thus, connectin is a novel cell adhesion molecule whose expression suggests a role in target recognition.

View details for Web of Science ID A1992JJ88600006

View details for PubMedID 1505024
ACUTE POSTTRANSFUSION FULMINANT HEPATITIS-C VIRUS-INFECTION - A CASE-REPORT GASTROENTEROLOGIA JAPONICA Woodfield, D. G. 1991; 26: 221-223
Abstract

A 21 year old woman developed acute fulminant hepatitis following a transfusion of hepatitis C virus (HCV) infected blood. The patient received a liver transplant and is now well and HCV antibody negative.

View details for Web of Science ID A1991FX75700054

View details for PubMedID 1909271
SOLUBILIZATION OF THE PLASMIN RECEPTOR FROM HUMAN CARCINOMA-CELLS BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS Burtin, P., Fondaneche, M. C. 1990; 170 (2): 748-754
Abstract

We could solubilize the plasmin receptor from the human colonic tumor cell line SW1116, using various detergents. Among these, Triton X100 and Sodium dodecyl sulfate were the most efficient. The solubilized receptor retained its plasmin binding activity, as judged by dot blot tests. After electrophoresis of the solubilisate in SDS polyacrylamide gels and transfer to nitrocellulose, specific plasmin fixation was still observed on autoradiograms as a sharp band in the 50K area. This band was not altered by treatment of the solubilisate with beta-mercaptoethanol.

View details for Web of Science ID A1990DR88200049

View details for PubMedID 2166506

Associate Professor of Ophthalmology at the Stanford University Medical Center

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