CRISPR-based genome editing in primary human pancreatic islet cells.
2021; 12 (1): 2397
Gene targeting studies in primary human islets could advance our understanding of mechanisms driving diabetes pathogenesis. Here, we demonstrate successful genome editing in primary human islets using clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein 9 (Cas9). CRISPR-based targeting efficiently mutated protein-coding exons, resulting in acute loss of islet beta-cell regulators, like the transcription factor PDX1 and the KATP channel subunit KIR6.2, accompanied by impaired beta-cell regulation and function. CRISPR targeting of non-coding DNA harboring type 2 diabetes (T2D) risk variants revealed changes in ABCC8, SIX2 and SIX3 expression, and impaired beta-cell function, thereby linking regulatory elements in these target genes to T2D genetic susceptibility. Advances here establish a paradigm for genetic studies in human islet cells, and reveal regulatory and genetic mechanisms linking non-coding variants to human diabetes risk.
View details for DOI 10.1038/s41467-021-22651-w
View details for PubMedID 33893274
SIX2 and SIX3 coordinately regulate functional maturity and fate of human pancreatic β cells.
Genes & development
The physiological functions of many vital tissues and organs continue to mature after birth, but the genetic mechanisms governing this postnatal maturation remain an unsolved mystery. Human pancreatic β cells produce and secrete insulin in response to physiological cues like glucose, and these hallmark functions improve in the years after birth. This coincides with expression of the transcription factors SIX2 and SIX3, whose functions in native human β cells remain unknown. Here, we show that shRNA-mediated SIX2 or SIX3 suppression in human pancreatic adult islets impairs insulin secretion. However, transcriptome studies revealed that SIX2 and SIX3 regulate distinct targets. Loss of SIX2 markedly impaired expression of genes governing β-cell insulin processing and output, glucose sensing, and electrophysiology, while SIX3 loss led to inappropriate expression of genes normally expressed in fetal β cells, adult α cells, and other non-β cells. Chromatin accessibility studies identified genes directly regulated by SIX2. Moreover, β cells from diabetic humans with impaired insulin secretion also had reduced SIX2 transcript levels. Revealing how SIX2 and SIX3 govern functional maturation and maintain developmental fate in native human β cells should advance β-cell replacement and other therapeutic strategies for diabetes.
View details for DOI 10.1101/gad.342378.120
View details for PubMedID 33446570
Aquagenic pruritus in an adolescent effectively managed with beta-alanine supplementation.
Aquagenic pruritus is a rare debilitating condition, which can be idiopathic, iatrogenic, or associated with systemic disease. In idiopathic cases, treatment can be challenging as options are limited and of variable efficacy. Here, we report the case of a teenage boy with refractory idiopathic aquagenic pruritus effectively managed with administration of beta-alanine supplementation, a treatment gaining traction in lay media but not yet reported in the medical literature. This report adds to the limited options published for treatment of idiopathic aquagenic pruritus in pediatric patients.
View details for DOI 10.1111/pde.14440
View details for PubMedID 33170524
The long dystrophin gene product Dp427 modulates retinal function and vascular morphology in response to age and retinal ischemia.
2019; 129: 104489
Mutations in dystrophin are the major cause of muscular dystrophies. Continuous muscular degeneration and late stage complications, including cardiomyopathy and respiratory insufficiency, dominate the clinical phenotype. Gene expression and regulation of the dystrophin gene outside of muscular tissue is far more complex. Multiple tissue-specific dystrophin gene products are widely expressed throughout the body, including the central nervous system and eye, predisposing affected patients to secondary complications in non-muscular tissues. In this study, we evaluated the impact of the full-length dystrophin gene product, Dp427, on retinal homeostasis and angiogenesis. Based on the clinical case of a Duchenne muscular dystrophy (DMD) patient who developed severe fibrovascular changes in the retina in response to hypoxic stress, we hypothesized that defects in Dp427 make the retina more susceptible to stresses such as ageing and ischemia. To further study this, a mouse strain lacking Dp427 expression (Mdx) was studied during retinal development, ageing and in the oxygen-induced retinopathy (OIR) model. While retinal vascular morphology was normal during development and ageing, retinal function measured by electroretinography (ERG) was slightly reduced in young adult Mdx mice and deteriorated with age. Mdx mice also had increased retinal neovascularization in response to OIR and more pronounced long-term deterioration in retinal function following OIR. Based on these results, we suggest that DMD patients with a mutation in Dp427 may experience disturbed retinal homeostasis with increasing age and therefore be prone to develop excessive retinal neovascular changes in response to hypoxic stress. DMD patients in late disease stages should, thus, be regularly examined to detect asymptomatic retinal abnormalities and prevent visual impairment.
View details for DOI 10.1016/j.neuint.2019.104489
View details for PubMedID 31199961