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Epstein-Barr Virus Latent Membrane Protein 1 Regulates Host B Cell MicroRNA-155 and Its Target FOXO3a via PI3K p110 alpha Activation FRONTIERS IN MICROBIOLOGY Hatton, O., Smith, M. M., Alexander, M., Mandell, M., Sherman, C., Stesney, M. W., Hui, S., Dohrn, G., Medrano, J., Ringwalt, K., Harris-Arnold, A., Maloney, E. M., Krams, S. M., Martinez, O. M. 2019; 10
View details for DOI 10.3389/fmicb.2019.02692

View details for Web of Science ID 000501847400001
Evaluating for Human Herpesvirus 6 in the Liver Explants of Children With Liver Failure of Unknown Etiology JOURNAL OF INFECTIOUS DISEASES Yang, C. H., Sahoo, M. K., Fitzpatrick, M., Lau, A. H., Pinsky, B. A., Martinez, O. M. 2019; 220 (3): 361?69
View details for DOI 10.1093/infdis/jiy644

View details for Web of Science ID 000477595700004
Biomarkers for PTLD diagnosis and therapies. Pediatric nephrology (Berlin, Germany) Martinez, O. M. 2019
Abstract

Post-transplant lymphoproliferative disorder (PTLD) represents a spectrum of lymphoproliferative disorders and is a serious complication of pediatric transplantation. The majority of PTLD are associated with Epstein Barr virus (EBV) and the characteristic EBV+ B cell lymphomas are the leading post-transplant malignancy in children. EBV+ PTLD remains a formidable issue in pediatric transplantation and is thought to result from impaired immunity to EBV as a result of immunosuppression. However, the key viral and immune factors that determine whether EBV+ PTLD develops remain unknown. Recently, there has been much interest in developing biomarkers in order to improve and achieve more personalized approaches, in the clinical diagnosis, management, and treatment of EBV+ PTLD. Here, we review the status of immune-, viral-, and B cell lymphoma-derived candidates for biomarkers of EBV+ PTLD.

View details for DOI 10.1007/s00467-019-04284-w

View details for PubMedID 31240394
Differential role of natural killer group 2D in recognition and cytotoxicity of hepatocyte-like cells derived from embryonic stem cells and induced pluripotent stem cells AMERICAN JOURNAL OF TRANSPLANTATION Cisneros, T., Dillard, D. W., Qu, X., Arredondo-Guerrero, J., Castro, M., Schaffert, S., Martin, R., Esquive, C. O., Krams, S. M., Martinez, O. M. 2019; 19 (6): 1652?62
View details for DOI 10.1111/ajt.15217

View details for Web of Science ID 000469275600010
Dual blockade of the PI3K/Akt/mTOR pathway inhibits posttransplant Epstein-Barr virus B cell lymphomas and promotes allograft survival AMERICAN JOURNAL OF TRANSPLANTATION Sang, A. X., McPherson, M. C., Ivison, G. T., Qu, X., Rigdon, J., Esquivel, C. O., Krams, S. M., Martinez, O. M. 2019; 19 (5): 1305?14
View details for DOI 10.1111/ajt.15216

View details for Web of Science ID 000471342300010
Genomic diversity of Epstein-Barr Virus in post-transplant lymphoproliferative disorder Martinez, O., Maloney, E., Busque, V., Hui, S., Esquivel, C. O., Krams, S. WILEY. 2019
View details for Web of Science ID 000485482200053
Characterization of genomic alterations in EBV plus PTLD Krams, S. M., Arvedson, M., Maloney, E., Balachandran, Y., McPherson, M., Boyd, S. D., Esquivel, C. O., Martinez, O. M. WILEY. 2019
View details for Web of Science ID 000485482200026
Differential role of natural killer group 2D in recognition and cytotoxicity of hepatocyte-like cells derived from embryonic stem cells and induced pluripotent stem cells. American journal of transplantation : official journal of the American Society of Transplantation and the American Society of Transplant Surgeons Cisneros, T., Dillard, D. W., Qu, X., Arredondo-Guerrero, J., Castro, M., Schaffert, S., Martin, R., Esquivel, C. O., Krams, S. M., Martinez, O. M. 2018
Abstract

Stem cell-based approaches have the potential to address the organ shortage in transplantation. Whereas both embryonic stem cells and induced pluripotent stem cells have been utilized as cellular sources for differentiation and lineage specification, their relative ability to be recognized by immune effector cells is unclear. We determined the expression of immune recognition molecules on hepatocyte-like cells (HLC) generated from murine embryonic stem cells and induced pluripotent stem cells, compared to adult hepatocytes, and we evaluated the impact on recognition by NK cells. We report that HLC lack MHC class I expression, and that embryonic stem cell derived-HLC have higher expression of the NK cell activating ligands Rae1, H60, and Mult1 than induced pluripotent stem cell-derived HLC and adult hepatocytes. Moreover, the lack of MHC class I render embryonic stem cell derived-HLC, and induced pluripotent stem cell derived-HLC, susceptible to killing by syngeneic and allogeneic NK cells. Both embryonic stem cell derived-HLC, and induced pluripotent stem cell derived-HLC, are killed by NK cells at higher levels than adult hepatocytes. Finally, we demonstrate that the NK cell activation receptor, NKG2D, plays a key role in NK cell cytotoxicity of embryonic stem cell derived-HLC, but not induced pluripotent stem cell-derived HLC. This article is protected by copyright. All rights reserved.

View details for PubMedID 30549427
Dual blockade of the PI3K/Akt/mTOR pathway inhibits post-transplant Epstein-Barr virus B cell lymphomas and promotes allograft survival. American journal of transplantation : official journal of the American Society of Transplantation and the American Society of Transplant Surgeons Sang, A. X., McPherson, M. C., Ivison, G. T., Qu, X., Rigdon, J., Esquivel, C. O., Krams, S. M., Martinez, O. M. 2018
Abstract

Post-transplant lymphoproliferative disorder (PTLD) is a serious complication of organ transplantation that often manifests as Epstein-Barr virus (EBV)-associated B cell lymphomas. Current treatments for PTLD have limited efficacy and can be associated with graft rejection or systemic toxicities. The mTOR inhibitor, rapamycin, suppresses tumor growth of EBV+ B cell lymphoma cells in vitro and in vivo; however, the efficacy is limited and clinical benefits of mTOR inhibitors for PTLD are variable. Here, we show constitutive activation of multiple nodes within the PI3K/Akt/mTOR pathway in EBV+ PTLD-derived cell lines. Inhibition of either PI3K or Akt, with specific inhibitors CAL-101 and MK-2206, respectively, diminished growth of EBV+ B cell lines from PTLD patients in a dose-dependent manner. Importantly, rapamycin combined with CAL-101 or MK-2206 had a synergistic effect in suppressing cell growth as determined by IC50 isobolographic analysis and Loewe indices. Moreover, these combinations were significantly more effective than rapamycin alone in inhibiting tumor xenograft growth in NOD-SCID mice. Finally, both CAL-101 and MK-2206 also prolonged survival of heterotopic cardiac allografts in C57BL/6 mice. Thus, combination therapy with rapamycin and a PI3K inhibitor, or an Akt inhibitor, can be an efficacious treatment for EBV-associated PTLD, while simultaneously promoting allograft survival. This article is protected by copyright. All rights reserved.

View details for PubMedID 30549430
Evaluating for human herpesvirus 6 in the liver explants of children with liver failure of unknown etiology. The Journal of infectious diseases Yang, C. H., Sahoo, M. K., Fitzpatrick, M., Lau, A. H., Pinsky, B. A., Martinez, O. M. 2018
Abstract

Background: Liver failure of unknown etiology (LFUE) has a transplant-free survival rate of less than 25%. Human herpesvirus 6 (HHV-6) may be associated with LFUE, but existing studies are limited by small sample size.Methods: We identified all children who underwent liver transplant for LFUE at a single quaternary children's hospital. 51/65 cases were able to be age-matched with controls (children who underwent liver transplant for metabolic liver disease). Quantitative polymerase chain reaction (PCR) for HHV-6 was performed on DNA from formalin-fixed paraffin-embedded liver explant tissue. Results: HHV-6 was detected in 34/51 cases (66.7%), and 19/51 controls (37.3%) (p=0.005). Average HHV-6 viral load was 213,207 copies/10 6 cells in positive cases (range: 7,293-1,102,030), and 38,115 copies/10 6 cells in positive controls (range: 1,382-122,375) (p=0.0008). HHV-6 was present significantly more often in cases compared to controls in patients less than 6 years in age; in particular, in patients less than 3 years in age, HHV-6 was present in 13/27 cases (48.1%), and 2/27 controls (7.4%) (p=0.0009).Conclusions: HHV-6 was detected in liver explants significantly more often and in higher quantities in children transplanted for LFUE compared to controls. This suggests HHV-6 should be evaluated for in young children who present with LFUE.

View details for PubMedID 30418598
Prospective Analysis of EBV plus PTLD in a Multi-Center Study of Pediatric Transplant Recipients Martinez, O. M., Krams, S. M., Lapasaran, M., Boyd, S. D., Bernstein, D., Twist, C., Weinberg, K., Gratzinger, D., Tan, B., Armstrong, B., Ikle, D., Brown, M., Robien, M., Esquivel, C. O. LIPPINCOTT WILLIAMS & WILKINS. 2018: S319
View details for Web of Science ID 000444541200511
Genomic Status of the Epstein Barr Virus and Virus-Associated PI3K/Akt/mTOR Pathway Dysregulation in Post-Transplant Lymphoproliferative Disorder McPherson, M., Balachandran, Y., Boyd, S. D., Zimmermann, H., Trappe, R. U., Esquivel, C. O., Krams, S. M., Martinez, O. M. LIPPINCOTT WILLIAMS & WILKINS. 2018: S95
View details for Web of Science ID 000444541200158
Delineation of the Viral and Host Cell Genomic Alterations in EBV-positive PTLD Balachandran, Y., McPherson, M., Boyd, S. D., Esquivel, C. O., Krams, S., Martinez, O. M. LIPPINCOTT WILLIAMS & WILKINS. 2018: S319
View details for Web of Science ID 000444541200512
Identifying shared patterns in the T cell receptor repertoire specific to IE-1 CMV Lucia Perez, M., Rubelt, F., Luque, S., Krams, S. M., Esquivel, C. O., Bestard, O., Martinez, O. M. LIPPINCOTT WILLIAMS & WILKINS. 2018: S141
View details for Web of Science ID 000444541200231
Application of Mass Cytometry for Analysis of the Alloimmune Response in a Model of Vascularized Composite Allotransplantation Harden, J. T., Sang, A. X., Qu, X., Esquivel, C., Martinez, O. M., Krams, S. M. LIPPINCOTT WILLIAMS & WILKINS. 2018: S198
View details for Web of Science ID 000444541200317
Epstein-Barr Virus Genome Variation in Post-Transplant Lymphoproliferative Disorder Maloney, E., Busque, V. A., Hui, S., Krams, S. M., Esquivel, C. O., Martinez, O. M. LIPPINCOTT WILLIAMS & WILKINS. 2018: S95
View details for Web of Science ID 000444541200157
Elucidation of the miRNome in EBV-positive and EBV-negative PTLD Balachandran, Y., Zimmermann, H., Esquivel, C. O., Krams, S., Trappe, R. U., Martinez, O. M. LIPPINCOTT WILLIAMS & WILKINS. 2018: S96
View details for Web of Science ID 000444541200160
NK Cell Recognition of Peptides Encoded by EBV Latent Cycle Proteins Mbiribindi, B., Moreno, C., Esquivel, C. O., Martinez, O. M., Krams, S. M. LIPPINCOTT WILLIAMS & WILKINS. 2018: S283
View details for Web of Science ID 000444541200451
Inhibition of Multiple Nodes in the PI3K/Akt/mTOR Pathway Synergistically Suppresses Post-Transplant B Cell Lymphomas Sang, A., McPherson, M., Ivison, G., Qu, X., Rigdon, J., Esquivel, C., Krams, S., Martinez, O. WILEY. 2018: 20
View details for Web of Science ID 000419034500015
Evaluating for human herpesvirus-6 in children with liver failure of unknown etiology Yang, C., Sahoo, M., Lau, A. H., Pinsky, B., Martinez, O. M. WILEY. 2017: 658A?659A
View details for Web of Science ID 000412089801360
STRATEGY FOR ANALYSIS OF DONOR-SPECIFIC ANTIBODY REPERTOIRE: PAIRED IMMUNOGLOBULIN HEAVY AND LIGHT CHAIN ANALYSIS OF INDIVIDUAL MEMORY B CELLS Lucia, M., Gomez, A., Bestard, O., Robinson, W., Martinez, O. WILEY. 2017: 103
View details for Web of Science ID 000411688501033
The Immune Response to Epstein Barr Virus and Implications for Posttransplant Lymphoproliferative Disorder. Transplantation Martinez, O. M., Krams, S. M. 2017
Abstract

Posttransplant lymphoproliferative disorder (PTLD) is a serious complication in organ transplant recipients and is most often associated with the Epstein Barr virus (EBV). EBV is a common gammaherpes virus with tropism for B lymphocytes and infection in immunocompetent individuals is typically asymptomatic and benign. However, infection in immunocompromised or immunosuppressed individuals can result in malignant B cell lymphoproliferations such as PTLD. EBV+ PTLD can arise following primary EBV infection, or because of reactivation of a prior infection, and represents a leading malignancy in the transplant population. The incidence of EBV+ PTLD is variable depending on the organ transplanted and whether the recipient has preexisting immunity to EBV but can be as high as 20%. It is generally accepted that impaired immune function due to immunosuppression is a primary cause of EBV+ PTLD. In this overview, we review the EBV life cycle and discuss our current understanding of the immune response to EBV in healthy, immunocompetent individuals, in transplant recipients, and in PTLD patients. We review the strategies that EBV utilizes to subvert and evade host immunity and discuss the implications for the development of EBV+ PTLD.

View details for DOI 10.1097/TP.0000000000001767

View details for PubMedID 28376031
Absence of miR-182 Augments Cardiac Allograft Survival. Transplantation Wei, L., Kaul, V., Qu, X., Xiong, X., Lau, A. H., Iwai, N., Martinez, O. M., Krams, S. M. 2017; 101 (3): 524-530
Abstract

MicroRNAs (miRNAs) are small noncoding RNA molecules that regulate the posttranscriptional expression of target genes and are important regulators in immune responses. Previous studies demonstrated that the miRNA, miR-182 was significantly increased during allograft rejection. Further, the transcription factor Forkhead box (FOX) protein 1, (FOXO1) was shown to be a target of miR-182. The aim of this study is to further examine the role of miR-182 in alloimmune responses.Transplantation of BALB/c cardiac allografts was performed in C57BL/6, miR-182, B6.129S-H2 (MHC II and CD4 T cell-deficient) and B6.129S2-Tap1 (MHC I and CD8 T cell-deficient) mice, with or without CTLA-4Ig administration. T cell phenotype, FOXO1 protein levels and graft infiltrating lymphocytes were determined in C57BL/6 or miR-182 mice by flow cytometric analysis, Western blot, and immunohistochemistry, respectively.We now show that T cells, mainly CD4 are the main cellular source of miR-182 during allograft rejection. In the absence of miR-182, CTLA-4Ig treatment significantly increased allograft survival (31.5 days C57BL/6 vs 60 days miR-182; P < 0.01). Further, CTLA4-Ig treatment inhibits miR-182 expression, increases FOXO1 levels, and reduces the percentage of CD4CD44 T cells after transplantation. Fewer T cells infiltrate the cardiac allografts, and memory T cells are significantly decreased in allograft recipients deficient in miR-182 with CTLA4-Ig treatment (P < 0.01).Our findings suggest that miR-182 contributes to the T-cell responses to alloantigen especially under costimulation blockade. Therapeutics that target specific miRNAs may prove beneficial in transplantation.

View details for DOI 10.1097/TP.0000000000001345

View details for PubMedID 27893611

View details for PubMedCentralID PMC5315626
Dynamics of Viral and Host Immune Cell MicroRNA Expression during Acute Infectious Mononucleosis. Frontiers in microbiology Kaul, V., Weinberg, K. I., Boyd, S. D., Bernstein, D., Esquivel, C. O., Martinez, O. M., Krams, S. M. 2017; 8: 2666
Abstract

Epstein-Barr virus (EBV) is the etiological agent of acute infectious mononucleosis (IM). Since acute IM is a self-resolving disease with most patients regaining health in 1-3 weeks there have been few studies examining molecular signatures in early acute stages of the disease. MicroRNAs (miRNAs) have been shown, however, to influence immune cell function and consequently the generation of antibody responses in IM. In this study, we performed a comprehensive analysis of differentially expressed miRNAs in early stage uncomplicated acute IM. miRNAs were profiled from patient peripheral blood obtained at the time of IM diagnosis and at subsequent time points, and pathway analysis performed to identify important immune and cell signaling pathways. We identified 215 differentially regulated miRNAs at the most acute stage of infection when the patients initially sought medical help. The number of differentially expressed miRNAs decreased to 148 and 68 at 1 and 2 months post-primary infection, with no significantly changed miRNAs identified at 7 months post-infection. Interferon signaling, T and B cell signaling and antigen presentation were the top pathways influenced by the miRNAs associated with IM. Thus, a dynamic and regulated expression profile of miRNA accompanies the early acute immune response, and resolution of infection, in IM.

View details for PubMedID 29379474

View details for PubMedCentralID PMC5775229
Identifying specificity groups in the T cell receptor repertoire. Nature Glanville, J., Huang, H., Nau, A., Hatton, O., Wagar, L. E., Rubelt, F., Ji, X., Han, A., Krams, S. M., Pettus, C., Haas, N., Arlehamn, C. S., Sette, A., Boyd, S. D., Scriba, T. J., Martinez, O. M., Davis, M. M. 2017
Abstract

T cell receptor (TCR) sequences are very diverse, with many more possible sequence combinations than T cells in any one individual. Here we define the minimal requirements for TCR antigen specificity, through an analysis of TCR sequences using a panel of peptide and major histocompatibility complex (pMHC)-tetramer-sorted cells and structural data. From this analysis we developed an algorithm that we term GLIPH (grouping of lymphocyte interactions by paratope hotspots) to cluster TCRs with a high probability of sharing specificity owing to both conserved motifs and global similarity of complementarity-determining region 3 (CDR3) sequences. We show that GLIPH can reliably group TCRs of common specificity from different donors, and that conserved CDR3 motifs help to define the TCR clusters that are often contact points with the antigenic peptides. As an independent validation, we analysed 5,711 TCR? chain sequences from reactive CD4 T cells from 22 individuals with latent Mycobacterium tuberculosis infection. We found 141 TCR specificity groups, including 16 distinct groups containing TCRs from multiple individuals. These TCR groups typically shared HLA alleles, allowing prediction of the likely HLA restriction, and a large number of M. tuberculosis T cell epitopes enabled us to identify pMHC ligands for all five of the groups tested. Mutagenesis and de novo TCR design confirmed that the GLIPH-identified motifs were critical and sufficient for shared-antigen recognition. Thus the GLIPH algorithm can analyse large numbers of TCR sequences and define TCR specificity groups shared by TCRs and individuals, which should greatly accelerate the analysis of T cell responses and expedite the identification of specific ligands.

View details for PubMedID 28636589
NKG2A-Expressing Natural Killer Cells Dominate the Response to Autologous Lymphoblastoid Cells Infected with Epstein-Barr Virus FRONTIERS IN IMMUNOLOGY Hatton, O., Strauss-Albee, D. M., Zhao, N. Q., Haggadone, M. D., Pelpola, J. S., Krams, S. M., Martinez, O. M., Blish, C. A. 2016; 7
Abstract

Epstein-Barr virus (EBV) is a human ?-herpesvirus that establishes latency and lifelong infection in host B cells while achieving a balance with the host immune response. When the immune system is perturbed through immunosuppression or immunodeficiency, however, these latently infected B cells can give rise to aggressive B cell lymphomas. Natural killer (NK) cells are regarded as critical in the early immune response to viral infection, but their role in controlling expansion of infected B cells is not understood. Here, we report that NK cells from healthy human donors display increased killing of autologous B lymphoblastoid cell lines (LCLs) harboring latent EBV compared to primary B cells. Coculture of NK cells with autologous EBV(+) LCL identifies an NK cell population that produces IFN? and mobilizes the cytotoxic granule protein CD107a. Multi-parameter flow cytometry and Boolean analysis reveal that these functional cells are enriched for expression of the NK cell receptor NKG2A. Further, NKG2A(+) NK cells more efficiently lyse autologous LCL than do NKG2A(-) NK cells. More specifically, NKG2A(+)2B4(+)CD16(-)CD57(-)NKG2C(-)NKG2D(+) cells constitute the predominant NK cell population that responds to latently infected autologous EBV(+) B cells. Thus, a subset of NK cells is enhanced for the ability to recognize and eliminate autologous, EBV-infected transformed cells, laying the groundwork for harnessing this subset for therapeutic use in EBV(+) malignancies.

View details for DOI 10.3389/fimmu.2016.00607

View details for Web of Science ID 000389857900001

View details for PubMedID 28018364

View details for PubMedCentralID PMC5156658
Epstein-Barr virus BARF1-induced NF kappa B/miR-146a/SMAD4 alterations in stomach cancer cells ONCOTARGET Kim, D. H., Chang, M. S., Yoon, C. J., Middeldorp, J. M., Martinez, O. M., Byeon, S., Rha, S. Y., Kim, S. H., Kim, Y. S., Woo, J. H. 2016; 7 (50): 82213-82227
Abstract

Epstein-Barr virus (EBV)-encoded BamHI-A rightward frame 1 (BARF1) is a putative viral oncogene in EBV-infected stomach cancer. The aim of the present study was to investigate BARF1-induced cellular protein and microRNA alterations. In this study, BARF1-expressing stomach cancer cells showed a high rate of proliferation, high levels of NF?B, and miR-146a upregulation, which was reversed by NF?B knockdown. During BARF1-induced NF?B upregulation, hCSF1 receptor level was unchanged. Knockdown of BARF1 in the naturally EBV-infected YCCEL1 stomach cancer cells suppressed cell proliferation, and downregulated NF?B and miR-146a. SMAD4 was identified as a miR-146a target and was downregulated in BARF1-expressing cells, whereas SMAD4 expression was restored by anti-miR-146a. Knockdown of BARF1 in YCCEL1 cells upregulated SMAD4, and this effect was reversed by miR-146a overexpression. Transfection of BARF1-expressing cells with pCEP4-SMAD4 abolished the cell proliferating effect of BARF1. In stomach cancer tissues, miR-146a was expressed at higher levels, and more frequent NF?B nuclear positivity immunohistochemically, but not of SMAD4 nuclear loss was found in the EBV-positive group compared with the EBV-negative group. In conclusion, EBV-encoded BARF1 promotes cell proliferation in stomach cancer by upregulating NF?B and miR-146a and downregulating SMAD4, thereby contributing to EBV-induced stomach cancer progression.

View details for DOI 10.18632/oncotarget.10511

View details for Web of Science ID 000391352800023

View details for PubMedID 27438138

View details for PubMedCentralID PMC5347686
Applying Mass Cytometry to the Analysis of Lymphoid Populations in Transplantation. American journal of transplantation Krams, S. M., Schaffert, S., Lau, A. H., Martinez, O. M. 2016
Abstract

Single-cell flow cytometric techniques have been indispensable to improving our understanding of the phenotype and function of immune cell subsets that are important in both rejection and tolerance after transplant. Mass cytometry, or cytometry by time of flight, is a single-cell-based platform that utilizes antibodies conjugated to rare heavy metal ions for analysis of cellular proteins by a time-of-flight mass spectrometer. This new technology allows for the evaluation of >40 simultaneous cellular parameters in a single sample because the limitation of spectral overlap, seen in conventional flow cytometry, is eliminated. In this review, we discuss the current state of mass cytometry, describe the advantages and disadvantages compared with multiparameter flow cytometry, introduce novel methods of high-dimensional data analysis and visualization, and review some recent studies using mass cytometry to profile the immune systems of healthy people and transplant recipients.

View details for DOI 10.1111/ajt.14145

View details for PubMedID 27888565
Mass cytometry reveals a distinct immunoprofile of operational tolerance in pediatric liver transplantation. Pediatric transplantation Lau, A. H., Vitalone, M. J., Haas, K., Shawler, T., Esquivel, C. O., Berquist, W. E., Martinez, O. M., Castillo, R. O., Krams, S. M. 2016
Abstract

Long-term IS in transplant patients has significant morbidity, poorer quality of life, and substantial economic costs. TOL, defined as graft acceptance without functional impairment in the absence of IS, has been achieved in some pediatric LT recipients. Using mass cytometry, peripheral blood immunotyping was performed to characterize differences between tolerant patients and patients who are stable on single-agent IS. Single-cell mass cytometry was performed using blood samples from a single-center pediatric LT population of operationally tolerant patients to comprehensively characterize the immune cell populations in the tolerant state compared with patients on chronic low-dose IS. Specific T-cell populations of interest were confirmed by flow cytometry. This high-dimensional phenotypic analysis revealed distinct immunoprofiles between transplant populations as well as a CD4(+) TOT (CD4(+) CD5(+) CD25(+) CD38(-/lo) CD45RA) that correlates with tolerance in pediatric LT recipients. In TOL patients, the TOT was significantly increased as compared to patients stable on low levels of IS. This TOT cell was confirmed by flow cytometry and is distinct from classic Treg cells. These results demonstrate the power of mass cytometry to discover significant immune cell signatures that have diagnostic potential.

View details for DOI 10.1111/petr.12795

View details for PubMedID 27781378
Harbingers of Long-term Graft Outcome: Go With the Flow? TRANSPLANTATION Martinez, O. M. 2016; 100 (9): 1799?1800
View details for PubMedID 27391194
MicroRNAs in EBV plus B cell lymphomas and secreted exosomes Kaul, V., Harris-Arnold, A., Esquivel, C., Martinez, O. M., Krams, S. M. LIPPINCOTT WILLIAMS & WILKINS. 2016: S257
View details for Web of Science ID 000436953200401
Natural Killer cells demonstrate functional and receptor diversity in response to alloantigen Hatton, O., Shawler, T., Blish, C., Martinez, O. M., Krams, S. M. LIPPINCOTT WILLIAMS & WILKINS. 2016: S121
View details for Web of Science ID 000436953200193
Generation of novel Cag-Luc-EGFP transgene-expressing murine embryonic stem cell derived hepatoblasts as a transplantable cell source Cisneros, T., Arredondo-Guerrero, J., Qu, X., Krams, S. M., Esquivel, C., Martinez, O. LIPPINCOTT WILLIAMS & WILKINS. 2016: S864
View details for Web of Science ID 000436953202146
Inhibition of the constitutively active PI3K/Akt/mTOR signaling pathway in epstein-barr virus B cell lymphomas demonstrate both antitumor and antirejection properties Sang, A., Ivison, G. T., Qu, X., Esquivel, C. O., Krams, S. M., Martinez, O. M. LIPPINCOTT WILLIAMS & WILKINS. 2016: S258
View details for Web of Science ID 000436953200402
NKG2A+natural killer cells dominate the response to latent Epstein-Barr virus infection Hatton, O., Strauss-Albee, D., Krams, S. M., Blish, C., Martinez, O. LIPPINCOTT WILLIAMS & WILKINS. 2016: S383
View details for Web of Science ID 000436953200598
Epstein-Barr virus-associated lymphoepithelial carcinoma after pediatric liver transplant LIVER TRANSPLANTATION Sang, A. X., Harris-Arnold, A., Kambham, N., Martinez, O. M., Krams, S. M., Strichartz, D., Esquivel, C. O. 2016; 22 (6): 849?53
View details for PubMedID 27065464

View details for PubMedCentralID PMC4882240
Liver microRNA Profile of Induced Allograft Tolerance. Transplantation Vitalone, M. J., Wei, L., Fujiki, M., Lau, A. H., Littau, E., Esquivel, C., Martinez, O. M., Krams, S. M. 2016; 100 (4): 781-790
Abstract

Although the liver is less immunogenic than other solid organs, most liver transplant recipients receive lifelong immunosuppression. In both experimental models and clinical transplantation, total lymphoid irradiation (TLI) has been shown to induce allograft tolerance. Our goal was to identify the microRNAs (miRNAs) expressed in tolerant liver allograft recipients in an experimental model of TLI-induced tolerance.To identify the miRNAs associated with TLI-induced tolerance, we examined syngeneic recipients (Lewis?Lewis) and allogeneic recipients (Dark Agouti?Lewis) of orthotropic liver transplants that received posttransplant TLI, allogeneic recipients that were not treated posttransplantation and experienced acute rejection, and native Dark Agouti livers. Quantitative-polymerase chain reaction miRNA array cards were used to profile liver grafts.We identified 12 miRNAs that were specifically and significantly increased during acute rejection. In early tolerance, 33 miRNAs were altered compared with syngeneic livers, with 80% of the miRNAs increased. In established tolerance, 42 miRNAs were altered. In addition, miR-142-5p and miR-181a demonstrated increased expression in tolerant livers (both early and established tolerance) as compared with syngeneic livers. A principal component analysis of all miRNAs assayed demonstrated a profile in established tolerance that was closely related to that seen in syngeneic livers.The miRNA profile of established tolerant allografts is very similar to syngeneic grafts, suggesting tolerance may be a return to an immunological state of quiescence.

View details for DOI 10.1097/TP.0000000000001105

View details for PubMedID 26950716
Post-transplant lymphoproliferative disorders NATURE REVIEWS DISEASE PRIMERS Dharnidharka, V. R., Webster, A. C., Martinez, O. M., Preiksaitis, J. K., Leblond, V., Choquet, S. 2016; 2
Abstract

Post-transplant lymphoproliferative disorders (PTLDs) are a group of conditions that involve uncontrolled proliferation of lymphoid cells as a consequence of extrinsic immunosuppression after organ or haematopoietic stem cell transplant. PTLDs show some similarities to classic lymphomas in the non-immunosuppressed general population. The oncogenic Epstein-Barr virus (EBV) is a key pathogenic driver in many early-onset cases, through multiple mechanisms. The incidence of PTLD varies with the type of transplant; a clear distinction should therefore be made between the conditions after solid organ transplant and after haematopoietic stem cell transplant. Recipient EBV seronegativity and the intensity of immunosuppression are among key risk factors. Symptoms and signs depend on the localization of the lymphoid masses. Diagnosis requires histopathology, although imaging techniques can provide additional supportive evidence. Pre-emptive intervention based on monitoring EBV levels in blood has emerged as the preferred strategy for PTLD prevention. Treatment of established disease includes reduction of immunosuppression and/or administration of rituximab (a B cell-specific antibody against CD20), chemotherapy and EBV-specific cytotoxic T cells. Despite these strategies, the mortality and morbidity remains considerable. Patient outcome is influenced by the severity of presentation, treatment-related complications and risk of allograft loss. New innovative treatment options hold promise for changing the outlook in the future.

View details for DOI 10.1038/nrdp.2015.88

View details for Web of Science ID 000381359800001

View details for PubMedID 27189056
Epstein-Barr Virus Modulates Host Cell MicroRNA-194 to Promote IL-10 Production and B Lymphoma Cell Survival AMERICAN JOURNAL OF TRANSPLANTATION Harris-Arnold, A., Arnold, C. P., Schaffert, S., Hatton, O., Krams, S. M., Esquivel, C. O., Martinez, O. M. 2015; 15 (11): 2814-2824
View details for DOI 10.1111/ajt.13375

View details for Web of Science ID 000363264800006

View details for PubMedID 26147452
NKp46 clusters at the immune synapse and regulates NK cell polarization FRONTIERS IN IMMUNOLOGY Hadad, U., Thauland, T. J., Martinez, O. M., Butte, M. J., Porgador, A., Krams, S. M. 2015; 6
Abstract

Natural killer (NK) cells play an important role in first-line defense against tumor and virus-infected cells. The activity of NK cells is tightly regulated by a repertoire of cell surface expressed inhibitory and activating receptors. NKp46 is a major NK cell-activating receptor that is involved in the elimination of target cells. NK cells form different types of synapses that result in distinct functional outcomes: cytotoxic, inhibitory, and regulatory. Recent studies revealed that complex integration of NK receptor signaling controls cytoskeletal rearrangement and other immune synapse-related events. However, the distinct nature by which NKp46 participates in NK immunological synapse formation and function remains unknown. In this study, we determined that NKp46 forms microclusters structures at the immune synapse between NK cells and target cells. Over-expression of human NKp46 is correlated with increased accumulation of F-actin mesh at the immune synapse. Concordantly, knock-down of NKp46 in primary human NK cells decreased recruitment of F-actin to the synapse. Live cell imaging experiments showed a linear correlation between NKp46 expression and lytic granules polarization to the immune synapse. Taken together, our data suggest that NKp46 signaling directly regulates the NK lytic immune synapse from early formation to late function.

View details for DOI 10.3389/fimmu.2015.00495

View details for Web of Science ID 000361914900001

View details for PubMedID 26441997

View details for PubMedCentralID PMC4585260
Orienting incoming PhD students through hands-on training in immunology research methods Sweere, J., Seong, Y., Schaffert, S., Spitzer, M., Martinez, O., Butte, M. AMER ASSOC IMMUNOLOGISTS. 2015
View details for Web of Science ID 000379404501191
Genomic Approaches to Unraveling the Diversity and Dynamics of the Human Virome AMERICAN JOURNAL OF TRANSPLANTATION Martinez, O. M., Ford, M. L. 2014; 14 (12): 2677
View details for DOI 10.1111/ajt.13064

View details for Web of Science ID 000345294300002
The interplay between Epstein-Barr virus and B lymphocytes: implications for infection, immunity, and disease. Immunologic research Hatton, O. L., Harris-Arnold, A., Schaffert, S., Krams, S. M., Martinez, O. M. 2014; 58 (2-3): 268-276
Abstract

Human B cells are the primary targets of Epstein-Barr virus (EBV) infection. In most cases, EBV infection is asymptomatic because of a highly effective host immune response, but some individuals develop self-limiting infectious mononucleosis, while others develop EBV-associated lymphoid or epithelial malignancies. The viral and immune factors that determine the outcome of infection are not understood. The EBV life cycle includes a lytic phase, culminating in the production of new viral particles, and a latent phase, during which the virus remains largely silent for the lifetime of the host in memory B cells. Thus, in healthy individuals, there is a tightly orchestrated interplay between EBV and the host that allows the virus to persist. To promote viral persistence, EBV has evolved a variety of strategies to modulate the host immune response including inhibition of immune cell function, blunting of apoptotic pathways, and interfering with antigen processing and presentation pathways. In this article, we focus on mechanisms by which dysregulation of the host B cell and immune modulation by the virus can contribute to development of EBV+ B cell lymphomas.

View details for DOI 10.1007/s12026-014-8496-1

View details for PubMedID 24619311
NK cells after transplantation: friend or foe. Immunologic research Hadad, U., Martinez, O., Krams, S. M. 2014; 58 (2-3): 259-267
Abstract

Natural killer (NK) cells are effector cells of the innate immune system that can lyse target cells without prior sensitization and have an important role in host defense to pathogens and transformed cells. A balance between negative and positive signals transmitted via germ line-encoded inhibitory and activating receptors controls the function of NK cells. Although the concept of "missing-self" would suggest that NK cells could target foreign allografts, the prevailing dogma has been that NK cells are not active participants in the mechanisms that culminate in the rejection of solid organ allografts. Recent studies, however, challenge this conclusion and instead implicate NK cells in contributing to both graft rejection and tolerance to an allograft. In this review, we highlight recent studies with the goal of understanding the complex NK cell interactions that impact alloimmunity.

View details for DOI 10.1007/s12026-014-8493-4

View details for PubMedID 24522700
Differential expression and functions of microRNAs in liver transplantation and potential use as non-invasive biomarkers. Transplant immunology Wei, L., Gong, X., Martinez, O. M., Krams, S. M. 2013; 29 (1-4): 123-129
Abstract

MicroRNAs (miRNAs) are important regulators in many biologic processes and have been implicated in the control of genes relevant to acute rejection and liver functions. Here we review the miRNAs specifically expressed in allografts during acute rejection and discuss potential roles for these miRNAs in liver dysfunction. We focus on miRNAs dysregulated both in the liver and in peripheral blood mononuclear cells and include a discussion of the potential for these miRNAs as non-invasive biomarkers to reflect liver status posttransplant.

View details for DOI 10.1016/j.trim.2013.08.005

View details for PubMedID 24001411
Natural Killer Cell-Activating Receptor NKG2D Mediates Innate Immune Targeting of Allogeneic Neural Progenitor Cell Grafts STEM CELLS Phillips, L. K., Gould, E. A., Babu, H., Krams, S. M., Palmer, T. D., Martinez, O. M. 2013; 31 (9): 1829-1839
Abstract

Cell replacement therapy holds promise for a number of untreatable neurological or psychiatric diseases but the immunogenicity of cellular grafts remains controversial. Emerging stem cell and reprogramming technologies can be used to generate autologous grafts that minimize immunological concerns but autologous grafts may carry an underlying genetic vulnerability that reduces graft efficacy or survival. Healthy allogeneic grafts are an attractive and commercially scalable alternative if immunological variables can be controlled. Stem cells and immature neural progenitor cells (NPC) do not express major histocompatibility complex (MHC) antigens and can evade adaptive immune surveillance. Nevertheless, in an experimental murine model, allogeneic NPCs do not survive and differentiate as well as syngeneic grafts, even when traditional immunosuppressive treatments are used. In this study, we show that natural killer (NK) cells recognize the lack of self-MHC antigens on NPCs and pose a barrier to NPC transplantation. NK cells readily target both syngeneic and allogeneic NPC, and killing is modulated primarily by NK-inhibiting "self" class I MHC and NK-activating NKG2D-ligand expression. The absence of NKG2D signaling in NK cells significantly improves NPC-derived neuron survival and differentiation. These data illustrate the importance of innate immune mechanisms in graft outcome and the potential value of identifying and targeting NK cell-activating ligands that may be expressed by stem cell derived grafts.

View details for DOI 10.1002/stem.1422

View details for Web of Science ID 000325255700010

View details for PubMedID 23733329
PI3K Inhibition Augments the Efficacy of Rapamycin in Suppressing Proliferation of Epstein-Barr Virus (EBV) plus B Cell Lymphomas AMERICAN JOURNAL OF TRANSPLANTATION Furukawa, S., Wei, L., Krams, S. M., Esquivel, C. O., Martinez, O. M. 2013; 13 (8): 2035-2043
Abstract

Posttransplant lymphoproliferative disorder (PTLD) continues to be a devastating and potentially life-threatening complication in organ transplant recipients. PTLD is associated with EBV infection and can result in malignant B cell lymphomas. Here we demonstrate that the PI3K/Akt/mTOR pathway is highly activated in EBV+ B cell lymphoma lines derived from patients with PTLD. Treatment with the mTORC1 inhibitor Rapamycin (RAPA) partially inhibited the proliferation of EBV+ B cell lines. Resistance to RAPA treatment correlated with high levels of Akt phosphorylation. An mTORC1/2 inhibitor and a PI3K/mTOR dual inhibitor suppressed Akt phosphorylation and showed a greater anti-proliferative effect on EBV+ B lymphoma lines compared to RAPA. EBV+ B cell lymphoma lines expressed high levels of PI3K?. We demonstrate that PI3K? is responsible for Akt activation in EBV+ B cell lymphomas, and that selective inhibition of PI3K? by either siRNA, or a small molecule inhibitor, augmented the anti-proliferative effect of RAPA on EBV+ B cell lymphomas. These results suggest that PI3K? is a novel, potential therapeutic target for the treatment of EBV-associated PTLD and that combined blockade of PI3K? and mTOR provides increased efficacy in inhibiting proliferation of EBV+ B cell lymphomas.

View details for DOI 10.1111/ajt.12328

View details for Web of Science ID 000322330000013

View details for PubMedID 23841834
Syk-Induced Phosphatidylinositol-3-Kinase Activation in EpsteinBarr Virus Posttransplant Lymphoproliferative Disorder AMERICAN JOURNAL OF TRANSPLANTATION Hatton, O., Lambert, S. L., Phillips, L. K., Vaysberg, M., Natkunam, Y., Esquivel, C. O., Krams, S. M., Martinez, O. M. 2013; 13 (4): 883-890
Abstract

Posttransplant lymphoproliferative disorder (PTLD)-associated Epstein-Barr virus (EBV)+ B cell lymphomas are serious complications of solid organ and bone marrow transplantation. The EBV protein LMP2a, a B cell receptor (BCR) mimic, provides survival signals to virally infected cells through Syk tyrosine kinase. Therefore, we explored whether Syk inhibition is a viable therapeutic strategy for EBV-associated PTLD. We have shown that R406, the active metabolite of the Syk inhibitor fostamatinib, induces apoptosis and cell cycle arrest while decreasing downstream phosphatidylinositol-3'-kinase (PI3K)/Akt signaling in EBV+ B cell lymphoma PTLD lines in vitro. However, Syk inhibition did not inhibit or delay the in vivo growth of solid tumors established from EBV-infected B cell lines. Instead, we observed tumor growth in adjacent inguinal lymph nodes exclusively in fostamatinib-treated animals. In contrast, direct inhibition of PI3K/Akt significantly reduced tumor burden in a xenogeneic mouse model of PTLD without evidence of tumor growth in adjacent inguinal lymph nodes. Taken together, our data indicate that Syk activates PI3K/Akt signaling which is required for survival of EBV+ B cell lymphomas. PI3K/Akt signaling may be a promising therapeutic target for PTLD, and other EBV-associated malignancies.

View details for DOI 10.1111/ajt.12137

View details for Web of Science ID 000316911900011

View details for PubMedID 23398911
MicroRNA 194 Regulates Apoptosis in Epstein Barr Virus plus B Cell Lymphomas Associated with Post-Transplant Lymphoproliferative Disorder. 13th American Transplant Congress (ATC) Harris-Arnold, A., Lambert, S., Krams, S., Martinez, O. WILEY-BLACKWELL. 2013: 96?96
View details for Web of Science ID 000318240300215
PI3K delta Inhibition Augments the Efficacy of mTOR Inhibitor Rapamycin on the Proliferation of Epstein-Barr Virus (EBV) plus B Cell Lymphomas. 13th American Transplant Congress (ATC) Furukawa, S., Hatton, O., Krams, S., Esquivel, C., Martinez, O. WILEY-BLACKWELL. 2013: 96?97
View details for Web of Science ID 000318240300216
Src Kinase and Syk Activation Initiate PI3K Signaling by a Chimeric Latent Membrane Protein 1 in Epstein-Barr Virus (EBV) plus B Cell Lymphomas PLOS ONE Hatton, O., Lambert, S. L., Krams, S. M., Martinez, O. M. 2012; 7 (8)
Abstract

The B lymphotrophic ?-herpesvirus EBV is associated with a variety of lymphoid- and epithelial-derived malignancies, including B cell lymphomas in immunocompromised and immunosuppressed individuals. The primary oncogene of EBV, latent membrane protein 1 (LMP1), activates the PI3K/Akt pathway to induce the autocrine growth factor, IL-10, in EBV-infected B cells, but the mechanisms underlying PI3K activation remain incompletely understood. Using small molecule inhibition and siRNA strategies in human B cell lines expressing a chimeric, signaling-inducible LMP1 protein, nerve growth factor receptor (NGFR)-LMP1, we show that NGFR-LMP1 utilizes Syk to activate PI3K/Akt signaling and induce IL-10 production. NGFR-LMP1 signaling induces phosphorylation of BLNK, a marker of Syk activation. Whereas Src kinases are often required for Syk activation, we show here that PI3K/Akt activation and autocrine IL-10 production by NGFR-LMP1 involves the Src family kinase Fyn. Finally, we demonstrate that NGFR-LMP1 induces phosphorylation of c-Cbl in a Syk- and Fyn-dependent fashion. Our results indicate that the EBV protein LMP1, which lacks the canonical ITAM required for Syk activation, can nevertheless activate Syk, and the Src kinase Fyn, resulting in downstream c-Cbl and PI3K/Akt activation. Fyn, Syk, and PI3K/Akt antagonists thus may present potential new therapeutic strategies that target the oncogene LMP1 for treatment of EBV+ B cell lymphomas.

View details for DOI 10.1371/journal.pone.0042610

View details for Web of Science ID 000307284100143

View details for PubMedID 22880054

View details for PubMedCentralID PMC3411813
Differential Expression of MicroRNAs During Allograft Rejection AMERICAN JOURNAL OF TRANSPLANTATION Wei, L., Wang, M., Qu, X., MAH, A., Xiong, X., Harris, A. G., Phillips, L. K., Martinez, O. M., Krams, S. M. 2012; 12 (5): 1113-1123
Abstract

MicrorRNA are small noncoding RNA molecules that regulate the posttranscriptional expression of target genes. In addition to being involved in many biologic processes, microRNAs are important regulators in innate and adaptive immune responses. Distinct sets of expressed microRNAs are found in different cell types and tissues and aberrant expression of microRNAs is associated with many disease states. MicroRNA expression was examined in a model of heterotopic heart transplantation by microarray analyses and a unique profile was detected in rejecting allogeneic transplants (BALB/c ? C57BL/6) as compared to syngeneic transplants (C57BL/6 ? C57BL/6). The microRNA miR-182 was significantly increased in rejecting cardiac allografts and in mononuclear cells that infiltrate the grafts. Forkhead box (FOX) proteins are a family of important transcription factors and FOXO1 is a target of miR-182. As miR-182 increases after transplant, there is a concomitant posttranscriptional decrease in FOXO1 expression in heart allografts that is localized to both the cardiomyocytes and CD3(+) T cells. The microRNA miR-182 is significantly increased in both peripheral blood mononuclear cells and plasma during graft rejection suggesting potential as a biomarker of graft status. Our results identify microRNAs that may regulate alloimmune responses and graft outcomes.

View details for DOI 10.1111/j.1600-6143.2011.03958.x

View details for Web of Science ID 000303235100010

View details for PubMedID 22300508

View details for PubMedCentralID PMC3461331
Modulation of IL-10 Specific B Cell microRNAs by Latent Membrane Protein 1 and Epstein-Barr Virus 24th Annual Meeting of the European-Association-for-Cardiothoracic-Surgery Harris-Arnold, A., Lambert, S., Krams, S., Martinez, O. WILEY-BLACKWELL. 2012: 162?162
View details for Web of Science ID 000303235501077
Emerging therapeutic strategies for Epstein-Barr virus+ post-transplant lymphoproliferative disorder PEDIATRIC TRANSPLANTATION Hatton, O., Martinez, O. M., Esquivel, C. O. 2012; 16 (3): 220-229
Abstract

De novo malignancies represent an increasing concern in the transplant population, particularly as long-term graft and patient survival improves. EBV-associated B-cell lymphoma in the setting of PTLD is the leading malignancy in children following solid organ transplantation. Therapeutic strategies can be categorized as pharmacologic, biologic, and cell-based but the variable efficacy of these approaches and the complexity of PTLD suggest that new treatment options are warranted. Here, we review current therapeutic strategies for treatment of PTLD. We also describe the life cycle of EBV, addressing the viral mechanisms that contribute to the genesis and persistence of EBV+ B-cell lymphomas. Specifically, we focus on the oncogenic signaling pathways activated by the EBV LMP1 and LMP2a to understand the underlying mechanisms and mediators of lymphomagenesis with the goal of identifying novel, rational therapeutic targets for the treatment of EBV-associated malignancies.

View details for DOI 10.1111/j.1399-3046.2012.01656.x

View details for Web of Science ID 000302619500013

View details for PubMedID 22353174
Changes in natural killer cell subsets in pediatric liver transplant recipients PEDIATRIC TRANSPLANTATION Pham, B., Piard-Ruster, K., Silva, R., Gallo, A., Esquivel, C. O., Martinez, O. M., Krams, S. M. 2012; 16 (2): 176-182
Abstract

NK cells are important in the immune response against tumors and virally infected cells. A balance between inhibitory and activating receptors controls the effector functions of NK cells. We examined the fate of circulating NK cells and the expression of the NK cell-activating receptors in pediatric liver transplant recipients. Blood specimens were collected from 38 pediatric liver transplant recipients before transplant, and at one wk, one, three, six, and nine months, and one yr post-transplant. PBMCs were isolated and analyzed for the levels of NK cell activation receptors NKp30, NKp46, and NKG2D in the CD56(dim) CD16(+) and CD56(bright) CD16(+/-) subsets of NK cells. We demonstrated that there is a significant decrease in the percentage of circulating NK cells post-transplant (pretransplant 7.69 � 1.54 vs. one wk post-transplant 1.73 � 0.44) in pediatric liver transplant recipients. Interestingly, NKp30 expression is significantly increased, while NKp46 and NKG2D levels remain stable on the NK cells that persist at one wk post-transplant. These data indicate that the numbers and subsets of circulating NK cells are altered in children after liver transplantation.

View details for DOI 10.1111/j.1399-3046.2012.01653.x

View details for Web of Science ID 000300709600017

View details for PubMedID 22360401

View details for PubMedCentralID PMC3306774
Syk Activation of Phosphatidylinositol 3-Kinase/Akt Prevents HtrA2-dependent Loss of X-linked Inhibitor of Apoptosis Protein (XIAP) to Promote Survival of Epstein-Barr Virus plus (EBV plus ) B Cell Lymphomas JOURNAL OF BIOLOGICAL CHEMISTRY Hatton, O., Phillips, L. K., Vaysberg, M., Hurwich, J., Krams, S. M., Martinez, O. M. 2011; 286 (43): 37368-37378
Abstract

B cell lymphoma survival requires tonic or ligand-independent signals through activation of Syk by the B cell receptor. The Epstein-Barr virus (EBV) protein latent membrane 2a (LMP2a), a mimic of the B cell receptor, provides constitutive survival signals for latently infected cells through Syk activation; however, the precise downstream mechanisms coordinating this survival response in EBV+ B cell lymphomas remain to be elucidated. Herein, we assess the mechanism of Syk survival signaling in EBV+ B cell lymphomas from post-transplant lymphoproliferative disorder (PTLD) to discover virally controlled therapeutic targets involved in lymphomagenesis and tumor progression. Using small molecule inhibition and siRNA strategies, we show that Syk inhibition reduces proliferation and induces apoptosis of PTLD-derived EBV+ B cell lines. Syk inhibition also reduces autocrine IL-10 production. Although Syk inhibition attenuates signaling through both the PI3K/Akt and Erk pathways, only PI3K/Akt inhibition causes apoptosis of PTLD-derived cell lines. Loss of the endogenous caspase inhibitor XIAP is observed after Syk or PI3K/Akt inhibition. The loss of XIAP and apoptosis that results from Syk or PI3K/Akt inhibition is reversed by inhibition of the mitochondrial protease HtrA2. Thus, Syk drives EBV+ B cell lymphoma survival through PI3K/Akt activation, which prevents the HtrA2-dependent loss of XIAP. Syk, Akt, and XIAP antagonists may present potential new therapeutic strategies for PTLD through targeting of EBV-driven survival signals.

View details for DOI 10.1074/jbc.M111.255125

View details for Web of Science ID 000296542400037

View details for PubMedID 21908615

View details for PubMedCentralID PMC3199484
HIGH DOSES OF CALCINEURIN INHIBITORS ARE ASSOCIATED WITH LOW LEVELS OF T REGULATORY CELLS IN PEDIATRIC LIVER TRANSPLANT RECIPIENTS. Piard-Ruster, K. S., Silva, R., Berquist, W., Pham, B., Gallo, A., Krams, S. M., Esquivel, C. O., Martinez, O. M. WILEY-BLACKWELL. 2011: 57?57
View details for Web of Science ID 000293251100066
Rejection of Small Intestinal Allografts Associates with a Unique Signature of microRNAs American Transplant Congress Krams, S. M., Wei, L., Pham, B., Harris, A., Castillo, R. O., Esquivel, C. O., Martinez, O. WILEY-BLACKWELL. 2011: 133?133
View details for Web of Science ID 000289318400339
Syk survival signals in Epstein-Barr virus plus B cell lymphomas Hatton, O., Lambert, S., Krams, S., Martinez, O. AMER ASSOC IMMUNOLOGISTS. 2011
View details for Web of Science ID 000209751708004
Latent membrane protein 1 of Epstein Barr Virus alters cellular microRNAs in B cell lymphomas Harris, A., Lambert, S., Krams, S., Martinez, O. AMER ASSOC IMMUNOLOGISTS. 2011
View details for Web of Science ID 000209751708102
Neural stem/progenitor cells are targets for natural killer cell-mediated killing Phillips, L., Gould, E., Palmer, T., Krams, S., Martinez, O. AMER ASSOC IMMUNOLOGISTS. 2011
View details for Web of Science ID 000209751708091
Neural Stem/Progenitor Cells Are Targets for Natural Killer Cell-Mediated Killing American Transplant Congress Phillips, L. K., GOULD, E. A., Palmer, T. D., Krams, S. M., Martinez, O. M. WILEY-BLACKWELL. 2011: 439?439
View details for Web of Science ID 000289318401623
Targeting Akt for Treatment of Post-Transplant Lymphoproliferative Disorder (PTLD) American Transplant Congress Hatton, O., Lambert, S., Krams, S. M., Esquivel, C. O., Martinez, O. M. WILEY-BLACKWELL. 2011: 137?137
View details for Web of Science ID 000289318400351
MHC Mismatch Inhibits Neurogenesis and Neuron Maturation in Stem Cell Allografts PLOS ONE Chen, Z., Phillips, L. K., Gould, E., Campisi, J., Lee, S. W., Ormerod, B. K., Zwierzchoniewska, M., Martinez, O. M., Palmer, T. D. 2011; 6 (3)
Abstract

The role of histocompatibility and immune recognition in stem cell transplant therapy has been controversial, with many reports arguing that undifferentiated stem cells are protected from immune recognition due to the absence of major histocompatibility complex (MHC) markers. This argument is even more persuasive in transplantation into the central nervous system (CNS) where the graft rejection response is minimal.In this study, we evaluate graft survival and neuron production in perfectly matched vs. strongly mismatched neural stem cells transplanted into the hippocampus in mice. Although allogeneic cells survive, we observe that MHC-mismatch decreases surviving cell numbers and strongly inhibits the differentiation and retention of graft-derived as well as endogenously produced new neurons. Immune suppression with cyclosporine-A did not improve outcome but non-steroidal anti-inflammatory drugs, indomethacin or rosiglitazone, were able to restore allogeneic neuron production, integration and retention to the level of syngeneic grafts.These results suggest an important but unsuspected role for innate, rather than adaptive, immunity in the survival and function of MHC-mismatched cellular grafts in the CNS.

View details for DOI 10.1371/journal.pone.0014787

View details for Web of Science ID 000289055700001

View details for PubMedID 21479168

View details for PubMedCentralID PMC3068158
Distinct Roles for the NK Cell-Activating Receptors in Mediating Interactions with Dendritic Cells and Tumor Cells JOURNAL OF IMMUNOLOGY Wai, L., Garcia, J. A., Martinez, O. M., Krams, S. M. 2011; 186 (1): 222-229
Abstract

NK cells are innate immune cells that are important in tumor immunity, but also have the ability to modulate the adaptive immune system through cytokine production or direct cell-cell interactions. This study investigates the interaction of NK cells with dendritic cells (DCs) and tumor cells, and the role of specific NK cell-activating receptors in this process. Primary rat NK cells and an NK cell line produced IFN-? when cocultured with either DCs or the rat hepatoma cell line McA-RH7777 (McA). This NK cell activation by DCs and McA required cell-cell contact and was dependent on distinct NK-activating receptors. Silencing NK cell expression of NKp46 and NKp30 significantly diminished DC- and McA-mediated NK cell IFN-? production, respectively. NK cells killed immature and mature DCs independently of NKp46, NKp30, and NKG2D; however, cytotoxicity against McA cells was dependent on NKp30 and NKG2D. Thus, we have shown in this study that NKp30 plays dual activating roles in NK-McA tumor interactions by mediating cytokine production and cytotoxicity. More importantly, NK cells are activated by both DCs and hepatoma cells to produce IFN-?, but require distinct NK cell-activating receptors, NKp46 and NKp30, respectively. Our data suggest that therapeutics could be developed specifically to target NK-DC interactions without compromising NK tumor immunity.

View details for DOI 10.4049/jimmunol.1002597

View details for Web of Science ID 000285688700029

View details for PubMedID 21106845

View details for PubMedCentralID PMC3433947
Toll-Like Receptor 4 Contributes to Small Intestine Allograft Rejection TRANSPLANTATION Krams, S. M., Wang, M., Castillo, R. O., Ito, T., Phillips, L., Higgins, J., Kambham, N., Esquivel, C. O., Martinez, O. M. 2010; 90 (12): 1272-1277
Abstract

Although outcomes for small intestine transplantation (SIT) have improved in recent years, allograft rejection rates remain among the highest of solid organ grafts. The high load of commensal bacteria in the small intestine may contribute through activation of the toll-like receptor (TLR) pathway. In this study, we examine the participation of TLR4 in acute allograft rejection in an orthotopic mouse model of SIT.Wild-type C57Bl/6 (H-2b) or TLR49(-/-) (H-2b) mice were transplanted with syngeneic (C57Bl/6), allogeneic (BALB/c; H-2d), or F1 (BALB/cxC57Bl/6; H-2d/b) vascularized, orthotopic small intestine grafts. Graft recipients were killed on days 2 to 6 posttransplant. Serum cytokines were measured by Luminex, and tissue was obtained for histology and quantitative real-time polymerase chain reaction.BALB/c grafts transplanted into C57Bl/6 recipients exhibited mixed inflammatory infiltrates, destruction of the mucosa, and significant apoptosis. TLR2 and TLR4 transcripts were modestly increased in syngeneic grafts on days 2 and 6 compared with native bowel, whereas TLR2 and TLR4 were significantly increased on days 2 and 6 in allogeneic grafts. Although fully mismatched and F1 grafts were rejected by C57Bl/6 recipients (mean survival time=8.2 and 9.3 days, respectively), graft survival was significantly prolonged in TLR4(-/-) recipients (mean survival time=10.6 and 14.3 days, respectively). Proinflammatory cytokines were markedly reduced in TLR4(-/-) graft recipients.Small intestine graft survival is prolonged in the absence of TLR4, suggesting that gut flora associated with the graft may augment alloimmune responses through TLR4. Thus, the TLR pathway may be a novel therapeutic target for improving SIT allograft survival.

View details for DOI 10.1097/TP.0b013e3181fdda0d

View details for Web of Science ID 000285377100006

View details for PubMedID 21197709

View details for PubMedCentralID PMC3799863
Acute Rejection of Small Intestine Allografts Is Associated With Increased Expression of Toll-like Receptors TRANSPLANTATION PROCEEDINGS Castillo, R. O., Wang, M., Ito, T., Higgins, J., Esquivel, C. O., Krams, S. M., Martinez, O. M. 2010; 42 (7): 2676-2678
Abstract

Although outcomes after intestinal transplantation have steadily improved owing to advances in immunosuppressive therapy, operative techniques, and postoperative medical management, rejection of the intestinal allograft continues to be a major clinical problem and constitutes the primary reason for graft loss. Although the adaptive immune system has been the major focus of investigation regarding regulation of rejection of the intestinal allograft, the role of the innate immune system has recently become of increased interest. We hypothesized that microbial products of the microflora associated with the intestinal allograft may engage the Toll-like receptor pathway of the innate immune system to potentiate alloimmune responses and rejection of the allograft. To investigate this, we established a murine model for orthotopic intestinal transplantation and allograft rejection. Using this model, we show that the expression of Toll-like receptor 2 is increased 50-fold and the expression of Toll-like receptor 4 is increased 200-fold during rejection of the allograft. We then performed survival studies that showed increased survival of mice, which had the Toll-like receptor knocked out. These preliminary studies suggest an important role for in innate immune system in acute rejection of the small intestinal allografts, and as such represents an emerging and promising area of investigation.

View details for DOI 10.1016/j.transproceed.2010.05.157

View details for Web of Science ID 000281942200052

View details for PubMedID 20832568
Identification of the rat NKG2D ligands, RAE1L and RRLT, and their role in allograft rejection EUROPEAN JOURNAL OF IMMUNOLOGY Zhuo, M., Fujiki, M., Wang, M., Piard-Ruster, K., Wai, L., Wei, L., Martinez, O. M., Krams, S. M. 2010; 40 (6): 1748-1757
Abstract

NKG2D is a receptor expressed by NK cells and subsets of T lymphocytes. On NK cells, NKG2D functions as a stimulatory receptor that induces effector functions. We cloned and expressed two rat NKG2D ligands, both members of the RAE1 family, RAE1L and RRLT, and demonstrate that these ligands can induce IFN-gamma secretion and cytotoxicity by rat NK cells. To examine changes in expression of NKG2D and the NKG2D ligands RAE1L and RRLT after transplantation, we used a Dark Agouti (DA)-->Lewis rat model of liver transplantation. NKG2D expression was significantly increased in allogeneic liver grafts by day 7 post-transplant. Ligands of NKG2D, absent in normal liver, were readily detected in both syngeneic and allogeneic liver grafts by day 1 post-transplant. By day 7 post-transplant, hepatocyte RAE1L and RRLT expression was significantly and specifically increased in liver allografts. In contrast to acute rejection that develops in the DA-->Lewis model, transplantation of Lewis livers into DA recipients (Lewis-->DA) results in spontaneous tolerance. Interestingly, expression of RAE1L and RRLT is low in Lewis-->DA liver allografts, but significantly increased in DA-->Lewis liver allografts undergoing rejection. In conclusion, our results suggest that expression of NKG2D ligands may be important in allograft rejection.

View details for DOI 10.1002/eji.200939779

View details for Web of Science ID 000279077200023

View details for PubMedID 20306467

View details for PubMedCentralID PMC3057641
MicroRNAs as Immune Regulators: Implications for Transplantation AMERICAN JOURNAL OF TRANSPLANTATION Harris, A., Krams, S. M., Martinez, O. M. 2010; 10 (4): 713-719
Abstract

The explosion of genetic information from recent advances in sequencing technologies, bioinformatics and genomics highlights the importance of understanding mechanisms involved in gene expression and regulation. Over the last decade, it has become clear that small ribonucleic acids (RNAs) are a central component of the cellular gene regulatory network. MicroRNAs (miRNAs) are a family of endogenous, small, noncoding single-stranded RNA of approximately 22 nucleotides in length that act as posttranscriptional gene regulatory elements. MicroRNAs can inhibit de novo protein synthesis by blocking translation through base-pairing with complementary messenger RNA (mRNA) and also suppress translation by promoting degradation of target mRNA. MicroRNAs are intimately involved in a variety of biologic processes including development, hematopoietic cell differentiation, apoptosis and proliferation. To date, over 800 human miRNAs have been identified, though the biologic function of only a fraction of miRNAs has been elucidated. Here, we discuss how miRNAs are produced, identified and quantitated, and focus on several key miRNAs that govern expression of genes relevant to allograft rejection, tolerance induction and posttransplant infection. Finally, we discuss potential ways in which the miRNA network can be modulated that ultimately may offer new strategies to promote long-term graft survival.

View details for DOI 10.1111/j.1600-6143.2010.03032.x

View details for Web of Science ID 000275768300004

View details for PubMedID 20199506
NK Cells Promote Acceptance of Liver Allografts Ito, T., Wang, M., Wai, L., Esquivel, C. O., Martinez, O. M., Krams, S. M. WILEY-BLACKWELL PUBLISHING, INC. 2010: 467
View details for Web of Science ID 000275921703199
The Role of NKG2D in Transplantation Wai, L., Wang, M., Ito, T., Piard-Ruster, K., Esquivel, C. O., Martinez, O. M., Krams, S. M. WILEY-BLACKWELL PUBLISHING, INC. 2010: 217
View details for Web of Science ID 000275921702016
The Effect of Calcineurin Inhibitor Dose on T Regulatory Cell Levels in Pediatric Liver Transplant Recipients. Piard-Ruster, K., Silva, R., Berquist, W., Pham, B., Gallo, A., Krams, S. M., Esquivel, C. O., Martinez, O. M. WILEY-BLACKWELL PUBLISHING, INC. 2010: 486
View details for Web of Science ID 000275921703268
Epstein-Barr Virus Latent Membrane Protein 1 Modulates Host MicroRNAs in B Cell Lymphomas Harris, A. C., Lambert, S. L., Krams, S. M., Martinez, O. M. WILEY-BLACKWELL PUBLISHING, INC. 2010: 295
View details for Web of Science ID 000275921702293
CD30 Activation of T Cells Causes Recruitment of TRAF Proteins and Induces P38-and Caspase-Dependent Apoptosis 10th American Transplant Congress Piard-Ruster, K. S., Hsieh, C. L., Krams, S. M., Martinez, O. M. WILEY-BLACKWELL. 2010: 364?364
View details for Web of Science ID 000275921702545
The Innate Immune Response Impairs Differentiation of Neural Progenitor Stem Cell Allografts 10th American Transplant Congress Phillips, L., Gould, E., Palmer, T., Krams, S. S., Martinez, O. M. WILEY-BLACKWELL. 2010: 365?365
View details for Web of Science ID 000275921702550
Effect of CP690,550 on the Jak/STAT Pathway in Adult Kidney Recipients. 10th American Transplant Congress Silva, R., Martinez, O. M., Krams, S. M., Busque, S. WILEY-BLACKWELL. 2010: 145?145
View details for Web of Science ID 000275921701356
Toll-Like Receptor 4 Contributes to Rejection of Small Intestine Allografts. 10th American Transplant Congress Krams, S. M., Wang, M., Castillo, R. O., Ito, T., Esquivel, C. O., Martinez, O. M. WILEY-BLACKWELL. 2010: 312?312
View details for Web of Science ID 000275921702359
Syk Inhibits Lymphoma Migration to the Lymph Node in a Xenotransplantaion Model of Epstein Barr Virus (EBV) plus Post-Transplant Lymphomproliferaive Disorder (PTLD) 10th American Transplant Congress Hatton, O., Lambert, S., Vaysberg, M., Krams, S. M., Esquivel, C. O., Martinez, O. M. WILEY-BLACKWELL. 2010: 62?62
View details for Web of Science ID 000275921701077
Natural Killer Cell Activation Receptor Expression Is Increased Post-Transplant in Pediatric Liver Transplant Recipients. 10th American Transplant Congress Pham, B., Silva, R., Piard-Ruster, K., Gallo, A., Esquivel, C. O., Martinez, O. M., Krams, S. M. WILEY-BLACKWELL. 2010: 486?486
View details for Web of Science ID 000275921703269
Induced Tolerance to Rat Liver Allografts Involves the Apoptosis of Intragraft T Cells and the Generation of CD4(+)CD25(+)FoxP3(+) T Regulatory Cells LIVER TRANSPLANTATION Fujiki, M., Esquivel, C. O., Martinez, O. M., Strober, S., Uemoto, S., Krams, S. M. 2010; 16 (2): 147-154
Abstract

Posttransplant total lymphoid irradiation is a nonmyeloablative regimen that has been extensively studied in rodent models for the induction of tolerance to bone marrow and solid organ allografts. Previous studies of experimental models and clinical transplantation have used total lymphoid irradiation in combination with anti-lymphocyte-depleting reagents and donor cell infusion to promote graft acceptance. In a rat model of orthotopic liver transplantation, we demonstrated that total lymphoid irradiation alone induced long-term graft survival. Apoptotic T cells were detected in markedly higher numbers in the livers of the total lymphoid irradiation-treated group in comparison with the control group of liver allograft recipients. Intragraft CD4(+)CD25(+)FoxP3(+) cells were increased in the total lymphoid irradiation group in the first week post-transplant and remained elevated in the graft and in the spleen. Importantly, the adoptive transfer of splenocytes from recipients that received posttransplant total lymphoid irradiation prolonged the survival of donor heart grafts, but not third-party heart grafts, whereas the depletion of CD4(+)CD25(+) cells from transferred splenocytes abrogated this prolongation. We conclude that posttransplant total lymphoid irradiation significantly increases the apoptosis of T cells in the liver graft and allows the accumulation of CD4(+)CD25(+)FoxP3(+) T regulatory cells, which facilitate the generation of donor-specific tolerance.

View details for DOI 10.1002/lt.21963

View details for Web of Science ID 000274437800005

View details for PubMedID 20104482

View details for PubMedCentralID PMC2937179
The Biology of Epstein-Barr Virus and Posttransplant Lymphoproliferative Disease POST-TRANSPLANT LYMPHOPROLIFERATIVE DISORDERS Martinez, O. M., Dharnidharka, V. R., Green, M., Webber, S. A. 2010: 29?43
View details for DOI 10.1007/978-3-642-01653-0_4

View details for Web of Science ID 000272722600004
Activation of the JAK/STAT Pathway in Epstein Barr Virus(+)-Associated Posttransplant Lymphoproliferative Disease: Role of Interferon-gamma AMERICAN JOURNAL OF TRANSPLANTATION Vaysberg, M., Lambert, S. L., Krams, S. M., Martinez, O. M. 2009; 9 (10): 2292-2302
Abstract

Epstein Barr virus (EBV) is associated with B-cell lymphomas in posttransplant lymphoproliferative disease (PTLD). Latent membrane protein 1 (LMP1), the major oncogenic protein of EBV, promotes tumorigenesis through activation of NF-kappaB, Erk, p38, JNK and Akt. The Jak/STAT signal transduction pathway is also constitutively active in PTLD-associated EBV(+) B-cell lymphomas. Here we determine the mechanism of Jak/STAT activation in EBV(+) B-cell lymphomas and the role of LMP1 in this process. Immunoprecipitation studies revealed no direct interaction of LMP1 and JAK3, but known associations between JAK3 and common gamma chain, and between LMP1 and TRAF3, were readily detected in EBV(+) B cell lines from patients with PTLD. An inducible LMP1 molecule expressed in EBV(-) BL41 Burkitt's cells demonstrated STAT activation only after prolonged LMP1 signaling. While LMP1 induced IFN-gamma production in BL41 cells, IFN-gamma receptor blockade and IFN-gamma neutralization prior to LMP1 activation markedly decreased STAT1 activation and expression of LMP1-driven IFN-gamma inducible genes. Understanding the mechanisms by which EBV induces cellular signal transduction pathways may facilitate development of new treatments for PTLD.

View details for DOI 10.1111/j.1600-6143.2009.02781.x

View details for Web of Science ID 000269801000013

View details for PubMedID 19656130
Natural Killer cell interactions with dendritic cells and tumor cells are mediated by different NK cell receptors Wai, L., Martinez, O. M., Krams, S. M. AMER ASSOC IMMUNOLOGISTS. 2009
View details for Web of Science ID 000209763603115
Insights into the Mechanism Responsible for Spontaneous Acceptance of Liver Allografts Zhuo, M., Ito, T., Wang, M., Esquivel, C. O., Martinez, O. M., Krams, S. M. AMER ASSOC IMMUNOLOGISTS. 2009
View details for Web of Science ID 000209763603136
Epstein-Barr Virus Latent Membrane Protein 1 Modulates Host MicroRNAs in B Cell Lymphomas Harris, A., Lambert, S., Krams, S., Martinez, O. M. AMER ASSOC IMMUNOLOGISTS. 2009
View details for Web of Science ID 000209763603146
Decreases in circulating CD4(+)CD25(hi)FOXP3(+) cells and increases in intragraft FOXP3(+) cells accompany allograft rejection in pediatric liver allograft recipients PEDIATRIC TRANSPLANTATION Stenard, F., Nguyen, C., Cox, K., Kambham, N., Umetsu, D. T., Krams, S. M., Esquivel, C. O., Martinez, O. M. 2009; 13 (1): 70-80
Abstract

We examined CD4(+)CD25(hi)FOXP3(+) cells Treg in children following liver transplantation and determined the relationship between Treg cell levels in the blood and in the graft. Peripheral blood was obtained from pediatric liver transplant patients at sequential time points: pre-transplant, one month, 3-4 months, 6-7 months, and 11-12 months post-transplant. PBMC were isolated, labeled for CD4, CD25 and FOXP3 expression and analyzed by flow cytometry for CD4(+)CD25(hi)FOXP3(+) cells. Sorted CD4(+)CD25(hi) cells were assessed for functional activity. Pretransplant blood levels of CD4(+)CD25(hi)FOXP3(+) Treg cells were not significantly different from post-transplant blood levels of CD4(+)CD25(hi)FOXP3(+) Treg cells. However, the blood levels of CD4(+)CD25(hi)FOXP3(+) Treg cells were significantly decreased during acute rejection compared with levels when graft function was stable. Immunohistochemistry revealed that FOXP3(+) cells were increased in the portal region of livers with histopathologic evidence of acute graft rejection compared with livers without evidence of rejection and were localized primarily within the inflammatory infiltrate. These data indicate that Treg cells are found at the site of allograft rejection and may play a role in regulation of alloreactivity. Moreover, monitoring peripheral CD4(+)CD25(hi)FOXP3(+) Treg cell levels may be useful in improving the post-transplant management of pediatric liver allograft recipients.

View details for DOI 10.1111/j.1399-3046.2008.00917.x

View details for PubMedID 18331536
Inflammation and the Innate Immune Response Impair Differentiation of Neural Progenitor Stem Cell Allografts. Phillips, L. K., Chen, Z., Palmer, T. D., Krams, S. M., Martinez, O. M. WILEY-BLACKWELL PUBLISHING, INC. 2009: 464
View details for Web of Science ID 000265068800963
Induced Tolerance to Liver Allografts Involves Apoptosis and the Generation of Regulatory T Cells Fujiki, M., Strober, S., Martinez, O., Esquivel, C., Krams, S. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2009: S76
View details for DOI 10.1016/j.clim.2009.03.223

View details for Web of Science ID 000266342300216
Insights into the Mechanism Responsible for Spontaneous Acceptance of Rat Liver Allografts. Ito, T., Wang, M., Zhuo, M., Esquivel, C. O., Martinez, O. M., Krams, S. WILEY-BLACKWELL PUBLISHING, INC. 2009: 631
View details for Web of Science ID 000265068801599
Natural Killer Cell Interactions with Dendritic Cells and Tumor Cells Are Mediated by Different NK Cell Receptors. Wai, L., Martinez, O. M., Krams, S. M. WILEY-BLACKWELL PUBLISHING, INC. 2009: 256
View details for Web of Science ID 000265068800226
Epstein-Barr Virus Latent Membrane Protein 1 Modulates Host MicroRNAs in B Cell Lymphomas Harris, A., Lambert, S., Krams, S., Martinez, O. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2009: S40
View details for DOI 10.1016/j.clim.2009.03.114

View details for Web of Science ID 000266342300109
Inflammation and the Innate Immune Response Impair Differentiation of Neural Progenitor Stem Cell Allografts Phillips, L., Chen, Z., Palmer, T., Krams, S., Martinez, O. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2009: S119
View details for DOI 10.1016/j.clim.2009.03.351

View details for Web of Science ID 000266342300343
Tumor-derived Variants of Epstein-Barr Virus Latent Membrane Protein 1 Induce Sustained Erk Activation and c-Fos JOURNAL OF BIOLOGICAL CHEMISTRY Vaysberg, M., Hatton, O., Lambert, S. L., Snow, A. L., Wong, B., Krams, S. M., Martinez, O. M. 2008; 283 (52): 36573-36585
Abstract

Latent membrane protein 1 (LMP1) of Epstein-Barr virus (EBV) is a proven oncogene that is essential for transformation of human B cells by the virus. LMP1 induces constitutive activation of several signal transduction pathways involving nuclear factor kappaB, phosphatidylinositol 3-kinase/Akt, and the mitogen-activated protein kinases (MAPK) p38, c-Jun N-terminal kinase (JNK), and extracellular signal-regulated kinase (Erk). Sequencing of LMP1 isolated from a panel of EBV+ B cell lymphomas identified three different variants of LMP1, each distinct from the B95.8 prototype isoform. All tumor variants of LMP1 as well as the B95.8 LMP1 isoform were able to induce rapid p38 phosphorylation as well as Akt and JNK activation. Additionally all variants showed similar ability to activate nuclear factor kappaB. In contrast, only tumor-derived LMP1 variants induced prolonged Erk activation and c-Fos expression. Sequence analysis revealed only two amino acids, 212 and 366, shared by the tumor variants but distinct from B95.8. Point mutation of either amino acids 212 (glycine to serine) or 366 (serine to threonine) from the B95.8 isoform to the tumor variant version of LMP1 was sufficient for gain of function characterized by sustained activation of Erk and subsequent c-Fos induction and binding to the AP1 site. Our results indicate that the enhanced ability of tumor-derived LMP1 to induce and stabilize the c-Fos oncogene can be localized to two amino acids in the C terminus of LMP1.

View details for DOI 10.1074/jbc.M802968200

View details for Web of Science ID 000261840500055

View details for PubMedID 18986987

View details for PubMedCentralID PMC2605991
Epstein-Barr virus, rapamycin, and host immune responses CURRENT OPINION IN ORGAN TRANSPLANTATION Krams, S. M., Martinez, O. M. 2008; 13 (6): 563-568
Abstract

To summarize recent advances that contribute to our understanding of the pathobiology of Epstein-Barr virus (EBV)-associated posttransplant lymphoproliferative disease (PTLD), the host immune response to virally infected B cells, and the molecular basis for the effects of mammalian target of rapamycin inhibitors on EBV+ B-cell lymphomas.Cytogenetic and genomic analyses support the concept that the underlying biology of EBV-associated PTLD is complex. Transplant recipients can generate and maintain significant populations of EBV-specific CD8+ memory T cells but the function of these cells may be impaired. EBV invokes multiple strategies to subvert and evade the host immune response. The phosphoinositide-3 kinase/Akt/mammalian target of rapamycin signal transduction pathway is a nexus for growth and survival signals in PTLD-associated EBV+ B-cell lymphomas.Multiple factors influence the development of EBV-associated PTLD including the host immune response to EBV, virally induced effects on the infected cell and the host immune system, and the type and intensity of immunosuppression.

View details for DOI 10.1097/MOT.0b013e3283186ba9

View details for Web of Science ID 000261755200001

View details for PubMedID 19060543

View details for PubMedCentralID PMC2650428
Molecular and immunologic mechanisms of cancer pathogenesis in solid organ transplant recipients AMERICAN JOURNAL OF TRANSPLANTATION Martinez, O. M., de Gruijl, F. R. 2008; 8 (11): 2205-2211
Abstract

The increased risk for the development of malignancies in transplant recipients is generally attributed to the debilitated immune system that results from chronic exposure to potent immunosuppressive drugs required to prevent graft rejection. While impaired immunity is clearly a key determinant, there is strong evidence that a constellation of other factors contribute to the pathogenesis of posttransplant cancers. In this article we discuss the underlying molecular and immunologic mechanisms that contribute to the development of de novo malignancies in transplant recipients, with particular focus on the two leading posttransplant neoplasia, skin cancer and Epstein-Barr virus (EBV)-associated posttransplant lymphoproliferative disorder (PTLD).

View details for DOI 10.1111/j.1600-6143.2008.02368.x

View details for Web of Science ID 000259937000008

View details for PubMedID 18801025
Syk drives Epstein Barr virus-infected B cell lymphoma growth and survival through activation of the PI3K/Akt pathway 8th American Transplant Congress Hatton, O., Lambert, S., Vaysberg, M., Sharman, J., Krams, S. M., Esquivel, C. O., Martinez, O. M. WILEY-BLACKWELL. 2008: 477?477
View details for Web of Science ID 000255763201545
PTLD derived tumor variants of EBV latent membrane protein 1 induce sustained ERK activation and cFos induction Vaysberg, M., Lambert, S. L., Krams, S. M., Martinez, O. M. WILEY-BLACKWELL. 2008: 308
View details for Web of Science ID 000255763200487
EBV-encoded LMP1 indirectly activates the Jak/STAT pathway through induction of IFN gamma Vaysberg, M., Lambert, S. L., Krams, S. M., Martinez, O. M. WILEY-BLACKWELL. 2008: 308
View details for Web of Science ID 000255763200486
Induction of the stimulatory receptor, NKG2D and its ligands after transplantation. Zhuo, M., Fujiki, M., Piard-Ruster, K., Martinez, O. M., Krams, S. M. BLACKWELL PUBLISHING. 2008: 493
View details for Web of Science ID 000255763201609
Investigating the role of toll-like receptor signaling in attenuating neurogenesis of Neural Stem/Progenitor Cells allografts in the adult hippocampus Phillips, L. K., Chen, Z., Carpentier, P., Palmer, T. D., Krams, S. M., Martinez, O. M. BLACKWELL PUBLISHING. 2008: 506
View details for Web of Science ID 000255763202049
EBV-encoded LMP1 indirectly activates the Jak/STAT pathway through production of IFN-gamma Vaysberg, M., Lambert, S. L., Krams, S. M., Martinez, O. M. FEDERATION AMER SOC EXP BIOL. 2008
View details for Web of Science ID 000208467808146
NK cell stimulatory receptors and ligands are induced post-transplantation Wai, L., Fujiki, M., Zhuo, M., Martinez, O. M., Krams, S. M. FEDERATION AMER SOC EXP BIOL. 2008
View details for Web of Science ID 000208467806788
Rapamycin, but not cyclosporine or FK506, alters natural killer cell function TRANSPLANTATION Wai, L., Fujiki, M., Takeda, S., Martinez, O. M., Krams, S. M. 2008; 85 (1): 145-149
Abstract

Infiltration of natural killer (NK) cells into solid organ allografts is observed in clinical and experimental transplantation. Studies suggest a role for NK cells in acute and chronic rejection of solid organ allografts; however, the effects of immunosuppressive agents on NK cells are not clearly established. Rat NK cell lines were analyzed for proliferation and cytotoxicity in the presence of cyclosporine, FK506, or rapamycin. Lewis recipients of DA liver allografts received immunosuppressive agents after transplantation. NK cells demonstrated robust function both in the absence and presence of cyclosporine and FK506. In contrast, rapamycin significantly inhibited proliferation and cytotoxicity of NK cells. NK cell numbers remained stable in graft recipients treated with cyclosporine and FK506, whereas there was a significant decrease in NK cells in rapamycin-treated recipients. These data indicate that immunosuppressive drugs have differential effects on NK cell function that may impact the immune response of transplant recipients.

View details for DOI 10.1097/01.tp.0000296817.28053.7b

View details for Web of Science ID 000252530600023

View details for PubMedID 18192925
Latent membrane protein 1 of EBV activates phosphatidylinositol 3-kinase to induce production of IL-10 JOURNAL OF IMMUNOLOGY Lambert, S. L., Martinez, O. M. 2007; 179 (12): 8225-8234
Abstract

EBV is a B lymphotrophic gamma-herpesvirus that is associated with multiple human malignancies, including posttransplant lymphoproliferative disorder. The EBV-encoded protein, latent membrane protein 1 (LMP1), is required for oncogenic transformation of human B cells by EBV. An important consequence of LMP1 expression in EBV-infected B cells is the induction of cellular IL-10, which acts as an autocrine growth factor for B cell lymphomas. However, the mechanisms by which LMP1 induces IL-10 are incompletely understood. We previously showed that rapamycin, a clinically relevant immunosuppressant and mammalian target of rapamycin inhibitor, could suppress IL-10 production by EBV-infected B cell lines. To test the hypothesis that PI3K, which acts upstream of mammalian target of rapamycin, might also be involved in LMP1-dependent IL-10 production, we generated B cell lines expressing signaling-inducible chimeric LMP1. Our results show that induced LMP1 signaling elicits both p38- and PI3K-dependent IL-10 production in EBV- B cells. Moreover, distinct regions of the LMP1 signaling tail are associated with p38- vs PI3K-dependent IL-10 induction. We also demonstrate that the LMP1-dependent p38 and PI3K activation regulates IL-10 induction through discrete mechanisms. Whereas p38 activation is critical for the phosphorylation of the transcription factor CREB, PI3K activation is required for the inactivation of glycogen synthase kinase 3beta (GSK3beta), an inhibitory kinase that can regulate CREB function. We find that GSK3beta regulates LMP1-dependent IL-10 induction, with GSK3beta inhibition by pharmacologic or small interfering RNA strategies enhancing LMP1-induced IL-10 induction. These findings demonstrate that LMP1 uses both p38 and PI3K activation for maximal up-regulation of IL-10.

View details for Web of Science ID 000251590000029

View details for PubMedID 18056366
Syk kinase inhibition by R406 inhibits BCR mediated B cell proliferation and interferes with BCR signal transduction in primary tumor samples 49th Annual Meeting of the American-Society-of-Hematology Sharman, J. P., Irish, J., Hatton, O., Martinez, O., Levy, R. AMER SOC HEMATOLOGY. 2007: 246A?246A
View details for Web of Science ID 000251100801071
Mutations to bid cleavage sites protect hepatocytes from apoptosis after ischemia/reperfusion injury TRANSPLANTATION Riddle-Taylor, E., Nagasaki, K., Lopez, J., Esquivel, C. O., Martinez, O. M., Krams, S. M. 2007; 84 (6): 778-785
Abstract

Apoptosis of hepatocytes contributes to many forms of liver pathology and can compromise liver function. Hepatocytes have been shown to require mitochondrial disruption to execute apoptosis, a process that is controlled by members of the Bcl-2 family. Bid is a proapoptotic Bcl-2 family member that is cleaved to its active form, tBid, by caspase 8 and granzyme B. Studies in the Bid-deficient mouse have established that hepatocytes require Bid to undergo apoptosis.We generated aspartic acid to glutamic acid mutations in the rat Bid protein, at the caspase 8 and granzyme B cleavage sites, and utilized recombinant adenoviruses to express this protein in hepatoma cells and in the livers of rats.Cells transduced with recombinant adenoviruses encoding Bid containing mutations to the caspase 8 and granzyme B cleavage sites are significantly protected from both tumor necrosis factor-alpha-induced and cell-mediated apoptosis. Protection occurs through a mechanism that includes decreased Bid cleavage, caspase activation, and mitochondrial membrane damage. Further, after warm ischemia/reperfusion injury, we show that rats expressing cleavage-resistant Bid in the liver display significantly less hepatocyte apoptosis as compared to control rat livers and this results in improved liver function and survival.Our results suggest that reagents that prevent the cleavage of Bid would be an effective strategy to inhibit hepatocyte apoptosis and decrease liver injury.

View details for DOI 10.1097/01.tp.0000281555.18782.2b

View details for Web of Science ID 000249841700017

View details for PubMedID 17893612
Peripheral regulatory T cells (CD4+CD25+FOXP3+) as a marker for rejection in pediatric liver transplant patients. 7th American Transplant Congress Stenard, F., Nguyen, C., Cox, K., Khambam, N., Krams, S., Esquivel, C., Martinez, O. WILEY-BLACKWELL. 2007: 278?279
View details for Web of Science ID 000246370200504
The effect of immunosuppressive agents on NK cell function. Wai, L., Fujiki, M., Takeda, S., Hsieh, C. L., Martinez, O. M., Krams, S. M. BLACKWELL PUBLISHING. 2007: 327
View details for Web of Science ID 000246370201127
EBV latent membrane protein 1 signals through PI3K survival pathways to induce IL-10 production in B cell lymphomas. Lambert, S. L., Martinez, O. M. WILEY-BLACKWELL PUBLISHING, INC. 2007: 334
View details for Web of Science ID 000246370201157
CD30 activation induces T cell apoptosis through the mitochondrial pathway. 7th American Transplant Congress Ruster, K. S., Hsieh, C. L., Krams, S. M., Martinez, O. M. WILEY-BLACKWELL. 2007: 334?334
View details for Web of Science ID 000246370201155
SYK is a novel target for treatment of Epstein Barr virus-infected B cell lymphomas in post-transplant lymphoproliferative disease. 7th American Transplant Congress Hatton, O., Vaysberg, M., Sharman, J., Krams, S. M., Esquivel, C. O., Martinez, O. M. WILEY-BLACKWELL. 2007: 317?318
View details for Web of Science ID 000246370201087
Mutations to bid cleavage sites protect hepatocytes from apoptosis following ischemia/reperfusion injury. 7th American Transplant Congress Riddle, E., Nagasaki, K., Stenard, F., Lopez, J., Martinez, O. M., Esquivel, C. O., Krams, S. M. WILEY-BLACKWELL. 2007: 435?435
View details for Web of Science ID 000246370201555
Rapamycin inhibits proliferation of Epstein-Barr virus-positive B-cell lymphomas through modulation of cell-cycle protein expression TRANSPLANTATION Vaysberg, M., Balatoni, C. E., Nepomuceno, R. R., Krams, S. M., Martinez, O. M. 2007; 83 (8): 1114-1121
Abstract

Posttransplant lymphoproliferative disease (PTLD) is a serious complication of solid organ and bone marrow transplantation and is closely associated with Epstein-Barr virus (EBV) infection. We have previously shown that rapamycin (RAPA) directly inhibits the in vitro and in vivo proliferation of EBV-infected B lymphoblastoid cell lines (SLCL), derived from patients with PTLD, by arresting cells in the G1 phase of the cell cycle. The aim of this study is to elucidate the mechanism by which RAPA causes cell cycle arrest in EBV+ B cells.SLCL were cultured without or with RAPA (10 ng/ml) and G1-associated cell cycle proteins were analyzed by immunoblot and densitometric analysis. CDK complexes were immunoprecipitated and incubated with retinoblastoma protein (Rb) substrate. Kinase activity of the complex was determined by Western blot with anti-phospho-Rb antibodies.We show that RAPA decreased both Cyclin D2 and Cyclin D3 protein levels. Furthermore, RAPA decreased the protein levels of cyclin dependent kinase 4 (CDK4) and increased the expression of the CDK inhibitor p27. In contrast, expression of the CDK inhibitor p21 was markedly inhibited by RAPA in the SLCL. Finally, in vitro kinase assays revealed that downstream hyperphosphorylation of Rb by CDK complexes was also decreased by RAPA.The results presented here elucidate key targets of RAPA-induced cell cycle arrest, provide insight into the growth pathways of EBV+ B-cell lymphomas, and demonstrate the potential for RAPA as a therapeutic option in the treatment of PTLD and other EBV+ lymphomas.

View details for DOI 10.1097/01.tp.0000260142.38619.9c

View details for Web of Science ID 000246234800017

View details for PubMedID 17452903
The effects of Immunosuppressive agents on NK cell function Wai, L., Fujiki, M., Takeda, S., Hsieh, C., Martinez, O. M., Krams, S. M. AMER ASSOC IMMUNOLOGISTS. 2007
View details for Web of Science ID 000209758203215
Latent Membrane Protein 1 of Epstein Barr Viruses Activates the PI3K Pathway to Induce Production of IL-10 Lambert, S. L., Martinez, O. M. AMER ASSOC IMMUNOLOGISTS. 2007
View details for Web of Science ID 000209758200278
Mutations to Bid cleavage sites protect hepatocytes from apoptosis following ischemia/reperfusion injury Krams, S. M., Nagasaki, K., Esquivel, C. O., Martinez, O. M., Riddle, E. AMER ASSOC IMMUNOLOGISTS. 2007
View details for Web of Science ID 000209758203227
Epstein-Barr virus: Evasive maneuvers in the development of PTLD AMERICAN JOURNAL OF TRANSPLANTATION Snow, A. L., Martinez, O. M. 2007; 7 (2): 271-277
Abstract

Epstein-Barr virus (EBV) infection is linked to approximately 90% of B-cell lymphomas associated with posttransplant lymphoproliferative disease (PTLD), a serious complication for immunosuppressed transplant recipients. In this paper, we review the myriad ways by which EBV can inadvertently drive the genesis and persistence of B-cell lymphomas, particularly when the antiviral immune response is compromised. Probing the basic mechanisms by which EBV infection proceeds and contributes to malignancy in such cases will hopefully improve our understanding and treatment of PTLD and other EBV-associated malignancies.

View details for DOI 10.1111/j.1600-6143.2006.01650.x

View details for Web of Science ID 000243440100003

View details for PubMedID 17229074
EBV can protect latently infected B cell lymphomas from death receptor-induced apoptosis JOURNAL OF IMMUNOLOGY Snow, A. L., Lambert, S. L., Natkunam, Y., Esquivel, C. O., Krams, S. M., Martinez, O. M. 2006; 177 (5): 3283-3293
Abstract

The relationship between EBV infection and sensitivity to death receptor (DR)-induced apoptosis is poorly understood. Using EBV- and EBV+ BJAB cells, we provide the first evidence that EBV can protect latently infected B cell lymphomas from apoptosis triggered through Fas or TRAIL receptors. Caspase 8 activation was impaired and cellular FLIP recruitment was enriched in death-inducing signaling complexes formed in EBV-infected BJAB cells relative to parent BJAB cells. Furthermore, latent membrane protein 1 expression alone could reduce caspase activation and confer partial resistance to DR apoptosis in BJAB cells. This protective effect was dependent on C-terminal activating region 2-driven NF-kappaB activation, which in turn up-regulated cellular FLIP expression in latent membrane protein 1+ BJAB cells. Thus, the ability of latent EBV to block DR apoptosis may help to ensure the survival of host cells during B cell differentiation, and contribute to the development of B cell lymphomas, especially in immunocompromised individuals.

View details for Web of Science ID 000240002800065

View details for PubMedID 16920969
NKp30 is a functional activation receptor on a subset of rat natural killer cells EUROPEAN JOURNAL OF IMMUNOLOGY Hsieh, C. L., Nagasaki, K., Martinez, O. M., Krams, S. M. 2006; 36 (8): 2170-2180
Abstract

NKp30 is a stimulatory receptor on human NK cells implicated in tumor immunity, and is capable of promoting or terminating dendritic cell maturation. To gain a better understanding of NKp30 biology, we have investigated the expression and function of rat NKp30 (rNKp30). We generated stable transfectants of rNKp30 in RNK16 cells, a rat NK lymphoma line, and used a novel panel of mAb against rNKp30 to study this receptor. Using agonistic rNKp30 mAb, we demonstrated that rNKp30 mediates robust IFN-gamma production and cytolytic responses from rNKp30-transfected RNK16 cells. We determined by flow cytometry that rNKp30 is expressed by a subset of primary NK cells isolated from the blood and spleen, and to a lesser extent also on liver NK cells. Stimulation of rNKp30 on primary NK cells led to IFN-gamma production. Liver NK cells expressed low levels of NKp30 and had reduced rNKp30-mediated IFN-gamma responses. During an alloimmune response in vivo, the proportion of the rNKp30(+) NK cell subset in the peripheral blood significantly increased, suggesting that rNKp30 may play an important role during alloactivation. Thus, our data demonstrate that NKp30 is indeed expressed in rodents and is a functional stimulatory receptor in a subset of rat NK cells.

View details for DOI 10.1002/eji.200635982

View details for Web of Science ID 000239855900017

View details for PubMedID 16821237
Reproduction and pregnancy in transplant recipients: current practices. Progress in transplantation McKay, D. B., Adams, P. L., Bumgardner, G. L., Davis, C. L., Fine, R. N., Krams, S. M., Martinez, O. M., Murphy, B., Pavlakis, M., Tolkoff-Rubin, N., Sherman, M. S., Josephson, M. A. 2006; 16 (2): 127-132
Abstract

Many transplant physicians are faced with questions from their patients about the safety and long-term consequences of pregnancy following transplantation. To better understand how pregnancies are managed and to clarify the outcome of pregnancy after transplantation, a survey questionnaire was developed and mailed to all medical and surgical directors of transplant centers throughout the United States; responses were obtained from 59.1% of the transplant centers. Although many opinions were collected, most respondents conceded that their opinions were based on personal experience rather than evidence-based. The underutilization of existing information was revealing and highlighted a need for an evidence-based approach to care of the pregnant transplant recipient and her offspring. The survey results, reported in this article, led to formation of a consensus conference to determine the optimal approach to pregnant transplant recipients and to define what is currently known and unknown about reproduction and transplantation.

View details for PubMedID 16789701
EBV+ B lymphoma cell lines from patients with post-transplant lymphoproliferative disease are resistant to TRAIL-induced apoptosis AMERICAN JOURNAL OF TRANSPLANTATION Snow, A. L., Vaysberg, M., Krams, S. M., Martinez, O. M. 2006; 6 (5): 976-985
Abstract

Lymphomas associated with post-transplant lymphoproliferative disease (PTLD) represent a significant complication of immunosuppression in transplant recipients. In immunocompetent individuals, EBV-specific cytotoxic T lymphocytes (CTL) prevent the outgrowth of activated B lymphoblasts through apoptosis induction. Soluble versions of TNF-related apoptosis-inducing ligand/Apo2 ligand (TRAIL) can induce apoptosis in numerous tumor cell types. Given the therapeutic potential of TRAIL, we examined the sensitivity of EBV+ spontaneous lymphoblastoid cell lines (SLCL) derived from patients with PTLD to treatment with soluble TRAIL. Despite abundant expression of TRAIL receptors (TRAIL-R), resistance to TRAIL-induced apoptosis was observed in all SLCL examined. This resistance could not be overcome by concomitant treatment with several pharmacological agents. Unlike BJAB positive control cells, for each SLCL tested, cleavage and activation of caspase 8 was inhibited due to failed recruitment of FADD and caspase 8 to TRAIL receptors upon stimulation. Further indicative of a proximal defect, TRAIL receptor aggregation could not be detected on the cell surface of SLCL following ligand engagement. These results suggest that the use of TRAIL for eliminating PTLD-associated tumors may be of limited clinical utility, and illustrate another mechanism by which EBV+ B lymphoma cells can evade tumor surveillance at the level of death receptor signaling.

View details for DOI 10.1111/j.1600-6143.2006.01295.x

View details for Web of Science ID 000236860700014

View details for PubMedID 16611333
CD72 down-modulates BCR-induced signal transduction and diminishes survival in primary mature B lymphocytes JOURNAL OF IMMUNOLOGY Li, D. H., Tung, J. W., Tarner, I. H., Snow, A. L., Yukinari, T., Ngernmaneepothong, R., Martinez, O. M., Parnes, J. R. 2006; 176 (9): 5321-5328
Abstract

CD72, a 45-kDa type II transmembrane glycoprotein carrying an ITIM motif, is believed to be an inhibitory coreceptor of the BCR. Mature B cells lacking CD72 show enhanced Ca(2+) mobilization and are hyperproliferative in response to BCR ligation. However, the signal transduction pathways downstream of BCR signaling that transmit the inhibitory effect of CD72 in mature B cells remain unknown. To address this question, we used hen egg lysozyme-specific BCR transgenic mice to elucidate the differential cell signaling between wild-type and CD72-deficient B cells in response to hen egg lysozyme Ag stimulation. Our results demonstrate that CD72 predominantly down-regulates the major signal transduction pathways downstream of the BCR, including NF-AT, NF-kappaB, ERK, JNK, p38-MAPK, and PI3K/Akt in mature B cells. CD72 ligation with anti-CD72 Ab (K10.6), which mimics the binding of CD100 (a natural ligand for CD72) to release the inhibitory function of CD72, augments cell proliferation, Ca(2+) flux, IkappaBalpha activation, and ERK MAPK activity upon Ag stimulation in wild-type B cells. In addition, we show direct evidence that CD72 promotes cell cycle arrest and apoptosis after Ag stimulation in mature B cells. Taken together, our findings conclude that CD72 plays a dominant role as a negative regulator of BCR signaling in primary mature B lymphocytes.

View details for Web of Science ID 000238768700028

View details for PubMedID 16621999
Latent membrane protein I of Epstein-Barr virus signals for IL-10 production in human B cell lymphomas Lambert, S., Martinez, O. M. AMER ASSOC IMMUNOLOGISTS. 2006: S167
View details for Web of Science ID 000238837101267
IL-10 induction by latent membrane protein 1 of Epstein Barr virus. Lambert, S., Martinez, O. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2006: S98?S99
View details for DOI 10.1016/j.clim.2006.04.176

View details for Web of Science ID 000237924300258
Abrogation of bid cleavage protects rat hepatocytes from apoptosis Riddle, E., Nagasaki, K., Martinez, O., Krams, S. JOHN WILEY & SONS INC. 2005: 502A
View details for Web of Science ID 000232480301321
IFN-gamma, produced by NK cells that infiltrate liver allografts early after transplantation, links the innate and adaptive immune responses AMERICAN JOURNAL OF TRANSPLANTATION Obara, H., Nagasaki, K., Hsieh, C. L., Ogura, Y., Esquivel, C. O., Martinez, O. M., Krams, S. M. 2005; 5 (9): 2094-2103
Abstract

The role of NK cells following solid organ transplantation remains unclear. We examined NK cells in acute allograft rejection using a high responder model (DA-->Lewis) of rat orthotopic liver transplantation. Recipient-derived NK cells infiltrated liver allografts early after transplantation. Since chemokines are important in the trafficking of cells to areas of inflammation, we determined the intragraft expression of chemokines known to attract NK cells. CCL3 was significantly increased in allografts at 6 h post-transplant as compared to syngeneic grafts whereas CCL2 and CXCL10 were elevated in both syngeneic and allogeneic grafts. CXCL10 and CX3CL1 were significantly upregulated in allografts by day 3 post-transplant as compared to syngeneic grafts suggesting a role for these chemokines in the recruitment of effector cells to allografts. Graft-infiltrating NK cells were shown to be a major source of IFN-gamma, and IFN-gamma levels in the serum were markedly increased, specifically in allograft recipients, by day 3 post-transplant. Accordingly, in the absence of NK cells the levels of IFN-gamma were significantly decreased. Furthermore, graft survival was significantly prolonged. These data suggest that IFN-gamma-producing NK cells are an important link between the innate and adaptive immune responses early after transplantation.

View details for DOI 10.1111/j.1600-6143.2005.00995.x

View details for Web of Science ID 000231023700003

View details for PubMedID 16095488

View details for PubMedCentralID PMC1473982
Reproduction and transplantation: Report on the AST consensus conference on reproductive issues and transplantation AMERICAN JOURNAL OF TRANSPLANTATION McKay, D. B., Josephson, M. A. 2005; 5 (7): 1592-1599
Abstract

It has been almost 50 years since the first child was born to a female transplant recipient. Since that time pregnancy has become common after transplantation, but physicians have been left to rely on case reports, small series and data from voluntary registries to guide the care of their patients. Many uncertainties exist including the risks that pregnancy presents to the graft, the patient herself, and the long-term risks to the fetus. It is also unclear how to best modify immunosuppressive agents or treat rejection during pregnancy, especially in light of newer agents available where pregnancy safety has not been established. To begin to address uncertainties and define clinical practice guidelines for the transplant physician and obstetrical caregivers, a consensus conference was held in Bethesda, Md. The conferees summarized both what is known and important gaps in our knowledge. They also identified key areas of agreement, and posed a number of critical questions, the resolution of which is necessary in order to establish evidence-based guidelines. The manuscript summarizes the deliberations and conclusions of the conference as well as specific recommendations based on current knowledge in the field.

View details for DOI 10.1111/j.1600-6143.2005.00969.x

View details for Web of Science ID 000229635100004

View details for PubMedID 15943616
Ifn-gamma, produced by NK cells recruited to liver allografts early after transplantation, links the innate and adaptive immune responses, produced by NK cells recruited to liver allografts early after transplantation, links the innate and adaptive immune responses. 6th American Transplant Congress Nagasaki, K., Obara, H., Hsieh, C., Esquivel, C. O., Martinez, O. M., Krams, S. M. WILEY-BLACKWELL. 2005: 434?434
View details for Web of Science ID 000229231601505
Prolonged graft survival and decreased expression of IFN-gamma in the absence of NK cells 6th American Transplant Congress Hsieh, C. L., Ohara, H., Nagasaki, K., Martinez, O. M., Krams, S. M. WILEY-BLACKWELL. 2005: 574?574
View details for Web of Science ID 000229231602457
Basic concepts in transplant immunology LIVER TRANSPLANTATION Martinez, O. M., Rosen, H. R. 2005; 11 (4): 370-381
View details for DOI 10.1002/lt.20406

View details for PubMedID 15776458
Mutations to the proapoptotic molecule, bid, protect Hepatocytes from apoptosis Riddle, E. D., Nagasaki, K., Martinez, O., Krams, S. FEDERATION AMER SOC EXP BIOL. 2005: A1438
View details for Web of Science ID 000227610903177
Latent membrane protein 1 of Epstein-Barr virus affects endogenous signaling pathways in human B cells Lambert, S., Martinez, O. FEDERATION AMER SOC EXP BIOL. 2005: A957?A958
View details for Web of Science ID 000227610706601
Identification, cloning, and characterization of a novel rat natural killer receptor, RNKp30: A molecule expressed in liver allografts TRANSPLANTATION Hsieh, C. L., Ogura, Y., Obara, H., Ali, U. A., Rodriguez, G. M., Nepomuceno, R. R., Martinez, O. M., Krams, S. M. 2004; 77 (1): 121-128
Abstract

As a component of the innate immune system, natural killer (NK) cells may play a significant role in the early events after solid-organ transplantation. Activated NK cells have been shown to infiltrate allografts in transplant models. To better understand NK cells and the role of NK cell receptors in transplantation, we have cloned and begun characterizing a novel rat molecule, rNKp30.RNKp30 cDNA was cloned by 5' rapid amplification of cDNA ends polymerase chain reaction (PCR) and reverse transcriptase (RT)-PCR from mononuclear cells infiltrating a rejecting liver allograft. Southern blot analysis was used to determine the rNKp30 gene copy number. RT-PCR and Northern blotting were used to examine rNKp30 RNA expression in NK cells, multiple tissues, and liver grafts. Immunocytochemistry, immunoprecipitation, and Western blot analysis with two anti-rNKp30 polyclonal antibodies, CA680 and CA1071, were performed. Tunicamycin and endoglycosidase treatments determined the extent of rNKp30 glycosylation.RNKp30 is homologous to human and macaque NKp30. It is a single copy gene with five identified single-nucleotide polymorphisms. RNKp30 is expressed by NK cells and is detectable as a single transcript by Northern blot in normal spleen, lymph node, and lung tissues. RNKp30 is a variably N-glycosylated cell surface molecule with a protein backbone of approximately 21 kDa. Elevated transcript expression of rNKp30 is detected in both rejected and spontaneously accepted liver allografts, but not in syngeneic or cyclosporine A-treated allografts.RNKp30 is a glycosylated surface NK cell receptor with limited polymorphism. This putative activation receptor is expressed in liver allografts and may participate in the innate immune response after transplantation.

View details for DOI 10.1097/01.TP.0000110423.27977.6F

View details for PubMedID 14724446
NK cells recruited to liver allografts are a source of IFN-gamma. 5th American Transplant Congress Obara, H., Martinez, O. M., Ogura, Y., Hsieh, C. L., Esquivel, C. O., Krams, S. M. WILEY-BLACKWELL. 2004: 249?249
View details for Web of Science ID 000221322500334
Rapamycin inhibits NK cell function. 5th American Transplant Congress Takeda, S., Hsieh, C., Ohara, H., Martinez, O. M., Krams, S. M. WILEY-BLACKWELL. 2004: 526?526
View details for Web of Science ID 000221322501339
NK receptor, rat NKp30, is a polymorphic glycoprotein involved in the immune response during allogeneic liver transplantation. 5th American Transplant Congress Hsieh, C. L., Ogura, Y., Obara, H., Ali, U., Su, W. W., Rodriguez, G., Nepomuceno, R., Martinez, O. M., Krams, S. M. WILEY-BLACKWELL. 2004: 510?510
View details for Web of Science ID 000221322501283
Rapamycin inhibits the interleukin 10 signal transduction pathway and the growth of Epstein Barr virus B-cell lymphomas CANCER RESEARCH Nepomuceno, R. R., Balatoni, C. E., Natkunam, Y., Snow, A. L., Krams, S. M., Martinez, O. M. 2003; 63 (15): 4472-4480
Abstract

EBV-infected B-cell lymphomas are a potentially life-threatening complication in bone marrow and solid organ transplant recipients. Immunosuppressive drugs required to prevent allograft rejection also impair anti-EBV T-cell immunity, thereby increasing the risk of EBV-associated disease. Here we demonstrate that the immunosuppressant rapamycin (RAPA) has a strong antiproliferative effect in vitro on B-cell lines derived from organ transplant recipients with EBV-associated posttransplant lymphoproliferative disorder (PTLD). Furthermore, RAPA significantly inhibits or delays the growth of solid tumors established from EBV-infected B-cell lines in a xenogeneic mouse model of PTLD. RAPA acts via cell cycle arrest, induction of apoptosis, and, most importantly, inhibition of interleukin 10 secretion, a necessary autocrine growth factor. The reduced interleukin 10 production is accompanied by corresponding decreases in the constitutive activation of the growth-promoting transcription factors signal transducer and activator of transcription 1 and 3. Thus, RAPA can limit B-cell lymphoma growth while simultaneously providing immunosuppression to prevent graft rejection in patients who are otherwise at risk for EBV-associated PTLD. Moreover, these findings may have application to other EBV-associated malignancies.

View details for Web of Science ID 000184562500028

View details for PubMedID 12907620
Depletion of an asialo GM1 cell population prolongs graft survival and leads to donor specific tolerance 90th Annual Meeting of the American-Association-for-Immunologists Obara, H., Hsieh, C. L., Martinez, O. M., Krams, S. M. FEDERATION AMER SOC EXP BIOL. 2003: C112?C112
View details for Web of Science ID 000182367000520
Bid cleavage in liver allograft rejection and hepatocyte apoptosis Riddle, E. D., Obara, H., Martinez, O. M., Krams, S. M. FEDERATION AMER SOC EXP BIOL. 2003: C112?C113
View details for Web of Science ID 000182367000521
Sirolimus inhibits, cell growth and the expression of cell cycle proteins in Epstein-Barr virus-associated B cell lymphomas Balatoni, C., Nepomuceno, R. N., Krams, S. M., Martinez, O. M. FEDERATION AMER SOC EXP BIOL. 2003: C116
View details for Web of Science ID 000182367000535
Epstein barr virus can protect latently infected B cell lymphomas from FasL- and TRAIL-mediated apoptosis Snow, A. L., Krams, S. M., Martinez, O. M. FEDERATION AMER SOC EXP BIOL. 2003: C325
View details for Web of Science ID 000182367001518
Rapamycin Inhibits the Interleukin 10 Signal Transduction Pathway and the Growth of Epstein Barr Virus B-cell Lymphomas Cancer Research Nepomuceno RR, Balatoni CE, Natkunam Y, Snow AL, Krams SM, Martinez OM 2003; 63 (15): 4472
Identification of Epstein-Barr virus-specific CD8(+) T lymphocytes in the circulation of pediatric transplant recipients TRANSPLANTATION Falco, D. A., Nepomuceno, R. R., Krams, S. M., Lee, P. P., DAVIS, M. M., Salvatierra, O., Alexander, S. R., Esquivel, C. O., Cox, K. L., Frankel, L. R., Martinez, O. M. 2002; 74 (4): 501-510
Abstract

Pediatric transplant recipients are at increased risk for Epstein Barr virus (EBV)-related B cell lymphomas. In healthy individuals, the expansion of EBV-infected B cells is controlled by CD8+ cytotoxic T cells. However, immunosuppressive therapy may compromise antiviral immunity. We identified and determined the frequency of EBV-specific T cells in the peripheral blood of pediatric transplant recipients.HLA-B*0801 and HLA-A*0201 tetramers folded with immunodominant EBV peptides were used to detect EBV-specific CD8+ T cells by flow cytometry in peripheral blood mononuclear cells from 24 pediatric liver and kidney transplant recipients. The expression of CD38 and CD45RO on EBV-specific, tetramer-binding cells was also examined in a subset of patients by immunofluorescent staining and flow cytometry.Tetramer-binding CD8+ T cells were identified in 21 of 24 transplant recipients. EBV-specific CD8+ T cells were detected as early as 4 weeks after transplant in EBV seronegative patients receiving an organ from an EBV seropositive donor. The frequencies (expressed as a percentage of the CD8+ T cells) of the tetramer-binding cells were HLA-B8-RAKFKQLL (BZLF1 lytic antigen peptide) tetramer, range=0.96 to 3.94%; HLA-B8-FLRGRAYGL (EBNA3A latent antigen peptide) tetramer, range=0.03 to 0.59%; and HLA-A2-GLCTLVAML (BMLF1 lytic antigen peptide) tetramer, range=0.06 to 0.76%. The majority of tetramer reactive cells displayed an activated/memory phenotype.Pediatric transplant recipients receiving immunosuppression can generate EBV-specific CD8+ T cells. Phenotypic and functional analysis of tetramer cells may prove useful in defining and monitoring EBV infection in the posttransplant patient.

View details for PubMedID 12352909
Constitutive activation of Jak/STAT proteins in Epstein-Barr virus-infected B-cell lines from patients with posttransplant lymphoproliferative disorder. TRANSPLANTATION Nepomuceno, R. R., Snow, A. L., Beatty, P. R., Krams, S. M., Martinez, O. M. 2002; 74 (3): 396-402
Abstract

Posttransplant lymphoproliferative disease (PTLD) is a major complication after bone marrow and solid organ transplantation. The disease encompasses a spectrum of abnormal, Epstein-Barr virus (EBV)-associated B-cell proliferations. We have previously shown that EBV-infected, spontaneous lymphoblastoid cell lines (SLCL) derived from PTLD patients require autocrine interleukin (IL)-10 to proliferate. To determine if cytokine signal transduction is involved in the autonomous growth of the SLCL, the activation states of the Jak/STAT signaling pathway proteins were analyzed in three different SLCL, termed JB7, MF4, and VB4.The tyrosine phosphorylation (P-tyr) states of the Janus kinases (Jaks) and signal transducers and activators of transcription (STAT) proteins were examined by immunoprecipitation and immunoblot. Activated STAT dimer formation was determined by electromobility shift assays.All three SLCL, but not the Daudi Burkitt's lymphoma B-cell line, expressed the four known Jak kinases constitutively tyrosine phosphorylated, with particularly high levels of P-tyr Jak1 in the JB7 line. STAT1 and STAT3, but not STAT2 or STAT5, are also constitutively activated in all SLCL. The ability of the activated STAT proteins to form DNA-binding dimers was confirmed by electromobility shift assay. The SLCL, but not the Daudi line, express activated STAT complexes composed of STAT1 and STAT3. Another EBV-infected B-cell line, isolated from a lymph node biopsy after kidney transplantation, is phenotypically similar to the other SLCL in both surface antigen and activated STAT1 and STAT3 expression.These data support the presence of a constitutively active autocrine signaling pathway consistent with IL-10 in the SLCL.

View details for Web of Science ID 000177496000017

View details for PubMedID 12177620
CD30 expression identifies the predominant proliferating T lymphocyte population in human alloimmune responses JOURNAL OF IMMUNOLOGY Chan, K. W., Hopke, C. D., Krams, S. M., Martinez, O. M. 2002; 169 (4): 1784-1791
Abstract

CD30 is an inducible member of the TNFR superfamily that is expressed on activated T and B cells and some lymphoid malignancies. We have previously shown that human CD30(+) T cells elicited with allogeneic APC are a major source of IFN-gamma and IL-5 production. In the present study we have used alloantigen, as well as anti-CD3 plus anti-CD28 mAb stimulation, to further characterize human CD30(+) T cells with respect to function and the expression of other activation-dependent cell surface molecules, including the related TNFR family members OX-40 and 4-1BB (CD137). Our results indicate that human CD30(+) T cells are a subset of activated T cells that also express CD25 and CD45RO. Moreover, we observed that allogeneic APC consistently induced a greater proportion of CD30(+) cells within the activated T cell population than did stimulation with plate-bound anti-CD3 plus anti-CD28 mAb or stimulation with soluble anti-CD3 plus anti-CD28 and autologous APC. The enhanced induction of CD30 expression by alloantigen was not common to other inducible TNFR family members because anti-CD3 plus anti-CD28 mAbs were far more effective in inducing expression of 4-1BB and OX-40. Furthermore, CD30 expression marked the predominant proliferating T cell population induced by alloantigen as determined by CFSE staining and flow cytometry. These results indicate that CD30, but not 4-1BB or OX-40, is preferentially induced by alloantigen, suggesting that CD30 may be important in human alloimmune responses.

View details for Web of Science ID 000177365800017

View details for PubMedID 12165500
NK cells and transplantation TRANSPLANT IMMUNOLOGY Hsieh, C. L., Obara, H., Ogura, Y., Martinez, O. M., Krams, S. M. 2002; 9 (2-4): 111-114
Abstract

The requirement for cytotoxic T lymphocytes during allograft rejection is controversial. We have demonstrated that CD8+ T cells are not essential for allograft rejection or for the induction of apoptosis in two experimental models of transplantation. To determine candidate cells types which may play a role in the events leading to graft rejection, the cellular composition of rejecting allografts was determined. We demonstrate that substantial numbers of NK cells, of recipient origin, infiltrate allografts as early as 12 h after transplantation. These NK cells produce cytokines and express cytotoxic mediators such as granzyme B and FasL. It is unknown which NK cell receptors are expressed and activated during transplantation. NK cells express multiple cell surface receptors, including MHC class I binding inhibitory receptors, which deliver a negative signal, and activation receptors, which stimulate cytokine secretion and cytotoxicity of NK cells. To begin to understand NK cell activation in the context of transplantation, we have recently cloned a novel rat immunoglobulin-like surface receptor from a rejecting liver allograft. Sequence analysis demonstrates that this putative activation receptor contains 71% identity to human NKp30 at the DNA level, suggesting that it is the rat homologue (rNKp30). Characterization of NK activation receptors may lead to better understanding of the interactions between the innate and adaptive immune responses in transplantation.

View details for Web of Science ID 000177317300007

View details for PubMedID 12180816
Cloning of a novel NK receptor, rat NKp30, from liver allografts and characterization of its expression Hsieh, C. L., Ogura, Y., Obara, H., Martinez, O., Krams, S. FEDERATION AMER SOC EXP BIOL. 2002: A1034?A1034
View details for Web of Science ID 000174593901701
Resistance to Fas and TRAIL-mediated apoptosis in PTLD-associated EBV plus B cell lymphomas Snow, A. L., Chen, L. J., Nepomuceno, R. R., Krams, S., Martinez, O. FEDERATION AMER SOC EXP BIOL. 2002: A1029
View details for Web of Science ID 000174593901678
Epstein-Barr virus infection is associated with endothelial bcl-2 expression in transplant liver allografts TRANSPLANTATION Millan, M. T., Natkunam, Y., Clarke-Katzenberg, R., Desai, D., Prapong, W., So, S. K., Esquivel, C. O., Sibley, R., Ferran, C., Martinez, O. M. 2002; 73 (3): 465-469
Abstract

In liver transplant recipients with Epstein-Barr virus (EBV) disease, we reported a low rate of acute rejection after stopping or markedly lowering immunosuppression. This observation led to the hypothesis that EBV, as a means of viral persistence, induces expression of antiapoptotic factors and these factors, in turn, confer protection to the transplanted organ. Bcl-2, an antiapoptotic factor induced by EBV in various host cells, is not normally expressed in the liver. We questioned whether bcl-2 is expressed in the transplanted liver and whether its expression is modified by EBV.Retrospective liver biopsy specimen from liver transplant patients diagnosed with EBV (n=12) were examined for the presence of bcl-2 by immunohistochemistry and compared with EBV (-) transplant (n=15), and nontransplant (n=13) livers.The most significant finding was the presence of endothelial bcl-2 expression in the majority of EBV (+) transplant samples examined (67%) and its relative absence in the other two groups (P<0.005). There was also bcl-2 expression in the hepatocytes and lymphocytes of the majority of transplant liver samples, irrespective of EBV status.We have identified a strong association between EBV infection and endothelial bcl-2 expression in transplant livers. We also found that transplantation, in itself, was associated with bcl-2 expression in the hepatocytes and lymphocytes of liver allografts.

View details for Web of Science ID 000174115400023

View details for PubMedID 11884946
Resistance to Fas-mediated apoptosis in EBV-infected B cell lymphomas is due to defects in the proximal Fas signaling pathway JOURNAL OF IMMUNOLOGY Snow, A. L., Chen, L. J., Nepomuceno, R. R., Krams, S. M., Esquivel, C. O., Martinez, O. M. 2001; 167 (9): 5404-5411
Abstract

Post-transplant lymphoproliferative disorder is characterized by the outgrowth of EBV-infected B cell lymphomas in immunosuppressed transplant recipients. Using a panel of EBV-infected spontaneous lymphoblastoid cell lines (SLCL) derived from post-transplant lymphoproliferative disorder patients, we assessed the sensitivity of such lymphomas to Fas-mediated cell death. Treatment with either an agonist anti-Fas mAb or Fas ligand-expressing cells identifies two subsets of SLCL based on their sensitivity or resistance to Fas-driven apoptosis. Fas resistance in these cells cannot be attributed to reduced Fas expression or to mutations in the Fas molecule itself. In addition, all SLCL are sensitive to staurosporine-induced cell death, indicating that there is no global defect in apoptosis. Although all SLCL express comparable levels of Fas signaling molecules including Fas-associated death domain protein, caspase 8, and caspase 3, Fas-resistant SLCL exhibit a block in Fas-signaling before caspase 3 activation. In two SLCL, this block results in impaired assembly of the death-inducing signaling complex, resulting in reduced caspase 8 activation. In a third Fas-resistant SLCL, caspase 3 activation is hindered despite intact death-inducing signaling complex formation and caspase 8 activation. Whereas multiple mechanisms exist by which tumor cells can evade Fas-mediated apoptosis, these studies suggest that the proximal Fas-signaling pathway is impeded in Fas-resistant post-transplant lymphoproliferative disorder-associated EBV(+) B cell lymphomas.

View details for Web of Science ID 000171858500080

View details for PubMedID 11673559
Apoptotic pathways in primary biliary cirrhosis and autoimmune hepatitis LIVER Fox, C. K., Furtwaengler, A., Nepomuceno, R. R., Martinez, O. M., Krams, S. M. 2001; 21 (4): 272-279
Abstract

Autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC) are two autoimmune diseases with unknown etiologies that primarily target the liver. In both diseases, liver lesions are accompanied by large infiltrates of mononuclear cells. The purpose of this study was to determine if either the Fas-mediated or the granule-exocytosis pathways contribute to apoptosis in these diseases.To determine the involvement of apoptosis in tissue injury we examined liver tissue for DNA fragmentation and morphological characteristics of apoptosis. The major cytotoxic pathways of activated lymphocytes were compared by quantitating the levels of transcripts for FasL and granzyme B, and expression was confirmed by immunoprecipitation of these molecules.In both diseases, apoptosis was observed. However, the main cell types undergoing apoptosis were hepatocytes in AIH, and biliary epithelial cells in PBC. In AIH the levels of FasL and granzyme B mRNA were increased over the levels detected in normal liver, while in PBC only the levels of granzyme B were elevated. Additionally, in AIH, the ratio of FasL transcripts to granzyme B transcripts was elevated, reflecting a possible increase in the relative contribution of FasL to the progression of the disease. Immunoprecipitation studies further support an increase in FasL protein expression in AIH.These data suggest that both FasL and granzyme B contribute to the apoptosis observed in AIH and PBC. However, FasL appears to play a more prominent role in the induction of hepatocyte apoptosis and tissue destruction in AIH.

View details for Web of Science ID 000170155900008

View details for PubMedID 11454191
Apoptosis and allograft rejection in the absence of CD8(+) T cells TRANSPLANTATION Ogura, Y., Martinez, O. M., Villanueva, J. C., Tait, J. F., Strauss, H. W., Higgins, J. P., Tanaka, K., Esquivel, C. O., Blankenberg, F. G., Krams, S. M. 2001; 71 (12): 1827-1834
Abstract

The requirement for cytotoxic T lymphocytes during allograft rejection is controversial. We previously demonstrated that CD8+ T cells are not necessary for allograft rejection or for the induction of apoptosis in rat small intestinal transplantation. In this study, we examined the mechanisms of apoptosis and rejection after liver transplantation in the absence of CD8+ T cells.Either Lewis or dark agouti rat liver grafts were transplanted into Lewis recipients to create syngeneic and allogeneic combinations. CD8+ T cells were depleted in an additional allogeneic group by treatment with OX-8 mAb on day -1 and day 1 after liver transplant.Apoptosis and rejection were observed in both the CD8+ T cell-depleted allogeneic and allogeneic grafts by hematoxylin and eosin staining, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling staining, and radiolabeled-annexin V in vivo imaging. Granzyme B and FasL were expressed in all allogeneic transplants, including those depleted of CD8+ T cells, indicating that a mononuclear cell other than a CD8+ T cell can be the source of these molecules during allograft rejection. Activation of the caspase cascade was detected in all rejecting allografts. Caspases 3, 8, and 9 were activated at similar significantly elevated levels in both allogeneic and CD8+ T cell-depleted liver grafts.These data indicate that in the absence of CD8+ T cells an alternative pathway, associated with granzyme B and FasL expression and activation of the caspase cascade, can mediate apoptosis and graft rejection.

View details for Web of Science ID 000169753800020

View details for PubMedID 11455265
Activated CD30+T cells proliferate in response to alloantigen and infiltrate rejecting allografts. Chan, K. W., Nepomuceno, R. R., Ogura, Y., Silva, R., Krams, S. M., Martinez, O. M. LIPPINCOTT WILLIAMS & WILKINS. 2000: S340?S340
View details for Web of Science ID 000086911800877
Radiolabeled annexin V imaging: Diagnosis of allograft rejection in an experimental rodent model of liver transplantation RADIOLOGY Ogura, Y., Krams, S. M., Martinez, O. M., Kopiwoda, S., Higgins, J. P., Esquivel, C. O., Strauss, H. W., Tait, J. F., Blankenberg, F. G. 2000; 214 (3): 795-800
Abstract

To assess the value of imaging rejection-induced apoptosis with technetium 99m and annexin V, a human protein-based radiopharmaceutical used in the diagnosis of acute rejection of a liver transplant, in a well-characterized rodent model of orthotopic liver transplantation.99mTc-radiolabeled annexin V was intravenously administered to six allografted (immunologically mismatched) and five isografted (immunologically matched) recipient rats on days 2, 4, and 7 after orthotopic liver transplantation. Animals were imaged 1 hour after injection of 0.2-2.0 mCi (8.0-74.0 MBq) of radiolabeled annexin V by use of clinical nuclear scintigraphic equipment.All animals in the allografted group demonstrated marked increases of 55% and 97% above the activity in the isografted group in hepatic uptake of annexin V on days 4 and 7, respectively. Severe acute rejection was histologically detected in all allografted livers on day 7. There was no histologic evidence of acute rejection in isografted animals. Dynamic hepatobiliary imaging with 99mTc and mebrofenin, an iminodiacetic acid derivative, demonstrated no correlation with the presence or absence of acute rejection or with annexin V uptake.Noninvasive imaging with radiolabeled annexin V is more sensitive and specific than imaging with 99mTc-mebrofenin in the diagnosis of acute rejection of a liver transplant.

View details for Web of Science ID 000085478800029

View details for PubMedID 10715048
Significance of detecting Epstein-Barr-Specific sequences in the peripheral blood of asymptomatic pediatric liver transplant recipients 3rd International Congress on Pediatric Transplantation Krieger, N. R., Martinez, O. M., Krams, S. M., Cox, K., So, S., Esquivel, C. O. JOHN WILEY & SONS INC. 2000: 62?66
Abstract

Pediatric allograft recipients are at increased risk for Epstein-Barr virus (EBV)-associated illnesses. The early identification and diagnosis of EBV-associated disorders is critical because disease progression can often be curtailed by modification of immunosuppression. We have previously shown that detection of EBV-specific sequences in the circulation by polymerase chain reaction (PCR) correlated well with the clinical symptoms of EBV infection. The purpose of the current study is to determine the significance of detecting EBV-specific sequences by PCR in asymptomatic pediatric liver transplant recipients. Peripheral-blood DNA was analyzed for the EBV genes, coding from the nuclear antigen 1 (EBNA-1) and the viral capsid antigen (gp220) by PCR. Samples from asymptomatic pediatric liver transplant recipients were analyzed from the immediate postoperative period and at 2- to 4-month intervals thereafter. We followed up 13 of these asymptomatic recipients who tested positive for EBV compared with 7 asymptomatic recipients who tested negative for EBV during the early posttransplantation period. Follow-up ranged from 1.5 to 4 years posttransplantation. Nine patients (69%) initially positive for EBV and asymptomatic ultimately developed symptoms of EBV infection, including fever, lymphadenopathy, rash, respiratory and gastrointestinal symptoms, and/or hepatitis. Five of these patients (56%) went on to develop posttransplant lymphoproliferative disorder based on histological examination of biopsied tissue and immunohistochemical identification of the EBV antigen/DNA in tissue. This is the first report suggesting that detection of EBV-specific sequences in the absence of symptoms may herald impending EBV-associated disorders. Thus, routine monitoring for circulating EBV sequences in asymptomatic recipients may be useful in the early identification of those at risk for developing EBV-associated disease and its ultimate prevention.

View details for Web of Science ID 000085673100008

View details for PubMedID 10648579
Involvement of Fas-Fas ligand interactions in graft rejection. International reviews of immunology Martinez, O. M., Krams, S. M. 1999; 18 (5-6): 527-546
Abstract

The Fas/Fas ligand (FasL) pathway has been shown to be important in T lymphocyte-mediated cell death and is a key peripheral immunoregulatory mechanism that limits expansion of antigen-activated lymphocytes. The expression of Fas by commonly transplanted organs such as the heart, lung, kidney, and liver suggests that these tissues may be targets of FasL-expressing allospecific cytotoxic T lymphocytes. In this review the current literature examining the Fas/FasL system as a potential cellular effector pathway in tissue injury is discussed. In addition to a deleterious role in destruction of graft tissue, Fas/FasL interactions may have a beneficial role in transplantation. Recent studies suggest that modulation of FasL in target tissue leads to deletion, via apoptosis, of graft infiltrating lymphoid cells. However, an equally compelling series of reports indicate that overexpression of FasL can lead to a heightened immune response. These data are reviewed in the context of strategies to achieve long term allograft survival.

View details for PubMedID 10672500
Posttransplant lymphoproliferative disorders and gastrointestinal manifestations of Epstein-Barr virus infection in children following liver transplantation TRANSPLANTATION Cao, S., Cox, K., Esquivel, C. O., Berquist, W., Concepcion, W., Ojogho, O., Monge, H., Krams, S., Martinez, O., So, S. 1998; 66 (7): 851-856
Abstract

Epstein-Barr virus (EBV) infection is common after liver transplantation in children and is associated with the risk of posttransplant lymphoproliferative disorders (PTLD).This retrospective study examined the frequency of gastrointestinal (GI) symptoms and the risk of PTLD in pediatric liver recipients who developed symptomatic EBV infection. We reviewed 172 children who received orthotopic liver transplants between March 1988 to December 1994. Twenty-two cases were retransplants. The mean age at transplantation was 3.7 years (range, 0.1-17 years). The immunosuppressive regimens consisted of induction therapy with Minnesota antilymphocyte globulin/antithymocyte globulin/OKT3 in most cases and maintenance therapy with prednisone and either cyclosporine or tacrolimus (FK506).After 1 year of minimum follow-up, 54 of 172 patients had symptomatic EBV infections (confirmed by serology, histology, or whole blood polymerase chain reaction. At the time of infection, 38.5% (21/54) had either diarrhea or GI bleeding or both. PTLD developed in 11 patients (6.4%). The incidence of PTLD was 42.9% (9/21) when GI bleeding or diarrhea was associated with EBV infections, compared with 6.1% (2/33) when EBV infection was not associated with GI symptoms. Seven of 10 (70%) patients with GI bleeding and 2 of 11 (18.2%) with diarrhea developed PTLD. Of seven patients examined by endoscopy for GI bleeding, two had biopsy-proven PTLD of the GI tract, whereas one of two patients examined by endoscopy for diarrhea had biopsy-proven PTLD.In summary, a high incidence of PTLD was found in patients who developed GI bleeding or diarrhea associated with EBV infection after pediatric liver transplantation. In these patients, endoscopy and biopsy may lead to early diagnosis of PTLD.

View details for Web of Science ID 000076585400007

View details for PubMedID 9798693
Characterization of allograft rejection in an experimental model of small intestinal transplantation. Journal of gastrointestinal surgery Hayashi, M., Martinez, O. M., Krams, S. M., BURNS, W., Esquivel, C. O. 1998; 2 (4): 325-332
Abstract

Graft rejection continues to be a major barrier to the success of clinical small intestinal transplantation. The objective of this study was to characterize histopathologic and immune parameters of allograft rejection in an experimental model of small intestinal transplantation. Heterotopic intestinal transplants were performed in allogeneic and isogeneic rat strain combinations. An additional group of allogeneic recipients was treated with tacrolimus (1 mg/kg/day) for 7 days beginning on posttransplant day 1. Recipients of allografts and isografts were killed on days 1 to 7 following transplantation, and tacrolimus-treated allograft recipients were killed on days 4 and 7. Grafts and native intestines were examined for histopathology and cytokine gene expression. Very early rejection was observed on posttransplant day 3 and severe rejection was apparent by day 7. The key histopathologic features of acute graft rejection including apoptosis, crypt epithelial cell injury, and an inflammatory infiltrate were uniformly identifiable on day 4 and progressed in severity through day 7. Interleukin (IL)-2, IL-4, IL-5, IL-6, interferon-gamma (IFN-gamma), and tumor necrosis factor-alpha mRNA were readily detectable in allografts on days 1 to 7. However, only IFN-gamma mRNA showed a significant early and sustained increase in allografts as compared to isografts and native intestine. Treatment of allograft recipients with tacrolimus abrogated the major histopathologic features of rejection and markedly inhibited IFN-gamma gene expression. These results indicate that graft rejection in small intestinal transplantation is characterized by a local and specific immune response marked by IFN-gamma production that results in crypt epithelial cell injury and apoptosis. Tacrolimus abrogates the histopathologic features of rejection in association with a marked inhibition of IFN-gamma gene expression.

View details for PubMedID 9841989
Effect of cyclosporine and tacrolimus on the growth of Epstein-Barr virus-transformed B-cell lines 16th Annual Meeting of the American-Society-of-Transplant-Physicians Beatty, P. R., Krams, S. M., Esquivel, C. O., Martinez, O. M. LIPPINCOTT WILLIAMS & WILKINS. 1998: 1248?55
Abstract

Transplant patients receiving immunosuppressive drugs are at increased risk for Epstein-Barr virus (EBV)-associated disorders including posttransplant lymphoproliferative disorder. The function of T lymphocytes, which are critical to preventing the expansion of EBV-infected B cells, is inhibited by immunosuppressive drugs. The purpose of this study was to determine whether immunosuppressive drugs have direct effects on EBV-infected B cells.The growth and proliferation of EBV-infected spontaneous lymphoblastoid cell lines (SLCLs), cultured in the presence or absence of cyclosporine (CsA) and tacrolimus (TAC), were measured by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and [3H]thymidine incorporation assays. The effect of CsA and TAC on the viability of SLCLs was determined by cell counts with trypan blue. Apoptosis of SLCLs was induced with an anti-Fas agonist monoclonal antibody in the presence or absence of CsA and TAC and measured by flow cytometry after terminal deoxynucleotidyl transferase end-labeling and propidium iodide staining.CsA and TAC, but not sirolimus, increased the growth of SLCLs. The increased growth in the presence of CsA and TAC was attributable to enhanced cell viability and not increased cell division of SLCLs. In addition, CsA and TAC inhibited Fas-mediated apoptosis of SLCLs.CsA and TAC enhance the survival of EBV-transformed B-cell lines. CsA and TAC promote or augment SLCL growth through protection from cell death but do not affect cell division. The inhibition of cell death by CsA and TAC may contribute to the expansion of EBV-infected cells in immunosuppressed individuals.

View details for Web of Science ID 000073676600017

View details for PubMedID 9603175
CD30 expression identifies a functional alloreactive human T-lymphocyte subset TRANSPLANTATION Martinez, O. M., Villanueva, J., Abtahi, S., Beatty, P. R., Esquivel, C. O., Krams, S. M. 1998; 65 (9): 1240-1247
Abstract

CD30 is a member of the tumor necrosis factor/nerve growth factor receptor family and has been proposed as a marker of specific cytokine-producing subsets in humans. Previous studies have examined the expression of CD30 on established T helper type 1 and T helper type 2 cell clones and the function of CD30+ cells after mitogenic stimulation. In this study, we examined the development and function of CD30+ T cells generated in response to alloantigen.Primary one-way mixed lymphocyte reactions were established, and the expression of CD30 on T lymphocytes was determined by immunofluorescence and flow cytometry. Fluorescence-activated cell sorting was utilized to define the cytokine profile of alloactivated CD30+ cells after restimulation with anti-CD3 monoclonal antibodies or alloantigen. The effect of cyclosporine on the development of CD30+ cells, and on cytokines produced by CD30+ T lymphocytes, in response to alloantigen was determined.CD30+ T lymphocytes could be detected on day 2 of mixed lymphocyte reactions and continued to increase in number and proportion through day 6. Both CD4 and CD8 T cells expressed CD30 after primary alloantigenic stimulation. CD30+ T cells are a subset of alloactivated T cells and are the major source of interferon-gamma and interleukin-5 produced in response to alloantigen. Cyclosporine partially, but not completely, inhibits the development of CD30+ cells, and has a greater effect on interferon-gamma production than on interleukin-5 production.CD30+ T lymphocytes may constitute an important immunoregulatory subset in human allograft rejection.

View details for Web of Science ID 000073676600016

View details for PubMedID 9603174
Allograft rejection after liver transplantation for autoimmune liver diseases LIVER TRANSPLANTATION AND SURGERY Hayashi, M., Keeffe, E. B., Krams, S. M., Martinez, O. M., Ojogho, O. N., So, S. K., Garcia, G., Imperial, J. C., Esquivel, C. O. 1998; 4 (3): 208-214
Abstract

Autoimmune liver diseases (AILD) may progress to liver failure, requiring liver transplantation as definitive therapy, and these immune-mediated disorders may predispose the patient to more frequent graft rejection. The objective of this study was to determine the effect of preexisting AILD on the incidence of allograft rejection after liver transplantation. Sixty-three patients who underwent liver transplantation between March 1988 and December 1994 for AILDs that included autoimmune hepatitis (AIH; n = 33) and primary biliary cirrhosis (PBC; n = 30) were retrospectively compared with 47 patients who underwent liver transplantation for alcoholic cirrhosis during the same time period. There was a lower incidence of acute allograft rejection in patients with AILD who received tacrolimus-based compared with cyclosporine-based immunosuppression (50% v 85.5%; P = .02). However, patients with AILDs overall had a higher incidence of acute rejection than patients with alcoholic cirrhosis (81% v 46.8%; P < .001), regardless of the type of immunosuppression. In addition, steroid-resistant rejection occurred more frequently in patients with AILDs than in patients with alcoholic cirrhosis (38.1% v 12.8%; P = .003). There was also a trend toward a higher incidence of chronic rejection in patients with AILDs compared with patients with alcoholic cirrhosis (11.1% v 2.1%), but this difference did not reach statistical significance. Patient and graft survivals at 1 and 3 years were similar between patients with AILDs and alcoholic liver disease. Compared with alcoholic cirrhosis, preexisting AILDs are associated with a higher incidence of acute allograft rejection and a trend toward more frequent chronic rejection.

View details for Web of Science ID 000077183800004

View details for PubMedID 9563959
CD8(+) cells are not necessary for allograft rejection or the induction of apoptosis in an experimental model of small intestinal transplantation JOURNAL OF IMMUNOLOGY Krams, S. M., Hayashi, M., Fox, C. K., Villanueva, J. C., Whitmer, K. J., BURNS, W., Esquivel, C. O., Martinez, O. M. 1998; 160 (8): 3673-3680
Abstract

Allospecific CTL can function as cellular effectors of solid organ graft rejection; however, the specific mechanisms of cell damage remain undetermined. In this study we examined the role of CD8+ T cells in apoptosis and rejection of small intestinal allografts. ACI rat intestinal grafts transplanted into Lewis rat recipients showed apoptosis of epithelial crypt cells on day 3 posttransplant as determined by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling staining. By day 7 numerous apoptotic crypt cells were detected in allografts, but were rarely observed in FK506-treated allograft recipients, isografts, or native intestine of allograft recipients. To further investigate the mechanism of rejection, recipient rats were depleted of CD8+ cells by treatment with OX-8 mAbs the day before and the day after transplantation of rat small intestinal allografts. Depletion of CD8+ cells from allograft recipients did not alter the tempo or the histologic features of rejection compared with those in the control (IgG-treated) group. Moreover, there was no difference in the number of apoptotic crypt epithelial cells in the grafts of control and CD8-depleted rats. Reverse transcriptase-PCR analyses determined there were similar levels of transcripts for Fas, Fas ligand, perforin, and granzyme B in control and CD8-depleted allograft recipients. By Western blot it was determined that the levels of Fas ligand protein were increased in the CD8-depleted group compared with those in control and FK506-treated allograft recipients. These data suggest that CD8 cells are not required for tissue injury or apoptotic cell death in small intestine allograft rejection.

View details for Web of Science ID 000072970400008

View details for PubMedID 9558067
Application of in situ hybridization technique for quantitative assessment of ongoing symptomatic Epstein-Barr virus infection after living related liver transplantation CLINICAL TRANSPLANTATION Egawa, H., Oh-Ishi, T., Arai, T., Inomata, Y., Uemoto, S., Asonuma, K., Kiuchi, T., Okajima, H., Matsui, O., Kawashima, N., Martinez, O. M., Tanaka, K. 1998; 12 (2): 116-122
Abstract

For quantitative assessment of ongoing symptomatic Epstein-Barr virus (EBV) infection in pediatric recipients of liver transplantation, we determined the number of peripheral blood mononuclear cells (PBMC) infected by EBV by in situ hybridization (ISH) and related the results with clinical courses of those patients. Twenty-four patients had symptomatic EBV infection between February 1995 and March 1996. Blood samples were obtained from these 24 patients at the time of acute phase, from 13 of them during convalescence, and 37 pediatric patients before transplantation. ISH was performed on the PBMC and polymerase chain reaction (PCR) on DNA from whole blood. Oligonucleotide probes for ISH were chosen from coding sequences of EBV-encoded small nuclear RNA 1 (EBER1). Results of ISH were reported in a number of cells expressing EBER1/5 x 104 PBMC (#EBER1). Fever, diarrhea, upper respiratory symptoms, pleural effusion, ascites, lymphadenopathy, and lymphoproliferative disease (LPD) accompanied with EBV infection proven by serology, viral-specific stain or PCR were regarded as EBV related diseases (EBVD). All samples with positive #EBER1 were accompanied by positive EBV PCR. #EBERI was 68.2 +/- 144.9 (mean +/- SD) ranging from 0 to 621 in the acute phase, 0.20 +/- 0.41 ranging from 0 to 2 in the convalescence phase, 0.27 +/- 0.77 in 23 preoperative patients with positive serology, and 0 in all 14 preoperative patients with negative serology. The #EBER1 in ongoing EBVD was significantly greater than that of patients in convalescence or before transplantation. Patients with #EBERI greater than 10 had a significantly lower chance of convalescence and a higher mortality than patients with #EBER 1 less than 10. We conclude that #EBER1 could be a specific and quantitative marker of EBVD and might predict progression to LPD.

View details for Web of Science ID 000073040900008

View details for PubMedID 9575399
Pathways of biliary epithelial cell apoptosis in primary biliary cirrhosis Fox, C. K., Martinez, O. M., Krams, S. M. FEDERATION AMER SOC EXP BIOL. 1998: A285?A285
View details for Web of Science ID 000076006401654
Human hepatocytes produce an isoform of Fas that inhibits apoptosis TRANSPLANTATION Krams, S. M., Fox, C. K., Beatty, P. R., Cao, S., Villanueva, J. C., Esquivel, C. O., Martinez, O. M. 1998; 65 (5): 713-721
Abstract

Fas (Apo-1/CD95), a member of the tumor necrosis factor receptor family, can mediate apoptosis when engaged by its ligand or by anti-Fas antibody. Fas is expressed by cells of the immune system and by some nonlymphoid tissues. Numerous studies have suggested that the Fas pathway may play a role in the rejection of allografts. Functional, soluble forms of the Fas receptor are produced by activated peripheral blood mononuclear cells and some transformed cell lines. The purpose of this study was to determine if soluble variants of Fas are produced in the liver and to determine if blockade of the Fas pathway, by liver-derived soluble Fas, inhibits Fas-mediated apoptosis.Liver and purified hepatocyte specimens were analyzed for Fas transcripts by reverse transcriptase-polymerase chain reaction with primers that span the transmembrane region of the molecule. Bile and cell lysates were analyzed for soluble Fas by specific enzyme-linked immunosorbent assay. Lysates were prepared from normal liver and hepatocytes and utilized to block Fas-mediated apoptosis of Jurkat cells as determined by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling and flow cytometry.A variant form of Fas is abundantly expressed in normal liver and purified hepatocytes. This variant form of Fas is expressed in all normal liver specimens but only in half of the liver specimens obtained during allograft rejection. The levels of soluble Fas diminish in patients undergoing liver allograft rejection in contrast to patients with stable grafts. Importantly, a soluble form of Fas is produced in the liver by hepatocytes and can specifically inhibit Fas-mediated apoptosis.These data raise the possibility that soluble Fas, produced by hepatocytes, may influence the immune response by blocking Fas-mediated apoptosis and, thus, may have a role in liver transplantation.

View details for Web of Science ID 000072573800019

View details for PubMedID 9521208
Apoptosis as a mechanism of tissue injury in liver allograft rejection SEMINARS IN LIVER DISEASE Krams, S. M., Martinez, O. M. 1998; 18 (2): 153-167
Abstract

Recent studies suggest that apoptosis is an important mechanism of cell death in the rejection of liver allografts and that infiltrating host lymphocytes mediate this process. The first section of this chapter addresses the cells and molecules that initiate the immune response following transplantation of a liver allograft. The recognition of donor alloantigens by infiltrating host lymphocytes stimulates a cascade of immune events which culminate in development of the effector cells that mediate tissue damage. Studies which demonstrate that apoptosis of hepatocytes and bile duct cells accompany allograft rejection are detailed in the second section of this chapter. The final section discusses the potential pathways which lead to apoptosis in liver allograft rejection. The contributions of the granule-exocytosis pathway, the Fas-mediated pathway, and cytokines to the induction of apoptosis in liver allografts are discussed. In addition, the concept that alloreactive graft infiltrating cells are deleted by apoptosis is presented. A further understanding of the mechanisms involved in apoptosis will lead to unique approaches toward the goal of achieving allograft tolerance.

View details for Web of Science ID 000079131100006

View details for PubMedID 9606812
Involvement of IL-10 in the autonomous growth of EBV-transformed B cell lines JOURNAL OF IMMUNOLOGY Beatty, P. R., Krams, S. M., Martinez, O. M. 1997; 158 (9): 4045-4051
Abstract

Immunocompromised individuals have an increased incidence of EBV-associated B cell lymphomas. The growth factors responsible for the unrestrained proliferation of these lymphomas have not yet been determined. In this study, spontaneous lymphoblastoid cell lines (SLCL) were derived without the addition of growth factors or virus from four patients with EBV-associated lymphoproliferative disorder. These cell lines were EBV transformed in vivo, and infection was verified through amplification of the viral gp220 gene. SLCL have an activated B cell phenotype (CD19+, CD21+, CD23+, CD38+, and CD40+) and produce IL-6, IL-10, TNF-alpha, and lymphotoxin-alpha. To determine whether these cytokines contribute to autonomous growth, neutralizing Abs for IL-10, IL-6, and TNF-alpha, a soluble TNFR:Fc fusion protein, and soluble IL-10R were used. These experiments established that, of the cytokines produced by SLCL, only IL-10 is an autocrine factor. IL-10 was produced by the majority of cells within each SLCL, and IL-10 secretion was concomitant with SLCL growth. Our findings demonstrate that IL-10 is utilized in the autonomous growth of EBV-related lymphomas and may be crucial in the development of lymphoproliferative disorder.

View details for Web of Science ID A1997WV76100005

View details for PubMedID 9126962
Cytokines are involved in the rejection of small intestine allografts VIth Meeting of the European-Intestinal-Transplantation-Study-Group (EITSG) Hayashi, M., Martinez, O. M., Krams, S. M., BURNS, W., Esquivel, C. O. ELSEVIER SCIENCE INC. 1997: 1802?
View details for Web of Science ID A1997WX49600060

View details for PubMedID 9142277
T cell receptor usage by cytotoxic T lymphocytes against autologous human melanoma ANTICANCER RESEARCH Leong, S. P., Liliental, J., Krams, S. M., Zhou, Y. M., GRANBERRY, M. E., Martinez, O. M. 1996; 16 (6B): 3355-3361
Abstract

T cell receptor (TCR) usage by tumor infiltrating lymphocytes (TIL) represents the host's response to autologous melanoma (AM). The goal of this study is it determine TCR usage by cytotoxic T lymphocytes (CTL) against AM.CTL were generated from three patients using lymphocytes from metastatic or tumor-draining lymph nodes by repeated in vitro sensitization (IVS). Total RNA was isolated from CTL and reverse-transcribed to cDNA. TCR usage was determined by polymerase chain reaction (PCR) using TCR primers.Cytolytic activity was non-specific within the first 2-4 weeks following IVS and TCR repertoire in these cultures revealed random V alpha and V beta gene usage. In contrast, by 6-10 weeks of culture, cytolysis was specifically directed against AM cells and such specific cytolysis was significantly correlated with TCR V alpha 1 (P < 0.001).TCR V alpha 1 is associated with a common restricted melanoma antigen.

View details for Web of Science ID A1996WB83600001

View details for PubMedID 9042192
Production of IL-4 and IL-10 does not lead to immune quiescence in vascularized human organ grafts TRANSPLANTATION Lang, T., Krams, S. M., Martinez, O. M. 1996; 62 (6): 776-780
Abstract

The delineation of T helper cell subsets into T helper type 1 (Th1) and T helper type 2 (Th2) populations based on the production of specific cytokines has been useful in understanding the regulation and progression of immune-based pathologies. In order to test the relevance of this concept to human solid organ transplantation, in situ and system Th1 and Th2 cytokine profiles were characterized in liver allograft recipients. Bile and serum samples obtained posttransplant were analyzed for the cytokines IL-2, IFN-gamma, IL-4, and IL-10. Significant elevations of IL-4 and IL-10 were measured at the site of graft rejection. In contrast, IL-2, IFN-gamma, and IL-4 were markedly elevated in the circulation during rejection. Analyses of sequential bile samples revealed that Th1 and Th2 cytokines showed similar kinetics of production in response to alloantigen. Taken together, these results indicate that acute rejection of human allografts can proceed [correction of procede] in the presence of minimal levels of the Th1 cytokines IL-2 and IFN-gamma and high levels of IL-4 and IL-10.

View details for Web of Science ID A1996VK61100014

View details for PubMedID 8824477
Elevations in IFN-gamma, IL-5, and IL-10 in patients with the autoimmune disease primary biliary cirrhosis: Association with autoantibodies and soluble CD30 CLINICAL IMMUNOLOGY AND IMMUNOPATHOLOGY Krams, S. M., Cao, S., Hayashi, M., Villanueva, J. C., Martinez, O. M. 1996; 80 (3): 311-320
Abstract

Antimitochondrial antibodies (AMA) which recognize the E2 component of the pyruvate dehydrogenase complex are found in virtually all patients with the autoimmune liver disease primary biliary cirrhosis (PBC). The factors that contribute to elevated AMA and the relationship of the autoantibodies to disease pathogenesis have not been elucidated. Since cytokines are important regulators of antibody production and isotype switching, the association of specific cytokines to antibody production was examined in patients with PBC. Elevations in IL-2, IFN-gamma, IL-4, IL-5, and IL-10 were detected in serum from patients with PBC. However, only IFN-gamma (6012 +/- 1128 pg/ml vs 147 +/- 89 pg/ml, P < 0.0001) and IL-5 (382 +/- 103 pg/ml vs 29 +/- 12 pg/ml, P < 0.001) were significantly elevated compared to normal controls. Moreover, there was a positive correlation in the levels of IFN-gamma, and to a lesser extent IL-5, with the levels of soluble CD30 (sCD30) in the circulation. The elevated levels of sCD30 detected in patients with PBC (194 +/- 29 U/ml vs 39 +/- 9 U/ml in normal controls) suggest that CD30+ cells may produce cytokines, which contribute to the immune abnormalities in patients with PBC.

View details for Web of Science ID A1996VG74900013

View details for PubMedID 8811053
Immunoregulatory cytokines in chronic hepatitis C virus infection: Pre- and posttreatment with interferon alfa HEPATOLOGY Cacciarelli, T. V., Martinez, O. M., Gish, R. G., Villanueva, J. C., Krams, S. M. 1996; 24 (1): 6-9
Abstract

T lymphocytes and immunoregulatory cytokines may be important in the host response to hepatitis C virus (HCV) infection. T-helper type 1 (Th1) cytokines (interleukin [IL]-2, interferon gamma [IFN-gamma]) are required for host antiviral immune responses, including cytotoxic T-cell generation and natural killer cell activation, while T-helper type 2 (Th2) cytokines (IL-4,IL-10) can inhibit the development of these effector mechanisms. In this study, the serum levels of Th1 and Th2 cytokines in patients (n = 23) infected with HCV were measured and compared with biochemical (alanine transaminase [ALT]) and viral (HCV RNA) indicators of infection. Serial cytokine levels were measured in a subset of 11 patients at 1 and 12 weeks during and at 1 week after interferon alfa (IFN-alpha) therapy (n = 33 samples). Levels of circulating IL-2, IL-4, IL-10, and IFN-gamma were significantly elevated in HCV patients versus normal controls (128 vs. 25 pg/mL, 3,045 vs. 29 pg/mL, 2,949 vs. 18 pg/mL, and 307 vs. 24 pg/mL respectively; P < .01). Treatment with IFN-alpha decreased the levels of IL-4 (321 +/- 224 pg/mL), and IL-10 (1,011 +/- 344 pg/mL), which paralleled a decrease in HCV RNA (114 +/- 27 vs. 25 +/- 20 Eq/ml X 10(5), pre- vs. post-IFN-alpha [12 weeks];P <.05). These findings indicate that an activated T-cell response, as manifest by increased circulating immunoregulatory cytokines, is present in patients with HCV liver disease. Furthermore, treatment with HCV liver disease. Furthermore, treatment with IFN-alpha diminishes the Th2 cytokine response. Thus, modulation of T-cell function and cytokine production may be one mechanism whereby IFN-alpha therapy results in reduced viral burden.

View details for Web of Science ID A1996UW63500002

View details for PubMedID 8707283
Elevations in IFN-gamma, IL-5, and IL-10 in patients with the autoimmune disease, primary biliary cirrhosis. Association with autoantibodies and soluble CD30. Krams, S. M., Cao, S., Hayashi, M., Esquivel, C. O., Martinez, O. M. W B SAUNDERS CO-ELSEVIER INC. 1996: A1240?A1240
View details for Web of Science ID A1996UF73704937
Apoptosis as a mechanism of cell death in liver allograft rejection (reply) TRANSPLANTATION Krams, S. M., Martinez, O. M. 1996; 61 (1): 168-169
View details for Web of Science ID A1996TQ20100035
Expression of cytokines and immune mediators during chronic liver allograft rejection 14th Annual Meeting of the American-Society-of-Transplant-Physicians Hayashi, M., Martinez, O. M., GARCIAKENNEDY, R., So, S., Esquivel, C. O., Krams, S. M. WILLIAMS & WILKINS. 1995: 1533?38
Abstract

To determine the immune processes involved in chronic liver allograft rejection (CR) we examined in situ cytokine production in tissue from 15 patients with both clinical and histopathological diagnoses of CR. Total RNA was isolated from liver samples, reverse-transcribed and analyzed by RT-PCR for the production of proinflammatory cytokines and immunoregulatory mediators. Transcripts for the Th1-like cytokines IL-2 and IFN-gamma were detected in 53.3% and 46.7% of CR grafts, while they were detected in only 16% and 0% of stable grafts, respectively. The cytotoxic T cell mediator granzyme B was expressed in the majority of liver grafts undergoing CR, but was expressed only in a minority of stable grafts (80% vs. 16%, P < 0.05). The T cell product IL-5 was also significantly upregulated in CR as compared with stable livers (80% vs. 16%, P < 0.01). Other Th2 cytokines--IL-4 and IL-10--and macrophage products--IL-1 beta, IL-6, IL-8, TGF-beta, and TNF-alpha--were not substantially upregulated in CR grafts as compared with stable grafts. PDGF-beta transcripts were detected in the majority of the CR grafts, but were not detected in stable liver grafts (73% vs. 0, P < 0.05). By immunohistochemical staining, we observed that CD3+CD4+, and CD3+CD4- T cells were detected in CR grafts along with CD20+ B cells and CD68+ macrophages. There was, however, a predominant infiltration of CD3+CD4+ lymphocytes. Taken together, these data suggest that infiltrating cells produce proinflammatory and immunoregulatory cytokines that have a role in mediating graft damage in CR.

View details for Web of Science ID A1995TN23000027

View details for PubMedID 8545886
Elevated biliary interleukin 5 as an indicator of liver allograft rejection. Transplant immunology Lang, T., Krams, S. M., Berquist, W., Cox, K. L., Esquivel, C. O., Martinez, O. M. 1995; 3 (4): 291-298
Abstract

Interleukin 5 (IL-5) is a T cell-derived cytokine that acts as a potent and specific eosinophil differentiation factor in humans. During liver allograft rejection, intragraft IL-5 mRNA and eosinophilia have been observed. The objective of this study was to correlate the levels of IL-5 in bile and serum with eosinophilia and allograft rejection in paediatric liver recipients. IL-5 levels were determined by ELISA (enzyme-linked immunosorbent assay) in bile (n = 85) and serum (n = 106) and obtained prospectively from 15 patients during the first 3 weeks post-transplantation. Biliary and serum IL-5 levels were significantly elevated during allograft rejection compared to IL-5 levels when no rejection was apparent or during infectious complications. The highest IL-5 levels were measured in the bile during the early rejection period (3 days prior to biopsy-proven rejection). Fifteen of 16 rejection episodes were marked by increases in IL-5 as revealed by analysis of sequential samples from individual patients. In all patients with rejection, elevations in serum IL-5 were associated with elevations in peripheral eosinophil counts. These results indicate that IL-5 is produced in the liver and may be a useful and specific marker of allograft rejection. Furthermore, these findings provide further evidence for a pathway of liver allograft rejection mediated by IL-5 activated eosinophils.

View details for PubMedID 8665147
THE USE OF NON-HEART-BEATING CADAVER DONORS IN EXPERIMENTAL LIVER-TRANSPLANTATION TRANSPLANTATION Tojimbara, T., Kennedy, R. G., BURNS, W., Hayashi, M., Krams, S., Martinez, O., So, S., Esquivel, C. O. 1995; 60 (10): 1179-1180
View details for Web of Science ID A1995TH32800020

View details for PubMedID 7482728
Expression of the cytotoxic T cell mediator granzyme B during liver allograft rejection. Transplant immunology Krams, S. M., Villanueva, J. C., QUINN, M. B., Martinez, O. M. 1995; 3 (2): 162-166
Abstract

Cytotoxic T lymphocytes (CTL) constitute a major component of the alloreactive response following organ transplantation. The molecular mechanisms of CTL killing remain to be determined but multiple candidate molecules involved in CTL-mediated cytotoxicity have been identified. Granzyme B, a serine protease, participates in perforin-dependent pathways of cytotoxicity and is necessary for induction of DNA fragmentation in target cells. In this study the expression of granzyme B in liver biopsies obtained from liver allograft recipients was determined by semiquantitative reverse transcriptase polymerase chain reaction. Biopsies were classified into four groups--no evidence of rejection, preservation injury, acute rejection, or resolving rejection--according to histopathological criteria. There was a significantly higher frequency of transcripts for granzyme B in the acute rejection group (82.8%) compared to the no rejection (20.0%), resolving rejection (12.5%) and preservation injury (0%) groups. Analysis of granzyme B gene expression in sequential samples from individual patients prior to, and after, treatment for rejection revealed an inverse correlation between granzyme B mRNA and response to treatment. These findings indicate that the cytopathic mediator granzyme B may participate in CTL-mediated cytotoxicity during liver allograft rejection.

View details for PubMedID 7582907
AN INCREASED INCIDENCE OF EPSTEIN-BARR-VIRUS INFECTION AND LYMPHOPROLIFERATIVE DISORDER IN YOUNG-CHILDREN ON FK506 AFTER LIVER-TRANSPLANTATION 13th Annual Meeting of the American-Society-of-Transplant-Physicians Cox, K. L., LAWRENCEMIYASAKI, L. S., GARCIAKENNEDY, R., Lennette, E. T., Martinez, O. M., Krams, S. M., Berquist, W. E., So, S. K., Esquivel, C. O. WILLIAMS & WILKINS. 1995: 524?29
Abstract

The incidence of Epstein-Barr virus (EBV) infection and lymphoproliferative disorder (LPD) was determined in a pediatric liver transplant population consisting of 51 children treated with FK506 and 91 treated with cyclosporine. The incidence of symptomatic EBV infection was 21.9% (23 of 105 cases) in children < 5 yr old and 10.8% (4 of 37 cases) in children 5 to 17 yr old as compared with 2.7% (9 of 323 cases) in adults (P < 0.0001). In the under 5 yr old group on cyclosporine, the incidences of EBV infection and LPD were 9 of 68 (13.2%) and 2 of 68 children, (2.9%), respectively. In contrast, in children under 5 yr old group on FK506, the incidences of EBV infection and LPD in the FK506 group were 14 of 37 (37.8%) and 7 of 37 children (18.9%), respectively. The difference between these two groups was statistically significant (P < 0.02). There were no cases of LPD in the 5-17 yr-old children on either cyclosporine (n = 23) or FK506 (n = 14). The incidence of EBV infections in the 5 to 17 yr age group, 17.4% on cyclosporine and 0% on FK506, was less than for the younger children on FK506 (37.8%). A total of 39% (9 of 23) of children under 5 yr old who had symptomatic EBV infections developed LPD, and 44% (4 of 9) with LPD died. The higher incidence of EBV infections and LPD in the younger children treated with FK506 was probably related to a greater intensity of immunosuppression for patients on FK506 than those on cyclosporine.

View details for Web of Science ID A1995QK22500015

View details for PubMedID 7533344
VIRAL AND IMMUNOLOGICAL ASPECTS OF EPSTEIN-BARR-VIRUS INFECTION IN PEDIATRIC LIVER-TRANSPLANT RECIPIENTS 13th Annual Meeting of the American-Society-of-Transplant-Physicians Martinez, O. M., Villanueva, J. C., LAWRENCEMIYASAKI, L., QUINN, M. B., Cox, K., Krams, S. M. WILLIAMS & WILKINS. 1995: 519?24
Abstract

Pediatric allograft recipients in particular are at increased risk for Epstein-Barr virus (EBV)-associated disorders. Early identification and diagnosis of EBV-associated disorders is critical, since disease progression can often be halted by reduction of immunosuppression. In this study we examined viral and immunologic parameters of EBV infection in the circulation of pediatric liver recipients to identify factors associated with disease. Peripheral blood DNA from pediatric liver recipients was analyzed by PCR for the EBV genes coding for the nuclear antigen 1 (EBNA-1) and the viral capsid antigen gp220. Sequences for these viral genes could be readily detected in the circulation of 36.5% of patients. Moreover, identification of the EBV genome was associated with symptomatic infection, suggesting that circulating EBV may be a useful marker of disease. Since EBV-infected B cells release the low-affinity IgE receptor (sCD23), we measured sCD23 in the circulation of pediatric liver recipients and found it to be elevated in patients with detectable virus or symptoms of infection. However, sCD23 was also elevated in cases where no EBV was detectable, suggesting that factors other than viral infection could stimulate release of sCD23. To further characterize the immune response to EBV infection, the peripheral levels of IL-4, IL-5, IL-10, and IFN-gamma were determined in pediatric liver recipients. Each of these cytokines was elevated in patients with symptoms or circulating virus compared with stable, age-matched liver recipients. IL-4, in particular, was significantly increased, indicating an important role for this cytokine in EBV infection. Together, these findings suggest that (1) monitoring circulating levels of EBV may be useful in patients at high risk and (2) cytokines that promote B cell growth and differentiation contribute to EBV-associated disorders.

View details for Web of Science ID A1995QK22500014

View details for PubMedID 7878757
APOPTOSIS AS A MECHANISM OF CELL-DEATH IN LIVER ALLOGRAFT-REJECTION 13th Annual Meeting of the American-Society-of-Transplant-Physicians Krams, S. M., Egawa, H., QUINN, M. B., Villanueva, J. C., GARCIAKENNEDY, R., Martinez, O. M. WILLIAMS & WILKINS. 1995: 621?25
Abstract

It is generally recognized that there are two mechanisms of cell death, apoptosis and necrosis. Apoptosis--programmed cell death--is involved in numerous states of physiological cell deletion. Recent studies have demonstrated that hepatocytes, under certain conditions, undergo apoptosis. The purpose of this work was to determine if apoptotic cell death is involved in liver allograft rejection. Groups of Lewis (RT1l) rats underwent orthotopic liver transplantation (OLT) from disparate DA (RT1a) or syngeneic Lewis rats. Liver samples were harvested at 1, 2, 3, 4, and 7 days posttransplant and analyzed for apoptotic cell death. Since the characteristics of apoptosis are difficult to discern using routine hematoxylin and eosin staining, we utilized a novel method that detects the classic indicator of apoptosis, nonrandom DNA degradation. Paraffin-embedded tissue sections were end-labeled with nonradioactive dUTP and detection of apoptotic bodies accomplished by immunoassay. The incidence of apoptotic cells increased steadily over time in allografts, in contrast to syngeneic grafts. In this study apoptotic cell death paralleled standard indicators of liver allograft rejection including pathology, mononuclear cell infiltration, and increases in liver enzymes. Moreover, increased expression of TGF-beta 1 correlated with apoptosis in liver allografts, supporting the previously described role for this cytokine in hepatocyte apoptosis. Our results demonstrate, for the first time, that apoptosis may be a mechanism of cell death in liver allograft rejection.

View details for Web of Science ID A1995QK22500031

View details for PubMedID 7878768
DIFFERENTIAL PATTERNS OF CIRCULATING INTERCELLULAR-ADHESION MOLECULE-1 (CICAM-1) AND VASCULAR CELL-ADHESION MOLECULE-1 (CVCAM-1) DURING LIVER ALLOGRAFT-REJECTION 13th Annual Meeting of the American-Society-of-Transplant-Physicians Lang, T., Krams, S. M., Villanueva, J. C., Cox, K., So, S., Martinez, O. M. WILLIAMS & WILKINS. 1995: 584?89
Abstract

During allograft rejection, adhesion molecules play an integral role in infiltration, activation, and binding of effector cells to target tissue. Some adhesion molecules, including ICAM-1 and VCAM-1, exist in soluble, circulating forms that retain ligand-binding activity. In the present study the levels of circulating ICAM-1 (cICAM-1) and VCAM-1 (cVCAM-1) were compared in the serum and bile of pediatric liver recipients. The cICAM-1 was significantly elevated in the serum during allograft rejection and infection relative to periods when no rejection was apparent. Biliary cICAM-1, however, was specifically elevated during rejection and not during infection or when no rejection was apparent. The cVCAM-1 levels were elevated in the serum during rejection compared with levels when no rejection was evident. In contrast, cVCAM-1 was not detected in the bile. Serum levels of both cICAM-1 and cVCAM-1 decreased rapidly following successful treatment for rejection, whereas elevated levels persisted, or increased, in ongoing rejection. The differential patterns of the circulating forms of ICAM-1 and cVCAM-1 were consistent with the membrane expression of these molecules during graft rejection. ICAM-1 expression was extensive on bile duct epithelium, endothelium, hepatocytes, and infiltrating leukocytes during rejection, while VCAM-1 was restricted to endothelium. These findings indicate that the release of circulating adhesion molecules is a prominent feature of liver allograft rejection. Measurement of these markers may be useful in distinguishing rejection from infection and in determining the efficacy of treatment for rejection.

View details for Web of Science ID A1995QK22500025

View details for PubMedID 7533349
MOLECULAR MARKERS OF EPSTEIN-BARR-VIRUS INFECTION IN THE CIRCULATION OF TRANSPLANT RECIPIENTS XVth World Congress of the Transplantation-Society Martinez, O. M., Villanueva, J. C., LAWRENCEMIYASAKI, L., QUINN, M. B., Gish, R., Cox, K., So, S., Esquivel, C. O., Krams, S. M. ELSEVIER SCIENCE INC. 1995: 1211?12
View details for Web of Science ID A1995QJ19900484

View details for PubMedID 7533372
DISTINCT PATTERNS OF TH2 CYTOKINE PRODUCTION DURING IMMUNE ACTIVATION IN PEDIATRIC LIVER ALLOGRAFT RECIPIENTS XVth World Congress of the Transplantation-Society Lang, T., Krams, S. M., Villanueva, J. C., Cox, K., So, S., Esquivel, C., Martinez, O. M. ELSEVIER SCIENCE INC. 1995: 1146?47
View details for Web of Science ID A1995QJ19900456

View details for PubMedID 7533367
CIRCULATING INTERCELLULAR-ADHESION MOLECULE-1 AND VASCULAR CELL-ADHESION MOLECULE-1 IN PEDIATRIC LIVER RECIPIENTS XVth World Congress of the Transplantation-Society Lang, T., Krams, S. M., Villanueva, J. C., So, S. K., Berquist, W. E., Cox, K. L., Esquivel, C. O., Martinez, O. M. ELSEVIER SCIENCE INC. 1995: 1148?49
View details for Web of Science ID A1995QJ19900457

View details for PubMedID 7533368
LONG-TERM NONRESPONSIVENESS TO A LIVER ALLOGRAFT MAY BE CYTOKINE-MEDIATED XVth World Congress of the Transplantation-Society Egawa, H., Martinez, O. M., QUINN, M. B., Villanueva, J. C., So, S., Esquivel, C. O., Krams, S. M. ELSEVIER SCIENCE INC. 1995: 241?42
View details for Web of Science ID A1995QJ19900085

View details for PubMedID 7533390
CHARACTERIZATION OF CYTOKINE EXPRESSION IN AN ANIMAL-MODEL OF ACUTE LIVER ALLOGRAFT-REJECTION XVth World Congress of the Transplantation-Society Egawa, H., Martinez, O. M., QUINN, M. B., So, S., Esquivel, C. O., Krams, S. M. ELSEVIER SCIENCE INC. 1995: 505?6
View details for Web of Science ID A1995QJ19900201

View details for PubMedID 7879079
APOPTOSIS AS A MECHANISM OF CELL-DEATH IN A RAT MODEL OF LIVER ALLOGRAFT-REJECTION XVth World Congress of the Transplantation-Society Krams, S. M., Egawa, H., QUINN, M. B., Martinez, O. M. ELSEVIER SCIENCE INC. 1995: 466?67
View details for Web of Science ID A1995QJ19900185

View details for PubMedID 7879064
INTERLEUKIN-12 - A POSSIBLE CYTOTOXIC T-LYMPHOCYTE DIFFERENTIATION FACTOR IN ALLOGRAFT RECIPIENTS XVth World Congress of the Transplantation-Society Gish, R. G., Krams, S. M., Martinez, O. M. ELSEVIER SCIENCE INC. 1995: 459?60
View details for Web of Science ID A1995QJ19900182

View details for PubMedID 7879061
ACUTE LIVER ALLOGRAFT-REJECTION IN THE RAT - AN ANALYSIS OF THE IMMUNE RESPONSE TRANSPLANTATION Egawa, H., Martinez, O. M., QUINN, M. B., Villanueva, J. C., So, S., Esquivel, C. O., Krams, S. M. 1995; 59 (1): 97-102
Abstract

Liver allografts are vigorously rejected in 9-12 days in Lewis recipients of fully histoincompatible DA livers. The purpose of this study was to examine the initial events in this cascade, specifically the role of CD4+ T helper cells. Lewis recipients of DA or Lewis livers were killed at days 1, 2, 3, 4, and 7 days after transplant. Indicators of acute liver rejection, including a marked inflammatory infiltrate and decreased liver function, progressed in untreated recipients of allografts. Splenocytes taken from allogeneic recipients on days 1-4 and 7 proliferated in response to donor and third-party stimulators, whereas graft-infiltrating cells did not respond to donor and third-party antigens until day 3 after transplant, but thereafter maintained a good response. To further characterize the host T helper cell response to liver allografts, cytokine expression was analyzed in graft tissue and in the periphery. IL-4 mRNA was present in both syngeneic and allogeneic liver grafts, while message for IL-10 was present early in all liver grafts but persisted only in allografts. In contrast, IL-2 and IFN-gamma transcripts were specific to rejecting allografts. Similar patterns of cytokine expression were observed in the spleen, indicating the immune response to the graft involves the peripheral lymphoid organs. Thus, the cytokine profile detected during liver allograft rejection is extremely similar to that observed in other experimental models of transplantation.

View details for Web of Science ID A1995QB42700017

View details for PubMedID 7839435
CYTOKINE PATTERNS AND CYTOTOXIC MEDIATORS IN PRIMARY BILIARY-CIRRHOSIS HEPATOLOGY Martinez, O. M., Villanueva, J. C., Gershwin, M. E., Krams, S. M. 1995; 21 (1): 113-119
Abstract

Primary biliary cirrhosis (PBC) is an autoimmune disease of the liver with unknown etiology. Autoreactive T lymphocytes that infiltrate the liver may play a major role in the bile duct damage that accompanies the disease. We hypothesized that cytokines produced by T lymphocytes and other cells are central to the disease process. Therefore, we used reverse transcription-polymerase chain reaction (PCR) and Southern hybridization to identify cytokine message directly from liver tissue of 11 patients with PBC and 5 patients with autoimmune hepatitis (AI-CAH). Messenger RNA (mRNA) for interleukin (IL)-2, IL-5, IL-6, interferon gamma (IFN-gamma), and transforming growth factor beta (TGF-beta) were detected in the majority of the specimens from patients with PBC. The presence of IL-5 was associated with PBC (P < .001, PBC vs. AI-CAH). Because IL-5 is a potent eosinophil differentiation factor, we looked for evidence of activated eosinophils within the infiltrate. We observed the deposition of the primary cytotoxic granule protein of eosinophils, major basic protein (MBP), within the portal region of livers from patients with PBC. Moreover, we detected message for a cytotoxic T-lymphocyte (CTL) granzyme in 87.5% of these livers indicating that mature CTL are present. Thus, we present evidence for two effector pathways that may contribute to the tissue damage observed in PBC and have identified message for cytokines that may regulate these pathways.

View details for Web of Science ID A1995RB78800019

View details for PubMedID 7806143
Liver transplantation at California Pacific Medical Center, San Francisco, California. Clinical transplants Esquivel, C. O., Martinez, O., Krams, S., Lim, J., So, S. K., Concepcion, W., Cox, K. L., Keeffe, E. B. 1994: 163-171
Abstract

A number of modifications in patient selection, operative technique, and immunosuppressive management have greatly contributed to the success of the liver transplant program at CPMC. Graft rejection and the timely detection of EBV infection are ongoing problems in hepatic transplantation that are foci of active research in our field. To address these issues, our group is investigating the activity of cytokines and adhesion molecules using sophisticated molecular techniques, and we are developing a sensitive assay for EBV markers in blood. These and other projects currently in progress will continue when we move our liver transplant program to Stanford University Medical Center in January 1995.

View details for PubMedID 7547535
NEW IMMUNOLOGICAL INSIGHTS INTO MECHANISMS OF ALLOGRAFT-REJECTION GASTROENTEROLOGY CLINICS OF NORTH AMERICA Krams, S. M., Ascher, N. L., Martinez, O. M. 1993; 22 (2): 381-400
Abstract

Our current understanding of liver allograft rejection indicates that multiple cellular interactions, involving a variety of cell-associated and soluble mediators, are critical to the response. The extravasation and localization of recipient immune cells to the allograft is dependent on recognition and interaction of complementary adhesion molecules expressed on circulating leukocytes and endothelium. Similar receptor-ligand pairs can also augment the binding of effector cells to target tissue within the allograft. Inflammatory mediators such as IL-1, IL-6, TNF-alpha, and IFN-gamma produced within the allograft can increase the local expression of adhesion molecules and thereby promote the entry of specific and nonspecific cells. The TCR expressed on T lymphocytes has the potential to recognize MHC antigens expressed on the allograft in many different forms. Thus, the T cell response to graft-associated alloantigens appears to be complex and dynamic. The production of T cell-derived cytokines is central to the activation and maturation of effector cells within the allograft. The identification of cytotoxic mediators such as serine protease and MBP within rejecting human allografts supports the role of cytotoxic T cells and eosinophils as effector cells. Undoubtedly, the development of genetically manipulated animal models will serve to further elucidate our understanding of the cellular mechanisms of graft rejection.

View details for Web of Science ID A1993LE17200013

View details for PubMedID 8509176
IL-2 AND IL-5 GENE-EXPRESSION IN RESPONSE TO ALLOANTIGEN IN LIVER ALLOGRAFT RECIPIENTS AND INVITRO 11TH ANNUAL MEETING OF THE AMERICAN SOC OF TRANSPLANT PHYSICIANS Martinez, O. M., VILLANEUVA, J. C., Lake, J., Roberts, J. P., Ascher, N. L., Krams, S. M. LIPPINCOTT WILLIAMS & WILKINS. 1993: 1159?66
Abstract

IL-2 and IL-5 gene expression in response to alloantigen was studied in liver allograft recipients and in an in vitro system. Seventy-seven sequential liver allograft biopsies from 22 patients were analyzed for IL-2 and IL-5 mRNA by polymerase chain reaction and Southern blot hybridization. Message for IL-5 was present in 74% of allografts with rejection, 46% of allografts with resolving rejection, and 33% of allografts with no evidence of rejection. The frequency of IL-5 transcripts in rejecting allografts was significantly different than the frequency of IL-5 transcripts in grafts without evidence of rejection (P = 0.003). Message for IL-2 was detected in 29% of rejecting allografts, 18% of allografts without evidence of rejection, and 43% of allografts with resolving rejection. There was no significant association between IL-2 gene expression and the histopathological status of the allograft. Interestingly, 9 of 15 biopsies that contained IL-2 message in the no rejection and resolving rejection categories went on to display rejection shortly thereafter. IL-2 and IL-5 gene expression rarely occurred simultaneously within allografts. An in vitro system consisting of irradiated, allogeneic stimulator cells and normal peripheral blood mononuclear cells as responders was established to further investigate alloantigen-driven IL-2 and IL-5 production. Both IL-2 and IL-5 were produced in response to alloantigen as determined by specific bioassays. Maximal levels of IL-5 activity in culture supernatants generally followed maximal IL-2 levels by 24 hr, but both IL-2 and IL-5 production were dramatically inhibited by CsA. Analysis of cytokine gene expression revealed that IL-2 transcription peaked within the initial 24 hr of culture, whereas IL-5 transcription was maximal at 120 hr of culture. The expression of a CTL-specific serine esterase gene was similar to IL-5 in that it was maximal during the latter phases of the culture period. Thus, both human IL-2 and IL-5 are produced in response to alloantigen and are inhibitable by CsA. These data suggest that IL-2 and IL-5 may participate in cellular pathways of tissue damage within the rejecting allograft.

View details for Web of Science ID A1993LD21200042

View details for PubMedID 8497897
EVIDENCE FOR A NONCLASSICAL PATHWAY OF GRAFT-REJECTION INVOLVING INTERLEUKIN-5 AND EOSINOPHILS 18TH ANNUAL MEETING OF THE AMERICAN SOC OF TRANSPLANT SURGEONS Martinez, O. M., Ascher, N. L., Ferrell, L., Villanueva, J., Lake, J., Roberts, J. P., Krams, S. M., LORBER WILLIAMS & WILKINS. 1993: 909?18
Abstract

The role of IL-5 and eosinophils in allograft rejection was studied in human liver allograft recipients. Liver allograft biopsies were analyzed for intragraft IL-5 gene expression, and the percentages of eosinophils and plasma cells within the portal infiltrate as well as peripheral eosinophil levels were determined. The majority of allografts with evidence of rejection had concomitant IL-5 mRNA and eosinophilia, while no resolving or nonrejecting allografts had simultaneous IL-5 mRNA and eosinophilia. In fact, rejecting liver allografts that contain IL-5 mRNA and eosinophils also contain infiltrating cells that produce the cytotoxic mediator major basic protein. In contrast, intragraft plasma cell and peripheral eosinophil levels did not correlate with the histopathologic status of the allograft. Cyclosporine and FK506 had similar effects on the frequency of IL-5 gene expression in rejecting and nonrejecting allografts. However, OKT3 appeared to profoundly modulate IL-5 gene expression, since 0 of 11 biopsies obtained during OKT3 treatment for rejection contained IL-5 transcripts. These observations raise the possibility of a cellular pathway of liver allograft rejection mediated by IL-5-activated eosinophils.

View details for Web of Science ID A1993KY80000042

View details for PubMedID 8475567
INTRAGRAFT EOSINOPHILIA AND INTERLEUKIN-5 MESSENGER-RNA ACCOMPANY LIVER ALLOGRAFT-REJECTION JEAN HAMBURGER MEMORIAL CONGRESS / 14TH INTERNATIONAL CONGRESS OF THE TRANSPLANTATION SOC Martinez, O. M., Krams, S. M., Villanueva, J. C., Ferrell, L., Lake, J., Roberts, J. P., Ascher, N. L. ELSEVIER SCIENCE INC. 1993: 126?27
View details for Web of Science ID A1993KN62100042

View details for PubMedID 8438249
T-CELL RECEPTOR-V-ALPHA GENE USE IN SEQUENTIAL LIVER ALLOGRAFT BIOPSIES JEAN HAMBURGER MEMORIAL CONGRESS / 14TH INTERNATIONAL CONGRESS OF THE TRANSPLANTATION SOC Krams, S. M., Martinez, O. M., Villanueva, J. C., Lake, J., Roberts, J. P., Ascher, N. L. ELSEVIER SCIENCE INC. 1993: 84?85
View details for Web of Science ID A1993KN62100023

View details for PubMedID 8438498
MOLECULAR ASPECTS OF EARLY STAGES OF BREAST-CANCER PROGRESSION Workshop on Chemoprevention of Breast Cancer Smith, H. S., Lu, Y., Deng, G. R., Martinez, O., Krams, S., Ljung, B. M., Thor, A., Lagios, M. WILEY-LISS. 1993: 144?152
Abstract

It is clear that breast cancer progression is associated with inactivation of a number of different recessive oncogenes. The most widely evaluated tumor suppressor gene, p53, is mutated in approximately 30-50% of sporadic breast cancers. Mutations usually occur early in malignant progression. Loss of heterozygosity (LOH) studies have identified numerous chromosomal regions where other recessive oncogenes relevant to breast cancer may be located. Each LOH is seen in a varying proportion of breast cancers and may appear either early or late in progression. High-grade ductal carcinoma in situ (DCIS) and invasive carcinoma have similar genetic lesions, showing that aberrations can occur before invasive disease. Direct evidence that the same aberrations can be acquired later in progression comes from a study of multiple metastases from the same patient; other studies found that primary invasive cancers are characterized by marked intratumor heterogeneity for each lesion examined. The model we propose to account for these results hypothesizes that multiple genetic lesions can accomplish each phenotype required for malignancy (i.e., dysregulated proliferation, invasion, angiogenesis, etc.) and that, for a given tumor, at least one aberrant gene for each phenotypic change is stochastically selected. Biological heterogeneity of breast cancer results from the stochastic acquisition of various genetic aberrations. We further propose that the lymphocytic reaction in high-grade DCIS may select for aggressive tumor subpopulations capable of escaping immune surveillance. Another aspect of tumor heterogeneity may be the multiple mechanisms employed by various tumors to escape immune surveillance.

View details for Web of Science ID A1993NE16900027

View details for PubMedID 8007693
INTRAGRAFT CYTOKINE PROFILE DURING HUMAN LIVER ALLOGRAFT-REJECTION 17TH ANNUAL MEETING OF THE AMERICAN SOC OF TRANSPLANT SURGEONS Martinez, O. M., Krams, S. M., Sterneck, M., Villanueva, J. C., Falco, D. A., Ferrell, L. D., Lake, J., Roberts, J. P., Ascher, N. L. WILLIAMS & WILKINS. 1992: 449?56
Abstract

Forty-three human liver allograft biopsies and normal liver were directly analyzed for inflammatory and immunoregulatory cytokine gene expression by polymerase chain reaction (PCR). IL-5 gene expression was predominantly present in biopsies from liver allografts with histopathological evidence of acute rejection. IL-2 gene expression was rarely observed in rejecting allografts or allografts without evidence of rejection. In contrast, IL-4 message was readily detectable in the majority of liver allografts regardless of clinical status. The inflammatory mediators IL-1 beta, TNF-alpha, and IL-6 were detected with similar frequency in rejecting allografts and allografts without evidence of rejection. These findings suggest that inflammatory and immunoregulatory cytokines are produced within the allograft. Moreover, IL-5 may play a role in the local mechanisms of liver allograft rejection.

View details for Web of Science ID A1992HD76500035

View details for PubMedID 1738940
CYTOKINE AND T-CELL RECEPTOR GENE-EXPRESSION AT THE SITE OF ALLOGRAFT-REJECTION 10TH ANNUAL MEETING OF THE AMERICAN SOC OF TRANSPLANT PHYSICIANS Krams, S. M., Falco, D. A., Villanueva, J. C., Rabkin, J., Tomlanovich, S. J., Vincenti, F., Amend, W. J., Melzer, J., Garovoy, M. R., Roberts, J. P., Ascher, N. L., Martinez, O. M. WILLIAMS & WILKINS. 1992: 151?56
Abstract

Intragraft cytokine and T cell receptor gene expression was analyzed in rejecting renal allografts by polymerase chain reaction (PCR). Message for IL-1 beta, IL-6, and TNF-alpha was detected in nephrectomy tissue with pathological evidence of acute or chronic rejection. Similarly, mRNA for both IL-6 and TNF-alpha was present in renal biopsies from acute rejecting kidneys. IL-2R, IL-4, and IL-5 mRNA was present in both rejecting and rejected kidney allografts, indicating that these cytokines may play a role in ongoing renal allograft rejection. Conversely, IL-2, IL-7, and IFN-gamma message was detected infrequently. In order to address the diversity of T cells in rejecting kidneys, we have analyzed the clonality of the TcR present within the allograft tissue. Rearranged TcR genes were identified in all allografts examined (n = 16) indicating the presence of T cells bearing the alpha/beta TcR. We have determined that there is a heterogeneous infiltration of T cells in the rejected allograft with TcR representing x = 7.47 +/- 2.4 families rearranged in samples obtained from nephrectomies, whereas x = 5.33 +/- 0.58 families were detected in samples obtained from biopsy tissue. These data indicate that (1) cytokines are produced locally which may contribute to graft cell destruction, (2) the heterogeneity of intragraft T cells during kidney allograft rejection may exist because nonspecific lymphocytes have been recruited to the site by locally produced cytokines or because T cells are responding to multiple epitopes or multiple donor antigens. Detection of intragraft cytokines and TcR may prove useful in elucidating the mechanism of rejection and therefore lead to improved immunosuppression.

View details for Web of Science ID A1992HA58300031

View details for PubMedID 1733064
MHC EXPRESSION ON HUMAN HEPATOCYTES BEFORE AND AFTER ISOLATION TRANSPLANTATION PROCEEDINGS Burke, E. C., Martinez, O. M., Freise, C. E., McVicar, J., Roberts, J. P., Ascher, N. L. 1991; 23 (1): 1428-1429
View details for Web of Science ID A1991EV39100213

View details for PubMedID 1989254
CHARACTERIZATION OF A CYCLOSPORINE-CONTAINING LIPOSOME 13TH INTERNATIONAL CONGRESS OF THE TRANSPLANTATION SOC Freise, C. E., Hong, K., Clemens, L. E., Papahadjopoulos, D., Burke, E., Martinez, O., Lake, J., Ascher, N. L., Roberts, J. P. ELSEVIER SCIENCE INC. 1991: 473?74
View details for Web of Science ID A1991EV39000186

View details for PubMedID 1824989
ALLOGENEIC HEPATOCYTES STIMULATE THE PRODUCTION OF IMMUNOREGULATORY MOLECULES IN MIXED LYMPHOCYTE HEPATOCYTE CULTURES 13TH INTERNATIONAL CONGRESS OF THE TRANSPLANTATION SOC Martinez, O. M., Burke, E. C., ALAN, T. J., Ascher, N. L. ELSEVIER SCIENCE INC. 1991: 805?6
View details for Web of Science ID A1991EV39000321

View details for PubMedID 1899316
RECOMBINANT IL-4 INHIBITS IL-6 SYNTHESIS BY ADHERENT PERIPHERAL-BLOOD CELLS-INVITRO LYMPHOKINE RESEARCH Gibbons, R., Martinez, O., Matli, M., Heinzel, F., Bernstein, M., Warren, R. 1990; 9 (3): 283-293
Abstract

IL-4 has been shown to participate in interregulatory networks with other cytokines including IL-2, IFN-gamma, IL-1 beta, and TNF-alpha. The ability of recombinant IL-4 (rIL-4) to modulate the synthesis and release of IL-6 was investigated in vitro. LPS-stimulated adherent cells (AC) from human peripheral blood secreted IL-6 as determined by ELISA and bioassay with the B9 hybridoma cell line. The secretion of IL-6 peaked between 12-16 h after LPS stimulation. The addition of rIL-4 inhibited LPS-induced IL-6 production measured at 8 h. The inhibitory effect of rIL-4 on IL-6 secretion was dose-dependent and occurred at doses as low as 0.001 U/ml (0.01 pg/ml). Recombinant IL-4 was added to the cultures for various periods and removed by aspiration of the medium and washing the cells. The subsequent LPS-induction of IL-6 secretion was reduced in cultures exposed to rIL-4 for as little as 5 minutes indicating that the effect of rIL-4 on the monocytes was rapid and irreversible. Northern blot analysis revealed that the effect of rIL4 on LPS-induced production of IL-6 by AC occurred at least in part at the level of transcription. In contrast, PMA-induced IL-6 gene transcription was not affected by rIL-4. These findings indicate that rIL-4 can regulate the synthesis of IL-6 by adherent peripheral blood mononuclear cells.

View details for Web of Science ID A1990DQ52300002

View details for PubMedID 2118969
ALTERED MONOCYTE FUNCTION IN UREMIA CLINICAL IMMUNOLOGY AND IMMUNOPATHOLOGY GIBBONS, R. A., Martinez, O. M., Garovoy, M. R. 1990; 56 (1): 66-80
Abstract

Uremia appears to suppress immune function predisposing patients to infections. When the defect in cellular immunity was studied by exposing mononuclear cells (MNC) from uremic patients and controls to tetanus toxoid, diptheria toxoid, or Candida albicans antigen in vitro, the uremic cells were far less responsive. Monocytes and T cells, which are both involved in the proliferative response to soluble antigens, were isolated from MNC of uremic patients and HLA class II matched controls and incubated with tetanus toxoid. Tetanus toxoid-pulsed uremic monocytes were unable to stimulate the proliferation of HLA identical control T lymphocytes. Lymphocytes from uremic patients, however, were stimulated by tetanus toxoid-pulsed control monocytes. Therefore, the ability of monocytes to function as accessory cells is severely affected by uremia. The uremic monocytes were FcR+, produced IL-1 beta, and expressed levels of HLA class II antigens comparable to controls. Although the biochemical defect in uremic monocytes remains unknown, the abnormality could explain many of the immunological changes of uremia.

View details for Web of Science ID A1990DL44700008

View details for PubMedID 2192826
IL-4 INHIBITS IL-2 RECEPTOR EXPRESSION AND IL-2-DEPENDENT PROLIFERATION OF HUMAN T-CELLS JOURNAL OF IMMUNOLOGY Martinez, O. M., Gibbons, R. S., Garovoy, M. R., Aronson, F. R. 1990; 144 (6): 2211-2215
Abstract

Recent studies have shown that IL-4 can affect lymphocyte responses to IL-2. To evaluate the effects of IL-4 on T cell responses to physiologically relevant stimuli, we studied normal human T cells cultured with a low concentration of anti-CD3 mAb and IL-2 in the presence and absence of added IL-4. The addition of IL-4 to cultures of T cells stimulated with anti-CD3 mAb and IL-2 reduced the proliferative response by 49 to 59%. The inhibitory effect was observed in 3-, 5-, and 7-day cultures. Inhibition was dose-dependent with maximal inhibition at concentrations greater than or equal to 5 to 10 U/ml IL-4. IL-4-mediated inhibition occurred early during the T cell response, inasmuch as addition of IL-4 after stimulation for 24 h did not result in significant inhibition. Phenotypic analyses of cells cultured in the presence of anti-CD3 mAb, IL-2, and IL-4 suggested that the mechanism of regulation by IL-4 involves the inhibition of IL-2R expression. The proportion of both CD4+ and CD8+ cells that expressed IL-2R in response to IL-2 was diminished in the presence of IL-4, although HLA-DR levels were unaffected. Soluble IL-2R was also reduced in supernatants of cultures stimulated with anti-CD3 mAb, IL-2, and IL-4 as compared to cultures stimulated with anti-CD3 mAb and IL-2. These findings indicate that when normal human T cells are stimulated in vitro in a manner that approximates a physiologic interaction with Ag in vivo, rIL-4 provides a potent inhibitory signal to IL-2 responsive cells that is likely mediated by IL-4-induced inhibition of IL-2R expression.

View details for Web of Science ID A1990CU25200026

View details for PubMedID 1968925
REDUCTION IN HLA-DR, HLA-DQ AND HLA-DP EXPRESSION BY LEU-M3+ CELLS FROM THE PERIPHERAL-BLOOD OF PATIENTS WITH THERMAL-INJURY CLINICAL AND EXPERIMENTAL IMMUNOLOGY Gibbons, R. A., Martinez, O. M., Lim, R. C., Horn, J. K., Garovoy, M. R. 1989; 75 (3): 371-375
Abstract

Monocytes that bear HLA Class II antigens, such as HLA-DR, HLA-DQ, or HLA-DP, are obligatory for many cell-mediated immunological processes. Patients with thermal injury suffer from hypoimmunity and are at risk for developing life-threatening septic episodes. To determine whether an alteration in expression of HLA Class II antigens is involved in the defect, monocytes from the peripheral blood of burn patients and controls were double-stained with anti-Leu-M3 and either anti-HLA-DR, HLA-DQ, or HLA-DP monoclonal antibodies. As analysed by flow cytometry the percentage of Leu-M3+ monocytes from the peripheral blood from patients and controls was the same. The percentage of Leu-M3+ monocytes bearing the HLA Class II antigens and the density of antigen on the monocytes, however, was significantly reduced post-burn compared with controls. In nearly all cases these changes were detected as early as 24 h post-burn before any drug therapy was implemented. In-vivo re-expression of normal levels of HLA Class II coincided with patient recovery. In-vitro exposure of post-burn Leu-M3+ cells to IFN-gamma for 72 h restored HLA Class II expression to control levels. It is possible that the reductions in HLA Class II expression may be involved in the general immunosuppression that follows thermal injury.

View details for Web of Science ID A1989T913300008

View details for PubMedID 2495202
RECOMBINANT INTERLEUKIN-4 AND PARTIALLY PURIFIED LOW-MOLECULAR-WEIGHT B-CELL GROWTH-FACTOR STIMULATE NORMAL RESTING HUMAN LYMPHOCYTES-T TRANSPLANTATION PROCEEDINGS Martinez, O., Gibbons, R., Marra, M., Garovoy, M. R. 1989; 21 (1): 74-76
View details for Web of Science ID A1989U152300023

View details for PubMedID 2784902

Olivia Martinez

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