Post Doctoral Research Associate
SCHULZE DIABETES INSTITUTE (SDI), Department of Surgery
University of Minnesota
T and B Cell Signatures Associated with Survival and Rejection of Intraportal Porcine Islet Xenografts in Immunosuppressed Cynomolgus Macaques
Amar Singh1, Wilma L Suarez-Pinzon1, Raza A Naqvi1, Christopher Burlak1, Melanie L Graham1, Bernhard J Hering1.
1Department of Surgery, Schulze Diabetes Institute, Minneapolis, MN, United States
Background: Increasing evidence indicates that both T and B lymphocytes regulate immunity to porcine islet xenografts in nonhuman primates. Improving our mechanistic understanding of the roles of T and B cell subsets in preclinical models of islet xenotransplantation and the effects of immunotherapies on these subsets will facilitate the development of refined and clinically applicable immunosuppressive protocols that promote the in-vivo expansion of regulatory phenotypes. Accordingly, the aim of this study was to compare the phenotypes of regulatory circulating T and B lymphocytes with regulatory functions and other presumably important T and B cell subsets in immunosuppressed cynomolgus macaques with stable long-term and short-term islet xenograft survival.
Methods: Intraportal porcine islet xenografts were performed in 7 diabetic cynomolgus monkeys. Immunosuppression was based on costimulation blocking biologics in all recipients. For retrospective comparisons, the recipients were divided into two cohorts (Cohort A, stable graft function ≥180 days, n=4; Cohort B, graft function <180 days, n=3). Polychromatic flow cytometry was used to determine the circulatory frequencies of the following immune cell subsets: Treg cells (CD4+CD25+FoxP3+CD127-), Natural Suppressor Cells (NSC; CD122+CD 8+), Naïve B cells (CD3-CD20+CD21+CD27-), Immature B cells ( CD3-CD19+CD27-IgM+), Mature B cells (CD3-CD19+CD27+ CD38+), Transitional B cells (CD3-CD20+CD24-CD10+), Breg cells (CD19+CD24hiCD38hi), and Activated Memory B cells (CD3-CD20+CD21+CD27+). Cell frequencies are presented as mean ± SD. Statistical significance was determined using unpaired t test with or without Welch’s correction.
Results: Pre-transplant, the frequencies of the studied immune cell subsets were not different between Cohort A and B recipients: Treg (22.9 ± 6.3 vs 18.1 ± 3.1), NSC (1.6 ± 0.56 vs 1.6 ± 1.0), Breg (1.3 ± 0.41 vs 1.6 ± 0.78), transitional B cells (2.7 ± 1.1 vs 2.8 ± 0.42), naïve/activated memory B cells (4.1 ± 1 vs 3.9 ± 0.45), and immature/mature B cells (1.51 ± 0.11 vs 2.1 ± 0.37) with p>0.05 for all comparisons. At day 110 ± 30 posttransplant, significantly higher frequencies of lymphocytes with regulatory phenotypes were found in Cohort A compared to Cohort B recipients: Treg cells (54.8 ± 17.1 vs 24.8 ± 3.1, P= 0.031), NSC (6.18 ± 2.8 vs 0.81 ± 0.25; p= 0.019), and Breg cells (6.71 ± 1.41 vs 2.51 ± 1.16; p=0.008). At day 110 ± 30 posttransplant, Cohort A compared to Cohort B recipients also showed significantly higher frequencies of transitional B cells (7.3 ± 2.6 vs 2.6 ± 0.7; p= 0.031), naïve/activated memory B cells (29.2 ± 16.7 vs 0.56 ± 0.47; p= 0.033), and immature/mature B cells (11.3 ± 6.2 vs 0.46 ± 0.16; p= 0.031). We also observed a substantial presence of intracellular IL-10 and TGF-β in regulatory T and B cells in Cohort A recipients.
Conclusions: Our preliminary findings suggest an instrumental role of lymphocytes with regulatory phenotypes in the maintenance of porcine islet xenografts in macaques immunosuppressed with costimulation blocking biologics. Furthermore, rejection in this setting is preceded by a substantial increase in the memory and activated B cells whereas sustained graft survival is associated with preponderance of immature and transitional B cells phenotypes. Ongoing studies are directed at identifying the transcripts in T and B cell subsets associated with stable islet xenograft function.
11:00 - 12:30
|Islet Immunity and Tolerance||T and B Cell Signatures Associated with Survival and Rejection of Intraportal Porcine Islet Xenografts in Immunosuppressed Cynomolgus Macaques||Plenary Room 1|
11:00 - 12:30
|Islet Immunity and Tolerance||Circulating CD8+ effector memory T cells – A candidate biomarker for imminent rejection of porcine islet xenografts in nonhuman primates||Plenary Room 1|