Islet Recipients Research Articles

Strategy for Clinical Setting of Co-transplantation of Mesenchymal Stem Cells and Pancreatic Islets.

Islet transplantation may be the most efficient therapeutic technique for patients with type 1 diabetes mellitus (T1DM). However, the clinical application of this method is faced with numerous limitations, including isolated islet apoptosis, recipient rejection, and graft vascular reconstruction. Mesenchymal stem cells (MSCs) possess anti-apoptotic, immunomodulatory, and angiogenic properties. Here, we review recent studies on co-culture and co-transplantation of islets with MSCs. We have summarized the methods of preparation of co-transplantation, especially the merits of co-culture, and the effects of co-transplantation. Accumulating experimental evidence shows that co-culture of islets with MSCs promotes islet survival, enhances islet secretory function, and prevascularizes islets through various pretransplant preparations. This review is expected to provide a reference for exploring the use of MSCs for clinical islet co-transplantation.

Human HLA-DR+CD27+ regulatory T cells show enhanced antigen-specific suppressive function.

Regulatory T cells (Tregs) have potential for the treatment of autoimmune diseases and graft rejection. Antigen specificity and functional stability are considered critical for their therapeutic efficacy. In this study, expansion of human Tregs in the presence of porcine PBMCs (xenoantigen-expanded Tregs, Xn-Treg) allowed the selection of a distinct Treg subset, coexpressing the activation/memory surface markers HLA-DR and CD27 with enhanced proportion of FOXP3+Helios+ Tregs. Compared with their unsorted and HLA-DR+CD27+ double-positive (DP) cell-depleted Xn-Treg counterparts, HLA-DR+CD27+ DP-enriched Xn-Tregs expressed upregulated Treg function markers CD95 and ICOS with enhanced suppression of xenogeneic but not polyclonal mixed lymphocyte reaction. They also had less Treg-specific demethylation in the region of FOXP3 and were more resistant to conversion to effector cells under inflammatory conditions. Adoptive transfer of porcine islet recipient NOD/SCID IL2 receptor γ-/- mice with HLA-DR+CD27+ DP-enriched Xn-Tregs in a humanized mouse model inhibited porcine islet graft rejection mediated by 25-fold more human effector cells. The prolonged graft survival was associated with enhanced accumulation of FOXP3+ Tregs and upregulated expression of Treg functional genes, IL10 and cytotoxic T lymphocyte antigen 4, but downregulated expression of effector Th1, Th2, and Th17 cytokine genes, within surviving grafts. Collectively, human HLA-DR+CD27+ DP-enriched Xn-Tregs expressed a specific regulatory signature that enabled identification and isolation of antigen-specific and functionally stable Tregs with potential as a Treg-based therapy.

310.4: Developing an algorithm for islet transplant dosing that considers both pancreas donor and islet recipient factors

310.4: Developing an algorithm for islet transplant dosing that considers both pancreas donor and islet recipient factors

Delayed graft rejection in autoimmune islet transplantation via biomaterial immunotherapy

The induction of operational immune tolerance is a major goal in beta-cell replacement strategies for the treatment of type 1 diabetes. Our group previously reported long-term efficacy via biomaterial-mediated programmed death ligand 1 (PD-L1) immunotherapy in islet allografts in nonautoimmune models. In this study, we evaluated autoimmune recurrence and allograft rejection during islet transplantation in spontaneous nonobese diabetic (NOD) mice. Graft survival and metabolic function were significantly prolonged over 60 days in recipients of syngeneic islets receiving the biomaterial-delivered immunotherapy, but not in control animals. The biomaterial-mediated PD-L1 immunotherapy resulted in delayed allograft rejection in diabetic NOD mice compared with controls. Discrimination between responders and nonresponders was attributed to the enriched presence of CD206+ program death 1+ macrophages and exhausted signatures in the cytotoxic T cell compartment in the local graft microenvironment. Notably, draining lymph nodes had similar remodeling in innate and adaptive immune cell populations. This work establishes that our biomaterial platform for PD-L1 delivery can modulate immune responses to transplanted islets in diabetic NOD mice and, thus, can provide a platform for the development of immunologic strategies to curb the allo- and autoimmune processes in beta-cell transplant recipients.

Mesenchymal Stem Cell in Pancreatic Islet Transplantation.

Pancreatic islet transplantation is a therapeutic option for achieving physiologic regulation of plasma glucose in Type 1 diabetic patients. At the same time, mesenchymal stem cells (MSCs) have demonstrated their potential in controlling graft rejection, the most fearsome complication in organ/tissue transplantation. MSCs can interact with innate and adaptive immune system cells either through direct cell-cell contact or through their secretome including exosomes. In this review, we discuss current findings regarding the graft microenvironment of pancreatic islet recipient patients and the crucial role of MSCs operation as cell managers able to control the immune system to prevent rejection and promote endogenous repair. We also discuss how challenging stressors, such as oxidative stress and impaired vasculogenesis, may jeopardize graft outcomes. In order to face these adverse conditions, we consider either hypoxia-exposure preconditioning of MSCs or human stem cells with angiogenic potential in organoids to overcome islets' lack of vasculature. Along with the shepherding of carbon nanotubes-loaded MSCs to the transplantation site by a magnetic field, these studies look forward to exploiting MSCs stemness and their immunomodulatory properties in pancreatic islet transplantation.

Matching for HLA-DR excluding diabetogenic HLA-DR3 and HLA-DR4 predicts insulin independence after pancreatic islet transplantation.

In pancreatic islet transplantation, the exact contribution of human leukocyte antigen (HLA) matching to graft survival remains unclear. Islets may be exposed to allogenic rejection but also the recurrence of type 1 diabetes (T1D). We evaluated the HLA-DR matching, including the impact of diabetogenic HLA-DR3 or HLA-DR4 matches. We retrospectively examined the HLA profile in 965 transplant recipients and 2327 islet donors. The study population was obtained from patients enrolled in the Collaborative Islet Transplant Registry. We then identified 87 recipients who received a single-islet infusion. Islet-kidney recipients, 2nd islet infusion, and patients with missing data were excluded from the analysis (n=878). HLA-DR3 and HLA-DR4 were present in 29.7% and 32.6% of T1D recipients and 11.6% and 15.8% of the donors, respectively. We identified 52 T1D islet recipients mismatched for HLA-DR (group A), 11 with 1 or 2 HLA-DR-matches but excluding HLA-DR3 and HLA- DR4 (group B), and 24 matched for HLA-DR3 or HLA-DR4 (group C). Insulin-independence was maintained in a significantly higher percentage of group B recipients from year one through five post-transplantation (p<0.01). At five-year post-transplantation, 78% of group B was insulin-independent compared to 24% (group A) and 35% (group C). Insulin-independence correlated with significantly better glycemic control (HbA1c <7%), fasting blood glucose, and reduced severe hypoglycemic events. Matching HLA-A-B-DR (≥3) independently of HLA- DR3 or HLA-DR4 matching did not improve graft survival. This study suggests that matching HLA-DR but excluding the diabetogenic HLA-DR3 and/or 4 is a significant predictor for long-term islet survival.

214.1: Strategies to Achieve Long-term Insulin Independence Using a Multimodal Approach to Beta-cell Replacement in Patients With Type I Diabetes

Purpose: Beta cell replacement, via islet or solid organ pancreas transplant, remains the best strategy to restore insulin independence in patients with Type 1 diabetes mellitus (T1DM). Insulin independence beyond ten years is limited, with calcineurin inhibitors (CNI) contributing to beta cell toxicity and impairing renal function. Here, we report long-term results for multimodal beta cell replacement including islet and pancreas after islet (PAI) transplant with CNI-free immunosuppression (IS) regimens based on Belatacept (BELA) and Efalizumab (EFA). Methods: Ten non-uremic patients with severe T1DM received their first islet cell transplant between 2007 and 2009 with CNI-sparing IS based on BELA (n=5) or EFA (n=5). Patients were monitored for a mean of 12.9 ± 1.1 years from islet transplant for beta cell function and renal function. Following islet failure, patients were considered for repeat islet infusion and/or PAI transplant. Circulating immunologic phenotype was characterized in the first year. Results: Seven of 10 patients (70%) maintained insulin independence with normalization of HbA1c (3 BELA, 4 EFA) at 10 years post-islet transplant; four patients (2 EFA, 2 BELA) achieved long-term insulin independence after one islet infusion and three additional patients (2 EFA, 1 BELA) following PAI transplantation (Figure 1, Figure 2). Six of 10 (60%) remain insulin independent currently, at a mean follow-up of 13.3 ± 1.1 years from islet transplant. Eight of 10 patients (80%) maintained renal function with less than 30% decrease in GFR and without progression to stage 4 or stage 5 CKD (Figure 2). One patient progressed to stage 4 CKD following initiation of tacrolimus for PAI, and another progressed to stage 5 CKD after islet failure and return to full insulin dependence. Patients receiving EFA demonstrated profound expansion of circulating CD4+Foxp3+ regulatory T cells (Tregs). One patient (EFA-4) demonstrated Tregs comprising 68% of all circulating CD4+ T cells, and remains insulin independent despite discontinuing all immunosuppression, thus demonstrating the first reported case of operational tolerance in islet transplantation. Conclusions: Our results describe a multi-modal, personalized approach to beta-cell replacement. In our series, repeat islet transplant is ineffective at maintaining long-term insulin independence. PAI results in durable insulin independence but is associated with deterioration of renal function secondary to CNI dependence. EFA-based immunosuppression resulted in significant expansion of Tregs following transplantation, with one islet recipient establishing operational tolerance.Juvenile Diabetes Research Foundation (4-2004-372). NIH P30 (DK63720). NIH UO1 (AIO65193). CRC UL1 (RR024131). NIH T32 (AI125222).

105.4: Impact of the COVID-19 Lockdown on Behavior, Stress, Anxiety and Glycemic Control in Patients With Beta Cell Transplantation

Introduction: Patients with severely complicated type 1 diabetes (T1D) who receive β-cell transplantation (Tx) have multiple risk factors for a severe course of coronavirus disease, including the use of immunosuppression (IS). Lockdown strategies implemented due to the COVID-19 pandemic are known to impact both mental and physical health, but this impact is expected to be even greater in patients at high risk for a severe course of COVID-19. We therefore aimed to investigate the behavioral, mental and physical implications of the nationwide lockdown in islet and pancreas transplant recipients (referred to in this abstract as β-cell Tx). Methods: In order to study the effect of the lockdown on glycemic control, all patients with T1D and an islet transplantation or pancreas transplantation with non-optimal graft function according to the Igls criteria using IS were eligible. As a control group, patients with T1D without IS were included. Lockdown behavior and self-reported changes in anxiety, stress, physical activity, weight, and glycemic control were assessed using questionnaires. HbA1c and continuous glucose monitoring (CGM) metrics during lockdown were compared to measurements before lockdown. Results: Islet and pancreas (β-cell) Tx recipients (n = 51, age 55 (48 – 59) years, BMI 23.3 (20.9 – 27.4) kg/m2, diabetes duration 42 (34 – 48) years) adhered more stringently to lockdown measures compared to patients with T1D alone (n = 272, age 53 (37 – 62) years, BMI 25.2 (23.0 – 28.0) kg/m2, diabetes duration 27 (15 – 39) years). In β-cell Tx recipients as compared to T1D, 52.1% vs 18.3% (p <0.001) reported not going out for groceries and 45.8% vs 14.0% (p <0.001) reported not leaving the house at all. Fear of coronavirus infection was higher in β-cell Tx recipients (VAS 5.0 (3.0 – 7.0) vs 3.0 (2.0 – 5.0), p=0.004) and glycemic control worsened during lockdown as assessed by HbA1c (ΔHbA1c +1.67 ± 8.74 vs -1.72 ± 6.15 mmol/mol, p=0.006) as well as CGM (Δ time in range β-cell Tx -4.5% (-6.0% – 1.5%) vs T1D 3.0% (-2.0% – 6.0%), p=0.038; Δ time above range β-cell Tx 5.5% (-0.5% – 7.5%) vs T1D -3.0% (-7.5% – 3.0%), p=0.025). Among β-cell Tx recipients, 29.2% self-reported more difficulty with glycemic control, 26.8% increased insulin use, 40.0% less physical activity, 41.7% weight gain, 29.2% increased anxiety and 33.3% increased stress since the start of lockdown. Having had a β-cell Tx was the most important predictor of not leaving the house during the COVID-19 lockdown. Conclusions: The COVID-19 pandemic and subsequent lockdown add additional fear of infection, deterioration of glycemic control and more stringent social isolation behavior when patients with T1D also have an islet or pancreas transplant. These patients undertake less physical activity and experience more weight gain, stress and anxiety. Health care professionals should be aware of these behavioral, mental and physical implications to be able to provide extra support.

207.5: Impact of HLA Donor Specific Antibodies in the Era of Cell Therapy: Clinical Trajectories in Islet Allotransplantation Illustrate a Challenging Model

Introduction: Islet transplantation is a unique paradigm in the case of organ transplantation, since multiple injections and donors are required to achieve complete insulin-independence. Preformed or de novo DSAs may target one or several donor islets, which complexifies the analysis of their impact. Our objective is to discriminate illustrative clinical trajectories which can occur after transplantation. Methods: Adult type 1 diabetic pancreatic islet recipients transplanted between January 2005 and December 2018, were included in a monocentric observational study. Only patients with available sera tested by Luminex for Class I and II anti-HLA antibodies during their whole follow-up were analyzed. Results: 32 patients had a complete HLA antibodies screening during follow-up. Among them, 7 patients presented with DSA. Five clinical trajectories were identified according to the MFI-value, and the presence of preformed DSA, de novo DSA, or both. To simplify, we provide specific clinical-based trajectories considering an islet transplantation with 1- high MFI preformed DSA, 2- high MFI de novo DSA, 3- low-MFI preformed DSA, 4-low de novo DSA, and 5- a de novo DSA which becomes a preformed DSA because of multiple islet infusions. We analyzed also the specific case of islet-after-kidney transplantation. Recipients transplanted with preformed DSA had all favorable islet transplantation outcomes. The prognosis of de novo DSA was associated with unfavorable outcomes when following markers of unsteady glycemic control. Conclusion: Clinical-based trajectories and evidence from the literature reveal that islet transplantation with preformed DSA, even with high MFI values, can be associated with favorable outcomes. The fate of recipient presenting with de novo DSA is more questionable as both clinical data and preclinical studies show separate outcomes. Clinical phenotype and unsteady glycemic control before the emergence of DSA may lead the clinician to detect rejection.I-SITE ULNE.

Prolonged Islet Allograft Function is Associated With Female Sex in Patients After Islet Transplantation.

Islet transplantation (ITx) has proved to be effective in preventing severe hypoglycemia and improving metabolic control in selected subjects with type 1 diabetes. Long-term graft function remains a challenge. Estrogens have been shown to protect β cells from metabolic stresses and improve revascularization of transplanted human islets in the mouse. We aimed to evaluate the influence of sex in allograft survival of ITx recipients. We analyzed a retrospective cohort of ITx recipients (n = 56) followed-up for up to 20 years. Allograft failure was defined as a stimulated C-peptide <0.3 ng/mL during a mixed-meal tolerance test. Subjects were divided into recipients of at least 1 female donor (group 1) and recipients of male donors only (group 2). Group 1 subjects (n = 25) were aged 41.5 ± 8.4 years and group 2 subjects (n = 22) 45.9 ± 7.3 years (P = 0.062). Female recipient frequency was 44.8% (n = 13) in group 1 and 55.2% (n = 16) in group 2 (P = 0.145). Group 2 developed graft failure earlier than group 1 (680 [286-1624] vs 1906 [756-3256] days, P = 0.038). We performed additional analyses on female recipients only from each group (group 1, n = 16; group 2, n = 20). Female recipients in group 1 exhibited prolonged allograft function compared with group 2, after adjustment for confounders (odds ratio, 28.6; 95% CI, 1.3-619.1; P < 0.05). Recipients of islets from at least 1 female donor exhibited prolonged graft survival compared with recipients of islets from exclusively male donors. In addition, female recipients exhibited prolonged survival compared with male recipients following ITx of at least 1 female donor.

Extended survival versus accelerated rejection of nonhuman primate islet allografts: Effect of mesenchymal stem cell source and timing

Mesenchymal stem cells (MSC) have been shown to be immunomodulatory, tissue regenerative, and graft promoting; however, several questions remain with regard to ideal MSC source and timing of administration. In this study, we utilized a rigorous preclinical model of allogeneic islet cell transplantation, incorporating reduced immune suppression and near to complete mismatch of major histocompatibility antigens between the diabetic cynomolgus monkey recipient and the islet donor, to evaluate both the graft promoting impact of MSC source, that is, derived from the islet recipient, the islet donor or an unrelated third party as well as the impact of timing. Co-transplant of MSC and islets on post-operative day 0, followed by additional IV MSC infusions in the first posttransplant month, resulted in prolongation of rejection free and overall islet survival and superior metabolic control for animals treated with recipient as compared to donor or third-party MSC. Immunological analyses demonstrated that infusion of MSC from either source did not prevent alloantibody formation to the islet or MSC donor; however, treatment with recipient MSC resulted in significant downregulation of memory T cells, decreased anti-donor T cell proliferation, and a trend toward increased Tregulatory:Tconventional ratios.

Adapting Protocols or Models for Use in Insulin-Requiring Diabetes and Islet Transplant Recipients.

The authors’ perspective is described regarding modifications made in their clinic to glucose challenge protocols and mathematical models in order to estimate insulin secretion, insulin sensitivity and glucose effectiveness in patients living with Insulin-Requiring Diabetes and patients who received Pancreatic Islet Transplants to treat Type I diabetes (T1D) with Impaired Awareness of Hypoglycemia. The evolutions are described of protocols and models for use in T1D, and Insulin-Requiring Type 2 Diabetes (T2D) that were the basis for studies in the Islet Recipients. In each group, the need for modifications, and how the protocols and models were adapted is discussed. How the ongoing application of the adaptations is clarifying the Islet pathophysiology in the Islet Transplant Recipients is outlined.

41-OR: Roles of Transcription Factor EB (TFEB) in ß-Cell Function and Survival Under Stress

Autophagy can help protect stressed β-cells. We investigated if disruption of Transcription Factor EB (Tfeb), a transcriptional master-regulator of lysosomes and autophagy, contributes to β-cell failure and death under various ER stress-inducing conditions. Short-term β-cell autophagy-deficiency (inducible Atg5 knockout) reduced β-cell survival under hypoxic stress in vitro and impaired the ability of syngeneic marginal mass islet grafts to maintain glucose homeostasis in streptozotocin (STZ)-induced diabetic mice, with &amp;gt;80% of recipients of Atg5-deficient islets returning to diabetes within 10 days post-transplant (vs. 30% of controls, p=0.05). Hypoxic culture activated pro-apoptotic ER stress pathways, and confocal analysis of islet cells from LC3-RFP-EGFP autophagy reporter mice demonstrated that autophagic flux was interrupted. Hypoxia-induced impairment of autophagy was associated with reduced lysosomal cathepsin B activity and a significant loss of Tfeb, both of which could be partially rescued by Torin-1. Lipotoxicity and thapsigargin-induced ER-stress similarly reduced Tfeb protein and cell viability, suggesting ER stress-specific mechanisms may drive dysregulation of Tfeb and lysosomes. Conversely, overexpression of Tfeb:GFP reduced thapsigargin-induced death in MIN6 cells. To directly study the consequences of Tfeb loss we generated mice with Ins1Cre-driven, β-cell-specific, Tfeb deletion (TfebβKO). Notably, both male and female TfebβKO mice had normal glucose tolerance, but cultured Tfeb KO β-cells showed impaired lysosomal cathepsin B activity and TfebβKO mice had worsened glucose tolerance compared to Tfeb flox littermates following injection with multiple low-doses STZ. In summary, autophagy protects β-cells under hypoxia but is impaired under prolonged hypoxic stress, possibly through Tfeb dysregulation. Our data further suggest loss of Tfeb is a common feature of ER stress and that this loss exacerbates stress-induced β-cell failure and death. Disclosure Y. Zou: None. D. Pasula: None. B. Verchere: Advisory Panel; Self; Integrated Nanotherapeutics Inc., Sirona Biochem, Stock/Shareholder; Self; Integrated Nanotherapeutics Inc. D. S. Luciani: None. Funding JDRF (2-2013-50)

Immunological Monitoring in Beta Cell Replacement: Towards a Pathophysiology-Guided Implementation of Biomarkers.

Grafted beta cells are lost because of recurrence of T1D and/or allograft rejection, two conditions diagnosed with pancreas graft biopsy, which is invasive and impossible in case of islet transplantation. This review synthetizes the current pathophysiological knowledge and discusses the interest of available immune biomarkers. Despite the central role of auto-(recurrence of T1D) and allo-(T-cell mediated rejection) immune cellular responses, the latter are not directly monitored in routine. In striking contrast, there have been undisputable progresses in monitoring of auto and alloantibodies. Except for pancreas recipients in whom anti-donor HLA antibodies can be directly responsible for antibody-mediated rejection, autoantibodies (and alloantibodies in islet recipients) have no direct pathogenic effect. However, their fluctuation offers a surrogate marker for the activation status of T cells (because antibody generation depends on T cells). This illustrates the necessity to understand the pathophysiology when interpreting a biomarker and selecting the appropriate treatment.

Intestinal Delivery of Proinsulin and IL-10 via Lactococcus lactis Combined With Low-Dose Anti-CD3 Restores Tolerance Outside the Window of Acute Type 1 Diabetes Diagnosis.

A combination treatment (CT) of proinsulin and IL-10 orally delivered via genetically modified Lactococcus lactis bacteria combined with low-dose anti-CD3 (aCD3) therapy successfully restores glucose homeostasis in newly diagnosed non-obese diabetic (NOD) mice. Tolerance is accompanied by the accumulation of Foxp3+ regulatory T cells (Tregs) in the pancreas. To test the potential of this therapy outside the window of acute diabetes diagnosis, we substituted autoimmune diabetic mice, with disease duration varying between 4 and 53 days, with syngeneic islets at the time of therapy initiation. Untreated islet recipients consistently showed disease recurrence after 8.2 ± 0.7 days, while 32% of aCD3-treated and 48% of CT-treated mice remained normoglycemic until 6 weeks after therapy initiation (P < 0.001 vs. untreated controls for both treatments, P < 0.05 CT vs. aCD3 therapy). However, mice that were diabetic for more than 2 weeks before treatment initiation were less efficient at maintaining normoglycemia than those treated within 2 weeks of diabetes diagnosis, particularly in the aCD3-treated group. The complete elimination of endogenous beta cell mass with alloxan at the time of diabetes diagnosis pointed toward the significance of continuous feeding of the islet antigen proinsulin at the time of aCD3 therapy for treatment success. The CT providing proinsulin protected 69% of mice, compared to 33% when an irrelevant antigen (ovalbumin) was combined with aCD3 therapy, or to 27% with aCD3 therapy alone. Sustained tolerance was accompanied with a reduction of IGRP+CD8+ autoreactive T cells and an increase in insulin-reactive (InsB12–20 or InsB13–2) Foxp3+CD4+ Tregs, with a specific accumulation of Foxp3+ Tregs around the insulin-containing islet grafts after CT with proinsulin. The combination of proinsulin and IL-10 via oral Lactococcus lactis with low-dose aCD3 therapy can restore tolerance to beta cells in autoimmune diabetic mice, also when therapy is started outside the window of acute diabetes diagnosis, providing persistence of insulin-containing islets or prolonged beta cell function.

Transplantation of PEGylated islets enhances therapeutic efficacy in a diabetic nonhuman primate model.

Islet cell transplantation can lead to insulin independence, reduced hypoglycemia, and amelioration of diabetes complications in patients with type 1 diabetes. The systemic delivery of anti-inflammatory agents, while considered crucial to limit the early loss of islets associated with intrahepatic infusion, increases the burden of immunosuppression. In an effort to decrease the pharmaceutical load to the patient, we modified the pancreatic islet surface with long-chain poly(ethylene glycol) (PEG) to mitigate detrimental host-implant interactions. The effect of PEGylation on islet engraftment and long-term survival was examined in a robust nonhuman primate model via three paired transplants of dosages 4300, 8300, and 10000islet equivalents per kg body weight. A reduced immunosuppressive regimen of anti-thymocyte globulin induction plus tacrolimus in the first posttransplant month followed by maintenance with sirolimus monotherapy was employed. To limit transplant variability, two of the three pairs were closely MHC-matched recipients and received MHC-disparate PEGylated or untreated islets isolated from the same donors. Recipients of PEGylated islets exhibited significantly improved early c-peptide levels, reduced exogenous insulin requirements, and superior glycemic control, as compared to recipients of untreated islets. These results indicate that this simple islet modification procedure may improve islet engraftment and survival in the setting of reduced immunosuppression.

217-OR: Islet Transplantation vs. Standard of Care for Type 1 Diabetes Complicated by Severe Hypoglycemia from the Collaborative Islet Transplant Registry (CITR) and T1D Exchange (T1DX) Registry

Islet transplantation is an emerging treatment for T1D, in particular for those with long standing disease experiencing severe hypoglycemia events (SHE). A phase 3 trial of islet transplantation in T1D complicated by SHE demonstrated &amp;gt; 80% of islet recipients achieved HbA1c &amp;lt; 7.0% and absence of SHE at 1 year. To better understand the long-term benefit for glycemic control and risk of immunosuppression to kidney function associated with islet transplantation vs. standard of care, we selected patients from CITR with at least one SHE in the year prior to transplant, and compared them to patients from T1DX with at least one SHE in the prior year and followed over 5 years. The cohorts from CITR (n = 58) and T1DX (n = 213) were aged 42 ± 9 and 43 ± 17 with disease duration 29 ± 10 and 26 ± 15 years (P &amp;lt; 0.01) and experienced 5 ± 11 and 2 ± 1 SHE (P &amp;lt; 0.01) in the year prior to baseline with HbA1c 7.3 ± 1.0 vs. 7.8 ± 1.5 (P &amp;lt; 0.001). Mean HbA1c decreased to 6.3% vs. ranging 7.8-7.6% over 5 years (P &amp;lt; 0.001), and the % experiencing recurrent SHE was less in CITR than T1DX (P &amp;lt; 0.01), such that 78-77% vs. 23-28% met the primary endpoint of HbA1c &amp;lt; 7.0% and absence of SHE (P &amp;lt; 0.001). Insulin requirements were slightly less at baseline in CITR than T1DX (0.52 ± 0.14 vs. 0.64 ± 0.31 U/kg/d; P &amp;lt; 0.001), and decreased to 0.06-0.17 vs. remaining 0.66-0.56 U/kg/d over 5 years (P &amp;lt;0.001) with 80-48% insulin independent in CITR. GFR was 92 ± 17 and 90 ± 29 ml/min/1.73 m2 at baseline, and decreased to 83-78 vs. 87-81 ml/min/1.73 m2 over 5 years (P &amp;lt; 0.001). These data demonstrate long-term durability of islet transplantation to achieve near-normal glycemic control in the absence of SHE more often than that observed with standard of care delivered by specialized diabetes practice. Initial reduction in GFR observed during the first-year post-transplant may be attributable to the initiation of immunosuppression, but was not associated with further decline during 5 year follow-up. Disclosure M.R. Rickels: None. N.C. Foster: None. C.M. Ballou: None. B.J. Hering: Other Relationship; Self; Diabetes-Free, Inc. R. Alejandro: None. R. Beck: Other Relationship; Self; Abbott Laboratories, Ascensia Diabetes Care, Bigfoot Biomedical, Dexcom, Inc., Insulet Corporation, Lilly Diabetes, Roche Diabetes Care, Tandem Diabetes Care. K. Miller: None. F.B. Barton: None. Funding National Institutes of Health; Leona M. and Harry B. Helmsley Charitable Trust

Peri-graft porcine-specific CD4+ FoxP3+ regulatory T cells by CD40-CD154 blockade prevented the rejection of porcine islet graft in diabetic mice.

Anti-CD154 monoclonal antibody (mAb) treatment has been known to have potential to induce immune tolerance in organ transplantation. Several studies have suggested the involvement of CD4+ regulatory T cells (Treg s) in xeno-immune tolerance. However, the characteristics of Treg s and the mechanisms of their regulatory functions in islet xenotransplantation have not been clearly defined. Adult porcine islet cells were isolated and purified, and were transplanted under the kidney capsule of diabetic C57BL/6J mice with the administration of 0.5mg/mouse of anti-CD154 mAb on 0, 1, 3, 5, and 7days post-transplantation (DPT). The graft survival was monitored by blood glucose level. The islet graft and recipients' cells were analyzed by immunohistochemistry (IHC), flow cytometry, enzyme-linked immunosorbent spot (ELISPOT) assay, and mixed-lymphocyte reaction. Short-term anti-CD154 mAb monotherapy enabled the porcine islet graft to survive indefinitely in diabetic mice (n=18). Immunohistochemical staining showed significantly higher ratio of CD4+ Foxp3+ Treg s in the peri-graft site, but not in the spleen and kidney-draining lymph node of the recipient mice. Depletion of Treg s evoked graft rejection, and adoptive transfer of Treg s from anti-CD154 mAb-treated recipients provided protection to the graft from rejection. These Treg s showed more potent suppressive capacity than those from the untreated control and were found to be porcine antigen-specific. In this study, we showed that anti-CD154 mAb monotherapy resulted in indefinite porcine islet graft survival in mice. The porcine-specific CD4+ Foxp3+ Treg s in the peri-graft site played the critical role in the protection of islet graft from rejection, which suggests a prospective immunosuppressive strategy for islet xenotransplantation.

Mesenteric lymph nodes as alternative site for pancreatic islet transplantation in a diabetic rat model

BackgroundIslet transplantation has progressively become a safe alternative to pancreas transplantation for the treatment of type 1 diabetes. However, the long-term results of islet transplantation could be significantly increased by improving the quality of the islet isolation technique even exploring alternative islet transplantation sites to reduce the number of islets required to mitigate hyperglycemia. The goal of the study was to test the lymph node as a suitable anatomical location for islet engraftment in a rodent model.MethodsForty Lewis rats, 6–8 weeks old, body weight 250–300 g, have been used as islet donors and recipients in syngeneic islet transplantation experiments. Ten rats were rendered diabetic by one injection of 65 mg/Kg of streptozotocin. After pancreas retrieval from non diabetic donors, islet were isolated and transplanted in the mesenteric lymph nodes of 7 diabetic rats. Rats were followed for 30 days after islet transplantation.ResultsA total of 7 islet transplantations in mesenteric lymph nodes have been performed. Two rats died 24 and 36 h after transplantation due to complications. No transplanted rat acquired normal glucose blood levels and insulin independence after the transplantation. However, the mean blood levels of glycemia were significantly lower in transplanted rats compared with diabetic rats (470.4 mg/dl vs 605 mg/dl, p 0.04). Interestingly, transplanted rats have a significant weight increase after transplantation compared to diabetic rats (mean value 295 g in transplanted rats vs 245 g in diabetic rats, p < 0.05), with an overall improvement of social activities and health. Immunohistochemical analysis of the 5 mesenteric lymph nodes of transplanted rats demonstrated the presence of living islets in one lymph node.ConclusionsAlthough islet engraftment in lymph nodes is possible, islet transplantation in lymph nodes in rats resulted in few improvements of glucose parameters.

Effects of remote ischaemic preconditioning on intraportal islet transplantation in a rat model.

Remote ischaemic preconditioning (RIPC), which is the intermittent interruption of blood flow to a site distant from the target organ, is known to improve solid organ resistance to ischaemia-reperfusion injury. This procedure could be of interest in islet transplantation to mitigate hypoxia-related loss of islet mass after isolation and transplantation. Islets isolated from control or RIPC donors were analyzed for yield, metabolic activity, gene expression and high mobility group box-1 (HMGB1) content. Syngeneic marginal mass transplantation was performed in four streptozotocin-induced diabetic groups: control, RIPC in donor only, RIPC in recipient only, and RIPC in donor and recipient. Islets isolated from RIPC donors had an increased yield of 20% after 24h of culture compared to control donors (P=0.007), linked to less cell death (P=0.08), decreased expression of hypoxia-related genes (Hif1a P=0.04; IRP94 P=0.008), and increased intra-cellular (P=0.04) and nuclear HMGB1. The use of RIPC in recipients only did not allow for reversal of diabetes, with increased serum HMGB1 at day 1; the three other groups demonstrated significantly better outcomes. Performing RIPC in the donors increases islet yield and resistance to hypoxia. Validation is needed, but this strategy could help to decrease the number of donors per islet recipient.