Peter Zucker

Neonatal activation of CD28 signaling overcomes T cell anergy and prevents autoimmune diabetes by an IL-4-dependent mechanism.

Optimal T cell responsiveness requires signaling through the T cell receptor (TCR) and CD28 costimulatory receptors. Previously, we showed that T cells from autoimmune nonobese diabetic (NOD) mice display proliferative hyporesponsiveness to TCR stimulation, which may be causal to the development of insulin-dependent diabetes mellitus (IDDM). Here, we demonstrate that anti-CD28 mAb stimulation restores complete NOD T cell proliferative responsiveness by augmentation of IL-4 production. Whereas neonatal treatment of NOD mice with anti-CD28 beginning at 2 wk of age inhibits destructive insulitis and protects against IDDM by enhancement of IL-4 production by islet-infiltrating T cells, administration of anti-CD28 beginning at 5-6 wk of age does not prevent IDDM. Simultaneous anti-IL-4 treatment abrogates the preventative effect of anti-CD28 treatment. Thus, neonatal CD28 costimulation during 2-4 wk of age is required to prevent IDDM, and is mediated by the generation of a Th2 cell-enriched nondestructive environment in the pancreatic islets of treated NOD mice. Our data support the hypothesis that a CD28 signal is requisite for activation of IL-4-producing cells and protection from IDDM.

ICA69null Nonobese Diabetic Mice Develop Diabetes, but Resist Disease Acceleration by Cyclophosphamide

ICA69 (islet cell Ag 69 kDa) is a diabetes-associated autoantigen with high expression levels in β cells and brain. Its function is unknown, but knockout of its Caenorhabditis elegans homologue, ric-19, compromised neurotransmission. We disrupted the murine gene, ica-1, in 129-strain mice. These animals aged normally, but speed-congenic ICA69null nonobese diabetic (NOD) mice developed mid-life lethality, reminiscent of NOD-specific, late lethal seizures in glutamic acid decarboxylase 65-deficient mice. In contrast to wild-type and heterozygous animals, ICA69null NOD congenics fail to generate, even after immunization, cross-reactive T cells that recognize the dominant Tep69 epitope in ICA69, and its environmental mimicry Ag, the ABBOS epitope in BSA. This antigenic mimicry is thus driven by the endogenous self Ag, and not initiated by the environmental mimic. Insulitis, spontaneous, and adoptively transferred diabetes develop normally in ICA69null NOD congenics. Like glutamic acid decarboxylase 65, ICA69 is not an obligate autoantigen in diabetes. Unexpectedly, ICA69null NOD mice were resistant to cyclophosphamide (CY)-accelerated diabetes. Transplantation experiments with hemopoietic and islet tissue linked CY resistance to ICA69 deficiency in islets. CY-accelerated diabetes involves not only ablation of lymphoid cells, but ICA69-dependent drug toxicity in β cells that boosts autoreactivity in the regenerating lymphoid system.

Characterization of the role of major histocompatibility complex in type 1 diabetes recurrence after islet transplantation.

Background. Major histocompatibility complex (MHC) molecules are essential determinants of &bgr;-cell destruction in type 1 diabetes (T1D). MHC class I- or class II-null nonobese diabetic (NOD) mice do not spontaneously develop autoimmune diabetes and are resistant to adoptive transfer of disease. Both CD4+ and CD8+ T cells are associated with graft destruction after syngeneic islet transplantation. MHC molecules within the graft (i.e., on &bgr;-cells or donor lymphocytes) may influence the interactions between antigen presenting cells and effector T cells and, therefore, the survival outcome of the graft. Methods. Donor islets from NOD mice deficient in one or both of &bgr;2-microglobulin and class II transactivator genes were transplanted into diabetic NOD mice. Immunohistochemistry was performed to identify the phenotype of infiltrating cells and to assess graft insulin production. The presence of cytokines in the grafts was assayed by reverse transcription polymerase chain reaction. Results. MHC class II-null islets demonstrated rates of rejection comparable with control wild-type (wt) islets. In contrast, MHC class I- and II-null islets demonstrated indefinite survival (over 100 days). Infiltrates of both failed and surviving grafts were comprised of cytotoxic lymphocytes (CTL), helper T cells, and macrophages. Grafts also showed the presence of both Th1- and Th2-type cytokines (interleukin [IL]-2, IL-4, IL-10, and interferon-&ggr;), independent of graft status. Conclusions. These results demonstrate the primary importance of MHC class I molecules in the pathogenesis of diabetes recurrence postislet transplantation. Conversely, MHC class II expression is not a necessary mechanistic component of transplant destruction. In addition, these results implicate MHC class I-restricted CTLs but not MHC class II-restricted T cells in disease recurrence.

Transplanted MHC class I-deficient nonobese diabetic mouse islets are protected from autoimmune injury in diabetic nonobese recipients.

The injury of transplanted islets may occur by both autoimmune and alloimmune processes directed against MHC targets. To examine the role of MHC class I in islet graft injury, we transplanted syngeneic and allogeneic &bgr;2-microglobulin-deficient islets into diabetic nonobese diabetic (NOD) mice. Loss of graft function was observed within 14 days using allogeneic C57BL/6 and BALB/c MHC class I deficient as well as wild-type MHC class I-bearing NOD donor islets. However, islets isolated from MHC class I-deficient NOD mice (NOD-B2 m-/-) survived indefinitely when transplanted under the kidney capsule of diabetic NOD recipients. Transplanted NOD-B2 m-/- islets were surrounded by a nondestructive periinsular infiltrate that expressed interleukin-4 in addition to interferon-&ggr;. These studies demonstrate the primary role of MHC class I molecules in causing autoimmune destruction or recurrent diabetes in transplanted islets.

Insulin Resistance Prevented by Portal Delivery of Insulin in Rats With Renal Subcapsular Islet Grafts

We determined the metabolic effects of insulin derived from renal subcapsular islet grafts, either with systemic delivery of insulin through renal venous drainage (REN) or with portal delivery of insulin after renal vein–to–superior mesenteric vein anastomosis (RMA), in streptozotocin-induced diabetic Lewis rats, in comparison with normal rats. After gavage glucose, the plasma glucose responses were similar to normal in REN and RMA rats; however, hyperinsulinemia occurred in REN rats (area under the concentration curves [AUCs] of insulin, 27 ± 3 nmol · l−1 · min) in comparison with RMA (14 ± 2) and normal rats (19 ± 2), P < 0.003, with no difference in C-peptide responses. The ratio of AUC C-peptide to AUC insulin was lower in REN (2.0 ± 0.2) than in RMA (3.4 ± 0.3) and normal animals (3.2 ± 0.3), P < 0.0005. In euglycemic-hyperinsulinemic clamp studies using the same insulin infusion rate (10 pmol · kg−1 · min−1), insulin resistance was found in REN animals (mean glucose infusion rate [GIR], REN: 7.5 ± 1.2; RMA: 12.0 ± 1.2; normal: 12.7 ± 1.0 mg · kg−1 · min−1; P < 0.008), with higher steady-state insulin levels in REN (554 ± 63 pmol/l) than in RMA (291 ± 26) and normal rats (269 ± 60), P < 0.0001. With matching steady-state insulin levels in RMA and REN rats during infusion of insulin at 20 pmol · kg−1 · min−1 in RMA rats (steady-state insulin 623 ± 64 pmol/l), GIR was 15.7 ± 0.7 mg · kg−1 · min−1. Thus, systemic delivery of insulin from islet grafts is associated with hyperinsulinemia, insulin resistance, and decreased metabolic clearance of insulin. These abnormalities are prevented by portal delivery of insulin from islet grafts in the same site. The findings are consistent with the hypothesis that portal delivery of insulin is important in maintenance of normal whole-body insulin sensitivity.

Role of donor MHC class III genes in the destruction of transplanted islets in NOD mice.

Abstract: MHC class III genes are important in immune regulation and inflammation, and the gene products of this region are well conserved between species. Their role in diabetes is, however, unknown. We used islets from NOD mice that lacked expression of both MHC class I and class II molecules to test the effect of class III differences on the injury of transplanted NOD islets. Loss of islet MHC class I was highly protective, while deletion of MHC class II had no benefit on islet survival. However the combined absence of both MHC class I and class II expression by NOD islets resulted in a delayed form of injury, when islets were transplanted to NOD mice. As neither MHC class I or II molecules were expressed by donor islet tissue, these results suggest a previously unrecognized and important contribution of MHC class III differences on islet injury following transplantation.