Excessive secretion of glucagon is a major contributor to the development of diabetic hyperglycemia. Secretion of glucagon is regulated by various nutrients, with glucose being a primary determinant of the rate of alpha cell glucagon secretion. The intra-islet action of insulin is essential to exert the effect of glucose on the alpha cells since, in the absence of insulin, glucose is not able to suppress glucagon release in vivo. However, the precise mechanism by which insulin suppresses glucagon secretion from alpha cells is unknown. In this study, we show that insulin induces activation of GABAA receptors in the alpha cells by receptor translocation via an Akt kinase-dependent pathway. This leads to membrane hyperpolarization in the alpha cells and, ultimately, suppression of glucagon secretion. We propose that defects in this pathway(s) contribute to diabetic hyperglycemia.

Type 2 diabetic subjects manifest both disordered insulin action and abnormalities in their pancreatic islet cells. Whether the latter represents a primary defect or is a consequence of the former is unknown. To examine the ␤-cell mass and function of islets from type 2 diabetic patients directly, we isolated islets from pancreata of type 2 diabetic cadaveric donors (n ‫؍‬ 14) and compared them with islets from normal donors (n ‫؍‬ 14) matched for age, BMI, and cold ischemia time. The total recovered islet mass from type 2 diabetic pancreata was significantly less than that from nondiabetic control subjects (256,260 islet equivalents [2,588 IEq/g pancreas] versus 597,569 islet equivalents [6,037 IEq/g pancreas]). Type 2 diabetic islets were also noted to be smaller on average, and histologically, islets from diabetic patients contained a higher proportion of glucagon-producing ␣-cells. In vitro study of islet function from diabetic patients revealed an abnormal glucosestimulated insulin release response in perifusion assays. In addition, in comparison with normal islets, an equivalent number of type 2 diabetic islets failed to reverse hyperglycemia when transplanted to immunodeficient diabetic mice. These results provide direct evidence for abnormalities in the islets of type 2 diabetic patients that may contribute to the pathogenesis of the disease. Diabetes 53: 624 -632, 2004

A complementary DNA for a glucagon-like peptide-1 receptor was isolated from a human pancreatic islet cDNA library. The isolated clone encoded a protein with 90% identity to the rat receptor. In stably transfected fibroblasts, the receptor bound I n normal individuals, oral glucose absorption induces a greater insulin secretory response than an isoglycemic intravenous glucose infusion. This greater effect of oral glucose results from the secretion of intestinal hormones, which, at the level of the pancreatic p-cells, potentiate glucose-induced insulin secretion (1,2). These

A complementary DNA for a glucagon-like peptide-1 receptor was isolated from a human pancreatic islet cDNA library. The isolated clone encoded a protein with 90% identity to the rat receptor. In stably transfected fibroblasts, the receptor bound [ 125 I]GLP-1 with high affinity (K^ = 0.5 nM) and was coupled to adenylate cyclase as detected by a GLP-1-dependent increase in cAMP production (EC 50 = 93 pM). Two peptides from the venom of the lizard Heloderma suspectum, exendin-4 and exendin-(9-39), displayed similar ligand binding affinities to the human GLP-1 receptor. Whereas exendin-4 acted as an agonist of the receptor, inducing cAMP formation, exendin-(9-39) was an antagonist of the receptor, inhibiting GLP-1-induced cAMP production. Because GLP-1 has been proposed as a potential agent for treatment of NIDDM, our present data will contribute to the characterization of the receptor binding site and the development of new agonists of this receptor. Diabetes 42:1678-82, 1993 I n normal individuals, oral glucose absorption induces a greater insulin secretory response than an isoglyce-mic intravenous glucose infusion. This greater effect of oral glucose results from the secretion of intestinal hormones, which, at the level of the pancreatic p-cells, potentiate glucose-induced insulin secretion (1,2). These

Summary Regulatory B cells (Bregs) have been shown to play a critical role in immune homeostasis and in autoimmunity models. We have recently demonstrated that combined anti-TIM-1 and anti-CD45RB antibody treatment results in tolerance to full MHC-mismatched islet allografts in mice by generating Bregs that are necessary for tolerance. Bregs are antigen-specific and are capable of transferring tolerance to untreated, transplanted animals. Here we demonstrate that adoptively transferred Bregs require the presence of Tregs to establish tolerance, and that adoptive transfer of Bregs increases the number of Tregs. Interaction with Bregs in vivo induces significantly more Foxp3 expression in CD4+CD25− T cells than with naive B cells. We also show that Bregs express the TGF-β associated latency-associated peptide (LAP) and that Breg-mediated graft prolongation post-adoptive transfer is abrogated by neutralization of TGF-β activity. Regulatory B cells, like regulatory T cells, demonstrate preferential expression of both CCR6 and CXCR3. Collectively, these findings suggest that in this model of antibody-induced transplantation tolerance, Bregs promote graft survival by promoting Treg development, possibly via TGF-β production.

A reduction in ␤-cell mass is an important causative factor in type 1 and type 2 diabetes. Glucagon-like peptide-1 (GLP-1) and the long-acting agonist exendin 4 (Ex-4) expand ␤-cell mass by stimulating neogenesis and proliferation. In the partial pancreatectomy (Ppx) model, exogenous Ex-4 promotes islet regeneration, leading to sustained improvement in glucose tolerance. In this study, we investigate the potential role of endogenous GLP-1 in islet growth. We examined ␤-cell mass regeneration after 70% Ppx in mice receiving the GLP-1 antagonist Ex9-39 and in GLP-1R ؊/؊ mice. In Ex9-39 -treated sham-operated mice, persistent fasting hyperglycemia was observed, but ␤-cell mass was not diminished. In pancreatectomized mice, persistent glucose intolerance was noted, but this was not further exacerbated by Ex9-39. Accordingly, ␤-cell mass recovery of Ppx mice was not impaired by Ex9-39. In contrast, GLP-1R؊/؊ CD1 mice showed worse glucose intolerance after Ppx compared with wild-type CD1 Ppx mice, and this correlated with a significant defect in ␤-cell mass regeneration. The recovery of ␤-cell mass differed markedly in the BALB/c and CD1 control mice, indicating a significant role of genetic background in the regulation of ␤-cell mass. These studies point to a role for endogenous GLP-1 in ␤-cell regeneration after Ppx in mice.

The role of B cells in transplant tolerance remains unclear. Although B cell depletion often prolongs graft survival, sometimes it results in more rapid rejection, suggesting that B cells may have regulatory activity. We previously demonstrated that tolerance induction by anti-CD45RB antibody requires recipient B cells. Here we show that anti-CD45RB in combination with anti-TIM-1 antibody has a synergistic effect, inducing tolerance in all recipients in a mouse islet allograft model. This effect depends on the presence of recipient B cells, requires B cell IL-10 activity, and is antigen-specific. These data suggest the existence of a regulatory B cell population that promotes tolerance via an IL-10-dependent pathway.

The recent success of islet transplantation using the Edmonton protocol involved the use of sirolimus, tacrolimus, and daclizumab for immunosuppression. Islets were infused into the portal circulation after transhepatic access. This protocol provided a unique opportunity to measure sirolimus and tacrolimus levels from the portal vein and compare them to systemic venous levels. A total of 11 portal venous samples with a corresponding peripheral venous sample were obtained from patients undergoing a first or second islet infusion and medication levels were obtained on both types of specimens. The portal-to-systemic drug level ratio ranged from 0.95 to 2.71 for sirolimus and 1.0 to 3.12 for tacrolimus. Given the potential toxicity of these agents to islets, the findings in this study may have implications for designing the next generation of immunosuppressive protocols for islet transplantation.