Stimulation of insulin secretion by the incretin hormones glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP) has been found to be diminished in type 2 diabetes. We hypothesized that this impairment is due to a defect at the receptor level induced by the diabetic state, particularly hyperglycemia. Gene expression of incretin receptors, GLP-1R and GIPR, were significantly decreased in islets of 90% pancreatectomized (Px) hyperglycemic rats, with recovery when glucose levels were normalized by phlorizin. Perifused islets isolated from hyperglycemic Px rats showed reduced insulin responses to GLP-1 and GIP. To examine the acute effect of hyperglycemia on incretin receptor expression, a hyperglycemic clamp study was performed for 96 h with reduction of GLP-1 receptor expression but increase in GIP receptor expression. Similar findings were found when islets were cultured at high glucose concentrations for 48 h. The reduction of GLP-1 receptor expression by high glucose was prevented by dominant-negative protein kinase C (PKC)␣ overexpression, whereas GLP-1 receptor expression was reduced with wild-type PKC␣ overexpression. Taken together, GLP-1 and GIP receptor expression is decreased with chronic hyperglycemia, and this decrease likely contributes to the impaired incretin effects found in diabetes. Diabetes

We describe a new technique for microencapsulation with high-mannuronic acid (high-M) alginate crosslinked with BaCl 2 without a traditional permselective component, which allows the production of biocompatible capsules that allow prolonged survival of syngeneic and allogeneic transplanted islets in diabetic BALB/c and NOD mice for >350 days. The normalization of the glycemia in the transplanted mice was associated with normal glucose profiles in response to intravenous glucose tolerance tests. After explantation of the capsules, all mice became hyperglycemic, demonstrating the efficacy of the encapsulated islets. The retrieved capsules were free of cellular overgrowth and islets responded to glucose stimulation with a 5-to 10-fold increase of insulin secretion. Transfer of splenocytes isolated from transplanted NOD mice to NOD/SCID mice adoptively transferred diabetes, indicating that NOD recipients maintained islet-specific autoimmunity. In conclusion, we have developed a simple technique for microencapsulation that prolongs islet survival without immunosuppression, providing complete protection against allorejection and the recurrence of autoimmune diabetes. Diabetes 50: 1698 -1705, 2001 I slet transplantation represents an important alternative for the treatment of type 1 diabetes but still requires immunosuppressive agents with their serious side effects (1). One approach to avoid such treatment is to protect islets from the host's immune system with a semipermeable, biocompatible membrane (2,3). Transplantation of islets contained in alginate-poly-L-lysine (PLL) capsules was first described by Lim and Sun (4). Numerous studies have shown successful reversal of diabetes by transplantation of islets enclosed in alginate-PLL capsules in streptozotocin-induced animals (5-9). However, limited success has been reported in spontaneously diabetic NOD mice, a model of autoimmune diabetes (10 -13). Various factors have been implicated in the failure of encapsulated islets. A cellular reaction surrounding the capsules has often been observed, which could lead to depletion of oxygen and nutrients (14) or production of toxic cytokines (15). This accumulation of cells could be due to an immune response to the contained islets or to bioincompatibility of the capsular materials (16,17). Failure might also be attributed to problems with ␤-cell viability in the capsules (18).Protection of porcine islets remains a goal of encapsulation, but xenografts might be more difficult to protect than allografts, as suggested by studies performed with permeable polymer membranes (19,20). This concept is important because of the improved prospects for obtaining an abundant supply of human ␤-cells from precursor cells (21,22). The goal of this study was to determine whether stable biocompatible alginate microcapsules without a permselective component, such as PLL or polyethylene-glycol, would be able to protect mouse islets against allorejection and autoimmunity. RESEARCH DESIGN AND METHODSIslet isolation. Islets were isolated from ma...

Differentiation and maturation of porcine neonatal pancreatic cell clusters (NPCCs) microencapsulated in barium alginate were assessed after transplantation into immunocompetent mice. Microencapsulated NPCCs were transplanted into the peritoneal cavity of streptozocin-induced diabetic B6AF1 mice (n ‫؍‬ 32). The microcapsules were removed at 2, 6, and 20 weeks and examined for cellular overgrowth, insulin content, and insulin secretory responses to glucose and glucose with theophylline. The differentiation, maturation, and proliferation of the ␤-cells in the NPCCs were assessed by immunohistochemistry. Blood glucose levels were normalized in 81% of the animals that received a transplant and remained normal until termination of the experiments at 20 weeks. Hyperglycemic blood glucose levels after explantation of the capsules confirmed the function of the encapsulated NPCCs. Insulin content of the encapsulated NPCCs was increased 10-fold at 20 weeks after transplantation compared with pretransplantation levels. A 3.2-fold increase of the ratio of the ␤-cell area to the total cellular area was observed at 20 weeks, demonstrating the maturation of NPCCs into ␤-cells. A s a result of recent progress (1), there is increased interest in islet transplantation as a potential therapy for type 1 diabetes. However, two major barriers must be overcome before islet transplantation can be provided for more patients: 1) the limited availability of human pancreatic tissue and 2) the need for permanent immunosuppression to prevent graft rejection and autoimmunity (2). Xenogeneic islets from pigs and cows (3-5) have been considered as potential sources of islets for transplantation. Many factors favor the use of pigs: the similar structure of porcine and human insulin, the comparable glucose levels, and that both pigs and humans are omnivores. Islet cells can be isolated in large numbers from adult (6 -9) or neonatal pigs (10,11). However, adult pig islets have proved to be difficult to isolate and tend to fare poorly in tissue culture, which has limited their use. Neonatal pancreatic cell clusters (NPCCs) contain a high proportion of islet precursor cells, can be maintained in culture, and differentiate into ␤-cells after transplantation (11). Naked (10,12) or microencapsulated NPCCs (13) have been shown to restore normoglycemia after transplantation into streptozotocin (STZ)-diabetic nude mice.The concept of a bioartificial pancreas, consisting of islets enclosed within immunobarrier membranes, provides a potential way to overcome the need for immunosuppression. Our group recently developed a promising encapsulation method that uses highly purified alginate cross-linked with BaCl 2 , without a separate permselective barrier, which protects islets against allorejection and autoimmunity (14). The aims of this study were to assess the protective capacity of simple barium-alginate capsules in a xenotransplantation model of NPCCs transplanted into STZ-induced diabetic immunocompetent mice and then to evaluate the growth, maturation, a...