The treatment of diabetes mellitus by transplantation of isolated pancreatic islets is an approach that remains the subject of research by a large number of investigators throughout the world. A crucial requirement for the success of this enterprise is the ability to prepare viable isolated islets in adequate quantity. Over the years numerous descriptions of procedures for islet isolation from the pancreas of experimental animals and of man have been advanced; each claiming to be an improvement on previous methods. Indeed, there certainly have been advances, although few techniques live up to the claims that are made in their support Part of the problem is the generally poor methodology

Insulin secretion was studied using adult rat intact islets, dissociated isolated islet cells, and isolated islet cells that had been allowed to reaggregate. The three preparations were maintained for 2 h in tisue culture medium at 37 C in order to allow for equilibration after the isolation procedures and to permit restoration of contacts between the reaggregated cells. The isolated cells appeared well preserved at the ultrastructural level. Evidence for intimate contacts between the reaggregated cells was demonstrated by the reappearance of gap junctions between adjacent cells. Basal insulin release (2.8 mM glucose) during 30 min was elevated in the isolated cell suspension but was restored to the level found in intact islets when isolated cells from the same population were reaggregated. The elevated basal release did not appear to be due to cell damage since there was a commensurate increase in the rate of insulin biosynthesis relative to intact islets. Although there was a marked stimulation of insulin release from the intact islets at 16.7 mM glucose, the response was smaller in the reaggregated cells and in the isolated cell suspension. All three preparations responded to elevated cAMP levels evoked by glucagon in the presence of 16.7 mM glucose. Similar results were obtained when insulin release was studied in various preparations derived from newborn rat pancreatic endocrine cells. Thus, basal insulin release was elevated in isolated cells, whereas cultures with cell contacts displayed lower basal insulin release. Taken together, the restoration of lower basal insulin release and the parallel appearance of contacts between the reaggregated cells suggest that these two phenomena are interrelated. Thus, cell contacts may be important in regulating basal insulin release. Whether such regulation is a consequence merely of cell association or of direct communication between cells remains to be established.

Clinical islet cell transplantation has resulted in insulin independence in a limited number of cases. Rejection, recurrence of autoimmunity, and impairment of normal islet function by conventional immunosuppressive drugs, e.g., steroids, tacrolimus, and cyclosporin A, may all contribute to islet allograft loss. Furthermore, intraportal infusion of allogeneic islets results in the activation of intrahepatic macrophages and endothelial cells, followed by production of proinflammatory mediators that can contribute to islet primary nonfunction. We reasoned that the beneficial effects of anti-CD154 treatment on autoimmunity, alloreactivity, and proinflammatory events mediated by macrophages and endothelial cells made it an ideal agent for the prevention of islet allograft failure. In this study, a nonhuman primate model (Papio hamadryas) was used to assess the effect of humanized anti-CD154 (hu5c8) on allogeneic islet engraftment and function. Nonimmunosuppressed and tacrolimus-treated recipients were insulin independent posttransplant, but rejected their islet allografts in 8 days. Engraftment and insulin independence were achieved in seven of seven baboon recipients of anti-CD154 induction therapy administered on days -1, 3, and 10 relative to the islet transplant. Three of three baboons treated with 20 mg/kg anti-CD154 induction therapy experienced delayed rejection episodes, first detected by elevations in postprandial blood glucose levels, on postoperative day (POD) 31 for one and on POD 58 for the other two. Re-treatment with three doses of anti-CD154 resulted in reversal of rejection in all three animals and in a return to normoglycemia and insulin independence in two of three baboons. It was possible to reverse multiple episodes of rejection with this approach. A loss of functional islet mass, as detected by reduced first-phase insulin release in response to intravenous glucose tolerance testing, was observed after each episode of rejection. One of two baboons treated with 10 mg/kg induction therapy became insulin independent post-transplant but rejected the islet graft on POD 10; the other animal experienced a reversible rejection episode on POD 58 and remained insulin independent and normoglycemic until POD 264. Two additional baboon recipients of allogeneic islets and donor bone marrow (infused on PODs 5 and 11) were treated with induction therapy (PODs -1, 3, 10), followed by initiation of monthly maintenance therapy (for a period of 6 months) on POD 28. Rejection-free graft survival and insulin independence was maintained for 114 and 238 days, with preservation of functional islet mass observed in the absence of rejection. Prevention and reversal of rejection, in the absence of the deleterious effects associated with the use of conventional immunosuppressive drugs, make anti-CD154 a unique agent for further study in islet cell transplantation.

Schizodeme analysis of Leishmania isolates and comparison with IOIJIe phenotypic techniques. In: Rioux lA, m. LeishmDnia. Taxonomie et phylogcn=. Applications ea>-qlidemio1ogiques. Monrpellicr. CNRSIINSERNIOMS. 1986: 57-65.

This report provides our initial experience in islet isolation and intrahepatic allotransplantation in 21 patients. In group 1, 10 patients underwent combined liver-islet allotransplantation following upper-abdominal exenteration for cancer. In group 2, 4 patients received a combined liver-islet allograft for cirrhosis and diabetes. One patients had plasma C-peptide greater than 3 pM and was therefore excluded from analysis. In group 3, 7 patients received 8 combined cadaveric kidney-islet grafts (one retransplant) for end-stage renal disease secondary to type 1 diabetes mellitus. The islets were separated by a modification of the automated method for human islet isolation and the preparation were infused into the portal vein. Immunosuppression was with FK506 (group 1) plus steroids (groups 2 and 3). Six patients in group 1 did not require insulin treatment for 5 to greater than 16 months. In groups 2 and 3 none of the patients became insulin-independent, although decreased insulin requirement and stabilization of diabetes were observed. Our results indicate that rejection is still a major factor limiting the clinical application of islet transplantation in patients with type 1 diabetes mellitus, although other factors such as steroid treatment may contribute to deteriorate islet engraftment and/or function.