Different effects of FK506, rapamycin, and mycophenolate mofetil on glucose-stimulated insulin release and apoptosis in human islets.

Pancreatic islet transplantation has the potential to be an effective treatment for type 1 diabetes mellitus. While recent improvements have improved 1-year outcomes, follow-up studies show a persistent loss of graft function/survival over 5 years. One possible cause of islet transplant failure is the immunosuppressant regimen required to prevent alloimmune graft rejection. Although there is evidence from separate studies, mostly in rodents and cell lines, that FK506 (tacrolimus), rapamycin (sirolimus), and mycophenolate mofetil (MMF; CellCept) can damage pancreatic beta-cells, there have been few side-by-side, multiparameter comparisons of the effects of these drugs on human islets. In the present study, we show that 24-h exposure to FK506 or MMF impairs glucose-stimulated insulin secretion in human islets. FK506 had acute and direct effects on insulin exocytosis, whereas MMF did not. FK506, but not MMF, impaired human islet graft function in diabetic NOD*scid mice. All of the immunosuppressants tested in vitro increased caspase-3 cleavage and caspase-3 activity, whereas MMF induced ER-stress to the greatest degree. Treating human islets with the GLP-1 agonist exenatide ameliorated the immunosuppressant-induced defects in glucose-stimulated insulin release. Together, our results demonstrate that immunosuppressants impair human beta-cell function and survival, and that these defects can be circumvented to a certain extent with exenatide treatment.

A multi-year analysis of islet transplantation compared with intensive medical therapy on progression of complications in type 1 diabetes.

BACKGROUND: We hypothesized that transplantation of islets into type 1 diabetics could improve outcomes of glucose metabolism, renal function, retinopathy, and neuropathy compared with intensive medical therapy. METHODS: We conducted a prospective, crossover, cohort study of intensive medical therapy (group 1) versus islet cell transplantation (group 2) in 42 patients. All were enrolled in group 1 then 31 crossed over with group 2 when islet donation became available. Transplantation was performed by portal venous embolization of more than 12,000 islet equivalents/kg body weight under cover of immunosuppression with antithymocyte globulin, tacrolimus, and mycophenolate. Outcome measures were HbA1c, change in glomerular filtration rate (GFR), progression of retinopathy, and change in nerve conduction velocity. This report details interim analysis of outcomes after 34+/-18 months (group 1) and 38+/-18 months (group 2). RESULTS: HbA1c (%) in group 1 was 7.5+/-0.9 versus 6.6+/-0.7 in group 2 (P<0.01). GFR (mL/min/month) declined in both groups (group 1 -0.45+/-0.7 vs. group 2 -0.12+/-0.7, P=0.1). Slope of the GFR decline in group 1 was significantly more than 0. Retinopathy progressed in 10 of 82 eyes in group 1 versus 0 of 51 in group 2 (P<0.01). Nerve conduction velocity (m/sec) remained stable in group 1 (47.8+/-5 to 47.1+/-5 m/sec) and group 2 (47.2+/-4.5 to 47.7+/-3.5). CONCLUSION: Islet transplantation yields improved HbA1c and less progression of retinopathy compared with intensive medical therapy during 3 years follow-up.

Reduced progression of diabetic retinopathy after islet cell transplantation compared with intensive medical therapy.

BACKGROUND: Diabetic retinopathy is a major complication of type 1 diabetes and remains a leading cause of visual loss. There have been no comparisons of the effectiveness of intensive medical therapy and islet cell transplantation on preventing progression of diabetic retinopathy. METHODS: The British Columbia islet transplant program is conducting a prospective, crossover study comparing medical therapy and islet cell transplantation on the progression of diabetic retinopathy. Progression was defined as the need for laser treatment or a one step worsening along the international disease severity scale. An interim data analysis was performed after a mean 36-month follow-up postislet transplantation and these results are presented. RESULTS: The medical and postislet transplant groups were similar at baseline. Subjects after islet transplantation had better glucose control than the medically treated subjects (mean HbA1c 6.7%+/-0.9% vs. 7.5+/-1.2, P<0.01) and were C-peptide positive. Progression occurred significantly more often in all subjects in the medical group (10/82 eyes, 12.2%) than after islet transplantation (0/51 eyes, 0%) (P<0.01). Considering only subjects who have received transplants, progression occurred in 6/51 eyes while on medical treatment and 0/51 posttransplant (P<0.02). CONCLUSIONS: Progression of diabetic retinopathy was more likely to occur during medical therapy than after islet cell transplantation.

Transplantation for the treatment of type 1 diabetes.

Transplantation of pancreatic tissue, as either the intact whole pancreas or isolated pancreatic islets has become a clinical option to be considered in the treatment of patients with type 1 insulin-dependant diabetes mellitus. A successful whole pancreas or islet transplant offers the advantages of attaining normal or near normal blood glucose control and normal hemoglobin A1c levels without the risks of severe hypoglycemia associate with intensive insulin therapy. Both forms of transplants are also effective at eliminating the occurrence of significant hypoglycemic events (even with only partial islet function evident). Whereas whole pancreas transplantation has also been shown to be very effective at maintaining a euglycemic state over a sustained period of time, thus providing an opportunity for a recipient to benefit from improvement of their blood glucose control, it is associated with a significant risk of surgical and post-operative complications. Islet transplantation is attractive as a less invasive alternative to whole pancreas transplant and offers the future promise of immunosuppression-free transplantation through pre-transplant culture. Islet transplantation however, may not always achieve the sustained level of tight glucose control necessary for reducing the risk of secondary diabetic complications and exposes the patient to the adverse effects of immunosuppression. Although recent advances have led to an increased rate of obtaining insulin-independence following islet transplantation, further developments are needed to improve the long-term viability and function of the graft to maintain improved glucose control over time.

Improved human pancreatic islet isolation for a prospective cohort study of islet transplantation vs best medical therapy in type 1 diabetes mellitus.

HYPOTHESIS: A local multiorgan donor pancreas procurement program can provide a source for optimized isolation of purified viable islets for transplantation into patients with type 1 diabetes mellitus receiving best medical therapy. DESIGN: Prospective before-after cohort study. SETTING: Tertiary referral center. PATIENTS: Glycemic control was assessed in 10 patients with diabetes-induced renal dysfunction who were enrolled in a best medical therapy program and then crossed over to islet transplantation. INTERVENTIONS: Thirty human pancreata were retrieved from local multiorgan donors and consecutively processed with intraductal collagenase perfusion, continuous digestion, and density gradient purification (group 1, n = 9) or similarly processed but impure tissue fractions cultured in vitro and then repurified to retrieve additional islets (group 2, n = 21). Islets were implanted by percutaneous portal embolization, providing more than 10 000 islet equivalents (IE) per kilogram of body weight (infusions from 1-3 donors per patient) under cover of antithymocyte globulin, sirolimus, or mycophenolate mofetil and tacrolimus. MAIN OUTCOME MEASURES: Islet yields, purity, and cell viability (caspase 3, terminal deoxynucleotidyl transferase-mediated biotin-deoxyuridine 5-triphosphate nick-end labeling stain, and insulin secretion in vitro) were compared. In patients, monitored metabolic parameters were C-peptide secretion, insulin requirements, glycemic excursion, and hemoglobin A(1c) (HbA(1c)). RESULTS: For group 1 vs group 2, no differences were observed in pancreas age (43 vs 44 years), cold storage (5 vs 4 hours), or weight (73 vs 82 g). Group 2 yielded 453 690 IE vs 214 109 IE in group 1 (P = .002). Grafts contained 50% or more endocrine cells in both groups. No difference occurred in cell viability or insulin secretion. Islets from 90% of group 2 pancreata met release criteria for transplantation. C-peptide secretion was detected in all recipients and persisted with a median follow-up to 12 months (range, 6-21 months) after full islet transplantation. Daily insulin dependence was reversed in all patients for at least 3 months. Five patients resumed small insulin doses. Compared with the best care program, all patients had improved metabolic stability. The mean +/- SE HbA(1c) level at entry into the study was 7.8% +/- 0.5%, and this decreased to 6.9% +/- 0.2% after best care (P = .38) and further to 6.2% +/- 0.2% at 6 months after transplantation (P = .002 vs entry; P = .15 vs best care; analysis of variance). CONCLUSIONS: Local pancreas donor retrieval with islet isolation and culture conditioning enabled an offer of islets for transplantation for 90% of consecutively processed pancreata. Isolated islets secreted insulin during prolonged follow-up after implantation into patients, yielding metabolic control comparable with that achieved by best medical therapy.

Helicobacter pylori infection targets adherens junction regulatory proteins and results in increased rates of migration in human gastric epithelial cells.

The human gastric pathogen Helicobacter pylori attaches to antral epithelial cells in vivo. Cultured human antral epithelial cells, AGS and NCI-N87 cell lines, were grown in the absence or presence of H. pylori and compared with respect to gene transcript levels, protein expression, organization of the actin cytoskeleton, and the regulation of cell migration. The Clontech Neurobiology array detected differentially expressed transcripts, while Western blots were used to investigate related changes in protein levels. Infection with H. pylori consistently upregulated annexin II, S100 A7, Rho-GTP, and IQGAP-1, whereas SSTR-1 was downregulated upon H. pylori infection. In the adherens junction, E-cadherin and IQGAP-1 were translocated from the plasma membrane to intracellular vesicles. The primary and NCI-N87 cells were similar with respect to cell-cell and cell-matrix adhesion and cell migratory behavior; in contrast the AGS cells were significantly different from the primary gastric epithelial cell preparations, and thus caution must be used when using this cell line for studies of gastric disease. These studies demonstrate a correlation between H. pylori infection and alterations to epithelial cell adhesion molecules, including increased levels of Rho-GTP and cell migration. These data indicate that destabilizing epithelial cell adherence is one of the factors increasing the risk of H. pylori-infected individuals developing gastric cancer.