Melittin disruption of raft and non-raft-forming biomimetic membranes: a study by quartz crystal microbalance with dissipation monitoring.

In this work we examine the role of lateral phase separation in cholesterol-containing biomimetic membranes on the disrupting action of melittin using a label-free surface-sensitive technique, quartz crystal microbalance with dissipation monitoring (QCM-D). Melittin disruption mechanisms depend strongly on the geometry of the lipid layer; however, despite the interplay between layer geometry/thickness and melittin activity, results indicate that the presence of lipid heterogeneity and lateral phase separation greatly influences the disrupting efficiency of melittin. In homogeneous non-raft forming membranes with high cholesterol content, melittin spontaneous activity is strongly delayed compared to heterogeneous raft-forming systems with the same amount of cholesterol. These results confirm the importance of lateral phase separation as a determinant factor in peptide activity. The information provided can be used for the design of more efficient antimicrobial peptides and the possibility of using a label-free approach for tailored-membranes and interactions with other types of peptides, such as amyloid peptides.

Combined islet and hematopoietic stem cell allotransplantation: a clinical pilot trial to induce chimerism and graft tolerance.

To prevent graft rejection and avoid immunosuppression-related side-effects, we attempted to induce recipient chimerism and graft tolerance in islet transplantation by donor CD34+hematopoietic stem cell (HSC) infusion. Six patients with brittle type 1 Diabetes Mellitus received a single-donor allogeneic islet transplant (8611 +/- 2113 IEQ/kg) followed by high doses of donor HSC (4.3 +/- 1.9 x 10(6) HSC/kg), at days 5 and 11 posttransplant, without ablative conditioning. An 'Edmonton-like' immunosuppression was administered, with a single dose of anti-TNFalpha antibody (Infliximab) added to induction. Immunosuppression was weaned per protocol starting 12 months posttransplant. After transplantation, glucose control significantly improved, with 3 recipients achieving insulin-independence for a short time (24 +/- 23 days). No severe hypoglycemia or protocol-related adverse events occurred. Graft function was maximal at 3 months then declined. Two recipients rejected within 6 months due to low immunosuppressive trough levels, whereas 4 completed 1-year follow-up with functioning grafts. Graft failure occurred within 4 months from weaning (478 +/- 25 days posttransplant). Peripheral chimerism, as donor leukocytes, was maximal at 1-month (5.92 +/- 0.48%), highly reduced at 1-year (0.20 +/- 0.08%), and was undetectable at graft failure. CD25+T-lymphocytes significantly decreased at 3 months, but partially recovered thereafter. Combined islet and HSC allotransplantation using an 'Edmonton-like' immunosuppression, without ablative conditioning, did not lead to stable chimerism and graft tolerance.

Simple measures to monitor beta-cell mass and assess islet graft dysfunction.

The aim of this study was to develop a simple test for the assessment of islet graft dysfunction based on measures involving fasting C-peptide. Calculations were made to account for the dependence of C-peptide secretion on glucose concentration (C-peptide/glucose ratio [CP/G]) and adjusted for renal function by calculating the C-peptide/glucose-creatinine ratio (CP/GCr). Values from 22 recipients were analyzed at different times post-last islet infusion. Receiver operating characteristic curves were used to determine which of these measures best predicts high 90-minute glucose (90 min-Glc; >10 mmol/L) after a Mixed Meal Tolerance Test (MMTT). In this initial analysis, CP/G was found to be superior predicting high 90 min-Glc with a larger area under the ROC curve than C-peptide (p = 0.01) and CP/GCr (p = 0.06). We then correlated C-peptide and CP/G with islet equivalents--IEQ/kg infused, 90 min-Glc after MMTT and clinical outcome (beta-score). C-peptide and CP/G in the first 3 months post-last islet infusion correlated with IEQ/kg infused. CP/G correlated with 90 min-Glc and beta-score. C-peptide and CP/G are good indicators of islet mass transplanted. CP/G is more indicative of graft dysfunction and clinical outcome than C-peptide alone. The ease of calculation and the good correlation with other tests makes this ratio a practical tool when monitoring and managing islet transplant recipients.

Continuous glucose monitoring system for early detection of graft dysfunction in allogenic islet transplant recipients.

BACKGROUND: There are no effective indicators of graft dysfunction in islet transplantation. This study evaluated the role of the Continuous Glucose Monitoring System (CGMS) as an early indicator of graft dysfunction in islet transplant recipients. METHODS: In 5 islet allograft recipients, we retrospectively determined the date of graft dysfunction: 3 fasting blood glucose levels >7.8 mmol/L (140 mg/dL) and/or 3 postprandial blood glucose levels >10 mmol/L (180 mg/dL) in 1 week. We then determined 2 time points in respect to graft dysfunction, 5 to 9 months before (time point A) and 2 to 3 months before (time point B). For these 2 time points, we assessed the following: HbA1c, C-peptide (CP), C-peptide glucose ratio (CPGR), 90-minute glucose from mixed meal tolerance test, and percentage of capillary blood glucose levels >7.8 mmol/L (%CBG >7.8) in a 15-day interval (1 week before and after CGMS placement). From the CGMS recordings, we calculated the glucose variability and the percentage of time spent in hyperglycemia >7.8 mmol/L (%HGT >7.8) and >10 mmol/L (%HGT >10). RESULTS: No difference was found between time points A and B for the following parameters: HbA1c, CP, CPGR, 90-minute glucose, %CBG >7.8, and %HGT >10. We observed a statistically significant increase from time point A to time point B in glucose variability (1.1 +/- 0.5 mmol/L to 1.6 +/- 0.6 mmol/L; P = .004), and in the %HGT >7.8 (11 +/- 12% to 22 +/- 18%; P = .036). CONCLUSION: Glucose variability and %HGT >7.8 determined using CGMS are useful as early indicators of graft dysfunction in islet transplant recipients. Further studies with larger sample sizes will help validate these observations.

C-peptide and glucose values in the peritransplant period after intraportal islet infusions in type 1 diabetes.

BACKGROUND: Successful islet allograft transplantation has been achieved worldwide. This study aimed at evaluating the relationship between peritransplant C-peptide (CP) values and long-term allograft function. METHODS: We measured CP-to-glucose ratio (CPGR) in intraportal samples pre- and postinfusion, and in peripheral circulation at baseline pretransplant and at 1, 3, 6, 12, 72 hours, 1 week, and 15 and 30 days after first and second infusion in 13 islet allograft recipients. Peritransplant treatment included intravenous (IV) 5% dextrose in saline in all patients. We compared portal CPGR to insulin reduction (%) at 30 days after each infusion, and at 1 year after second infusion. RESULTS: CPGR peaked between the immediate postinfusion and 3 hours and decreased at 12 hours. At 1 week, CPGR was 0.76 +/- 0.45 and 1.44 +/- 0.37 after first and second infusion, respectively. CPGR at 30 days after second infusion doubled compared to first infusion (P < .001). There was no correlation between peak CPGR and insulin reduction percent at any time point. One patient experienced hypoglycemia (47 mg/dL) 1 hour after second infusion. CONCLUSIONS: There was no relationship between the CP values in the peritransplant period and long-term graft function or success rate. The early peak in the C-peptide levels is indicative of a significant insulin release after each islet infusion. For this reason, it is important to carefully monitor serum glucose levels in the peritransplant period (hourly for the first 6 hours) and to maintain an IV glucose infusion to avoid hypoglycemia.