National Institutes of Health-Sponsored Clinical Islet Transplantation Consortium Phase 3 Trial: Manufacture of a Complex Cellular Product at Eight Processing Facilities.

Eight manufacturing facilities participating in the National Institutes of Health-sponsored Clinical Islet Transplantation (CIT) Consortium jointly developed and implemented a harmonized process for the manufacture of allogeneic purified human pancreatic islet (PHPI) product evaluated in a phase 3 trial in subjects with type 1 diabetes. Manufacturing was controlled by a common master production batch record, standard operating procedures that included acceptance criteria for deceased donor organ pancreata and critical raw materials, PHPI product specifications, certificate of analysis, and test methods. The process was compliant with Current Good Manufacturing Practices and Current Good Tissue Practices. This report describes the manufacturing process for 75 PHPI clinical lots and summarizes the results, including lot release. The results demonstrate the feasibility of implementing a harmonized process at multiple facilities for the manufacture of a complex cellular product. The quality systems and regulatory and operational strategies developed by the CIT Consortium yielded product lots that met the prespecified characteristics of safety, purity, potency, and identity and were successfully transplanted into 48 subjects. No adverse events attributable to the product and no cases of primary nonfunction were observed.

Assessment of ß-cell mass and α- and ß-cell survival and function by arginine stimulation in human autologous islet recipients.

We used intravenous arginine with measurements of insulin, C-peptide, and glucagon to examine ß-cell and α-cell survival and function in a group of 10 chronic pancreatitis recipients 1-8 years after total pancreatectomy and autoislet transplantation. Insulin and C-peptide responses correlated robustly with the number of islets transplanted (correlation coefficients range 0.81-0.91; P < 0.01-0.001). Since a wide range of islets were transplanted, we normalized the insulin and C-peptide responses to the number of islets transplanted in each recipient for comparison with responses in normal subjects. No significant differences were observed in terms of magnitude and timing of hormone release in the two groups. Three recipients had a portion of the autoislets placed within their peritoneal cavities, which appeared to be functioning normally up to 7 years posttransplant. Glucagon responses to arginine were normally timed and normally suppressed by intravenous glucose infusion. These findings indicate that arginine stimulation testing may be a means of assessing the numbers of native islets available in autologous islet transplant candidates and is a means of following posttransplant α- and ß-cell function and survival.

Are the current Indian growth charts really representative? Analysis of anthropometric assessment of school children in a South Indian district.

BACKGROUND: India currently is posed by the double threat of thinness and overweight/obesity among children. Different growth charts have taken different population and give different cut-off points to assess these conditions. OBJECTIVE: The objective of this study is to assess the anthropometry of school children, 5-18 years of age and thereby estimate the prevalence of childhood thinness, overweight and obesity. To analyze how the study population compares with that of Agarwal's growth chart. MATERIALS AND METHODS: The anthropometric measurements of all the students who were studying from 1(st) to 12(th) standards were taken from 27 randomly selected Government and private schools. Prevalence of thinness, overweight and obesity were assessed using two standards - Indian standard given by Agarwal and International Standards given by International Obesity Task Force (IOTF). RESULTS: The prevalence of thinness, overweight and obesity among 18,001 students enrolled as per Indian standard were 12.2%, 9.5% and 3% and as per International standard were 15.3%, 8.1% and 2.6% respectively. The mean and the 95(th) percentile values of body mass index for both boys and girls at all ages in this study are falling short of Agarwal's and IOTF values. Using international cut-offs as well as Indian cut-offs given by Agarwal, underestimate the prevalence of obesity among boys and girls of all age groups. CONCLUSION: This study shows that under and over-nutrition among school children is in almost equal proportions. There is an underestimation of obesity among children whenever an Indian or an International growth chart is used. Thus, this study brings out the need for a really representative growth chart.

Metabolic assessment prior to total pancreatectomy and islet autotransplant: utility, limitations and potential.

Islet autotransplant (IAT) may ameliorate postsurgical diabetes following total pancreatectomy (TP), but outcomes are dependent upon islet mass, which is unknown prior to pancreatectomy. We evaluated whether preoperative metabolic testing could predict islet isolation outcomes and thus improve assessment of TPIAT candidates. We examined the relationship between measures from frequent sample IV glucose tolerance tests (FSIVGTT) and mixed meal tolerance tests (MMTT) and islet mass in 60 adult patients, with multivariate logistic regression modeling to identify predictors of islet mass ≥2500 IEQ/kg. The acute C-peptide response to glucose (ACRglu) and disposition index from FSIVGTT correlated modestly with the islet equivalents per kilogram body weight (IEQ/kg). Fasting and MMTT glucose levels and HbA1c correlated inversely with IEQ/kg (r values -0.33 to -0.40, p ≤ 0.05). In multivariate logistic regression modeling, normal fasting glucose (<100 mg/dL) and stimulated C-peptide on MMTT ≥4 ng/mL were associated with greater odds of receiving an islet mass ≥2500 IEQ/kg (OR 0.93 for fasting glucose, CI 0.87-1.0; OR 7.9 for C-peptide, CI 1.75-35.6). In conclusion, parameters obtained from FSIVGTT correlate modestly with islet isolation outcomes. Stimulated C-peptide ≥4 ng/mL on MMTT conveyed eight times the odds of receiving ≥2500 IEQ/kg, a threshold associated with reasonable metabolic control postoperatively.

Rapid quantitative assessment of the pig pancreas biopsy predicts islet yield.

BACKGROUND: The cost of islet procurement from donor pigs is increased by the use of organs that produce low yields. We developed an assessment system using dithizone-stained pig pancreas biopsies to enable the preselection of donor organs. METHODS: Pig pancreas biopsy slices were soaked in dithizone solution. The islets were evaluated before islet isolation by converting the islet counts (IC) to islet equivalents (IE), and then determining the IE/cm(2), IE/IC, % islets >150 microm, and % islets >200 microm. These parameters were evaluated in 3 different areas of the pancreas (duodenal, splenic, and connecting lobe; n = 42 each). Stepwise multivariate linear regression analysis was performed to assess for correlations with islet yield and decide which area of the pancreas had the most predictive value. To identify other predictors, including donor and islet isolation variables, we performed binary logistic regression analysis with significant variables from the univariate analysis (n = 67). For this analysis, the pigs were categorized into high (n = 23) and low (n = 44) yield groups. RESULTS: Stepwise multivariate linear regression analysis revealed that IE/cm(2) of the splenic lobe significantly predicted islet yield. Binary logistic regression analysis indicated that the IE/mm(2) of the splenic lobe was the only parameter that significantly correlated with successful pig islet isolations (P = .01; odds ratio 3.605). Variables associated with donor and islet isolation, such as age, gender, ischemic time, or enzyme lot, were not significantly correlated with islet yield. CONCLUSION: Our study suggests that the islet distribution of splenic lobe biopsies can be a reliable predictor of islet yield from pig pancreata.

Establishment of fluorescein diacetate and ethidium bromide (FDAEB) assay for quality assessment of isolated islets.

One of the most important factors in clinical islet transplantation is isolation of a great number of islets with good viability. According to viability assessments of isolated islets, the static incubation test and the perifusion test of islets, which are used retrospectively, take much time and need various apparatus. But viability assessments of isolated islets for clinical islet transplantation require a simple, rapid, sensitive, and prospective method. We have developed a microfluorometric viability assay for isolated human, porcine, and dog islets of pancreata using fluorescein diacetate and ethidium bromide (FDAEB). Fluorescein diacetate (FDA) causes live cells to fluoresce green under blue light excitation (490 nm) and ethidium bromide (EB) causes dead cells to fluoresce red. In this study, we investigated the applicability of FDAEB staining to quality assessment of isolated islets for clinical use by correlation with the counting method with insulin secretion of islets. Discrimination of living from dead islets by insulin secretion correlated well with viability as determined by FDAEB staining. The proportion of living islets within isolated canine islets, as measured by microfluorometric counting, was found to correlate highly significantly on low-temperature (24 degrees C) culture (R = 0.831, p < 0.001) and on 37 degrees C culture (R = 0.553, p < 0.05) with the insulin contents of the same islets. Therefore, it is possible to differentiate degrees of viability, and a scoring system is described for this purpose. The FDAEB assay prospectively and easily provides a rapid, accurate, and objective measurement of the proportion of living cells and dead cells in isolated islets for clinical islet transplantation.