Islet graft response to transplantation injury includes up-regulation of protective as well as apoptotic genes

Pancreatic islets are particularly vulnerable in the initial days after transplantation when multiple factors converge to damage the islet graft. The aim of this study was to investigate the expression profile of genes involved in damage and protection of beta-cells in the initial days after syngeneic islet transplantation. We studied the expression of a set of selected genes involved in apoptosis (Bcl2, Bclx(L), Bax, Bad, Bid, and CHOP), cytokine defense, (SOCS-1 and SOCS-3), or free radical protection (Hmox1, Cu/Zn-SOD, Mn-SOD, and Hsp70). Because hyperglycemia has deleterious effects on islet transplantation outcome, we studied its effect on the expression of these genes. Five hundred islets were syngeneically transplanted under the kidney capsule of normoglycemic or streptozotocin-induced diabetic Lewis rats. Gene expression was analyzed by quantitative real-time RT-PCR in grafts 1, 3, and 7 days after transplantation, and in freshly isolated islets. The expression of proapoptotic genes Bid and CHOP, as well as protective genes Bclx(L), Socs1, Socs3, Hmox1, and MnSod, was maximally increased 1 day after transplantation, and in most cases it remained increased 7 days later, indicating the presence of a protective response against cell damage. In contrast, the expression of Bcl2, Bax, Bad, Cu/ZnSod, and Hsp70 genes did not change. Hyperglycemia did not modify the expression of most studied genes. However, MnSod and Ins2 expression was increased and reduced, respectively, on day 7 after transplantation to diabetic recipients, suggesting that hyperglycemia increased oxidative stress and deteriorated beta-cell function in transplanted islets

A role for the host in the roadmap to diabetes stem cell therapy

Stem cells represent an unlimited source for cell therapy (1), and considerable efforts have been made to overcome barriers to introducing this revolutionary therapy into clinical practice. Briefly, the following actions must be taken: 1) design in vitro differentiation strategies to generate either mature postmitotic b-cells or b-cell progenitors that may be safely implanted into the host (e.g., without uncontrolled proliferation), 2) devise selection methods to produce a pure b-cell population, 3) validate standard characterization protocols to determine the real differentiation stage of the cells ready to be transplanted, 4) obtain encapsulation devices to implant the cells, 5) develop preclinical controls in representative animal models, and 6) de fine cell-host interactions (for a recent review see ref. 2

A model for human islet transplantation to immunodeficient streptozotocin-induced diabetic mice

Streptozotocin (STZ) is a cytotoxic glucose analogue that causes beta cell death and is widely used to induce experimental diabetes in rodents. The sensitivity of beta cells to STZ is species-specific and human beta cells are resistant to STZ. In experimental islet transplantation to rodents, STZ-diabetes must be induced before transplantation to avoid destruction of grafted islets by STZ. In human islet transplantation, injection of STZ before transplantation is inconvenient and costly, since human islet availability depends on organ donation and frail STZ-diabetic mice must be kept for unpredictable lapses of time until a human islet preparation is available. Based on the high resistance of human beta cells to STZ, we have tested a new model for STZ-diabetes induction in which STZ is injected after human islet transplantation. Human and mouse islets were transplanted under the kidney capsule of athymic nude mice, and 10-14 days after transplantation mice were intraperitoneally injected with five consecutive daily doses of STZ or vehicle. Beta-cell death increased and beta-cell mass was reduced in mouse islet grafts after STZ injection. In contrast, in human islet grafts beta cell death and mass did not change after STZ injection. Mice transplanted with rodent islets developed hyperglycemia after STZ-injection. Mice transplanted with human islets remained normoglycemic and developed hyperglycemia when the graft was harvested. STZ had no detectable toxic effects on beta cell death, mass and function of human transplanted islets. We provide a new, more convenient and cost-saving model for human islet transplantation to STZ-diabetic recipients in which STZ is injected after islet transplantation

Role of blood glucose in cytokine gene expression in early syngeneic islet transplantation

In islet transplantation, local production of cytokines at the grafted site may contribute to the initial nonspecific inflammation response. We have determined whether the metabolic condition of the recipient modulates the cytokine expression in islet grafts in the initial days after transplantation. Normoglycemic and hyperglycemic streptozotocin-diabetic Lewis rats were transplanted with 500 syngeneic islets, an insufficient beta cell mass to restore normoglycemia in hyperglycemic recipients. The expression of IL-1beta, TNF-alpha, IFN-gamma, IL-6, IL-10, and IL-4 genes was determined by real-time PCR in freshly isolated islets, in 24-h cultured islets and in islet grafts on days 1, 3, and 7 after transplantation. IL-1beta mRNA was strongly and similarly increased in normoglycemic and hyperglycemic groups on days 1, 3, and 7 after transplantation compared with freshly isolated and cultured islets. TNF-alpha mRNA was also strongly increased on day 1, and it remained increased on days 3 and 7. IL-6 and IL-10 were not detected in freshly isolated islets, but their expression was clearly enhanced in 24-h cultured islets and islet grafts. IL-6 was further increased in hyperglycemic grafts. IL-10 expression was increased in both normoglycemic and hyperglycemic grafts on day 1 after transplantation, and remained increased in hyperglycemic grafts compared to 24-h cultured islets. IFN-gamma mRNA was barely detected in a few grafts, and IL-4 mRNA was never detected. Thus, the inflammatory response in islet grafts was maximal on day 1 after transplantation, it was sustained, although at lower levels, on days 3 and 7, and it was partly enhanced by hyperglycemia

Gastrin treatment stimulates beta cell regeneration and improves glucose tolerance in 95% pancreatectomized rats

β-Cell mass reduction is a central aspect in the development of type 1 and type 2 diabetes, and substitution or regeneration of the lost β-cells is a potentially curative treatment of diabetes. To study the effects of gastrin on β-cell mass in rats with 95% pancreatectomy (95%-Px), a model of pancreatic regeneration, rats underwent 95% Px or sham Px and were treated with [15 leu] gastrin-17 (Px+G and S+G) or vehicle (Px+V and S+V) for 15 d. In 95% Px rats, gastrin treatment reduced hyperglycemia (280 ± 52 mg vs. 436 ± 51 mg/dl, P < 0.05), and increased β-cell mass (1.15 ± 0.15 mg)) compared with vehicle-treated rats (0.67 ± 0.15 mg, P < 0.05). Gastrin treatment induced β-cell regeneration by enhancing β-cell neogenesis (increased number of extraislet β-cells in Px+G: 0.42 ± 0.05 cells/mm(2) vs. Px+V: 0.27 ± 0.07 cells/mm(2), P < 0.05, and pancreatic and duodenal homeobox 1 expression in ductal cells of Px+G: 1.21 ± 0.38% vs. Px+V: 0.23 ± 0.10%, P < 0.05) and replication (Px+G: 1.65 ± 0.26% vs. S+V: 0.64 ± 0.14%; P < 0.05). In addition, reduced β-cell apoptosis contributed to the increased β-cell mass in gastrin-treated rats (Px+G: 0.07 ± 0.02%, Px+V: 0.23 ± 0.05%; P < 0.05). Gastrin action on β-cell regeneration and survival increased β-cell mass and improved glucose tolerance in 95% Px rats, supporting a potential role of gastrin in the treatment of diabetes

Diminished fraction of blockable ATP-sensitive K+ channels in islets transplanted into diabetic mice

The reasons for the poor outcome of islet transplantation in diabetic patients are not well known; a better understanding of the pathophysiology of transplanted islets is needed. To study the mechanism coupling secretagogue stimuli with insulin release in transplanted islets, we determined the effects of glucose, tolbutamide, and carbamylcholine on the beta-cell membrane potential and cytosolic calcium concentrations ([Ca2+]i) of islets syngeneically transplanted into normal and streptozocin-induced diabetic mice. In both groups, normoglycemia was maintained after transplantation. Islets transplanted into normal recipients showed similar changes in beta-cell membrane potential and [Ca2+]i oscillations to those in control islets. In contrast, when islets were transplanted into diabetic mice, bursts of electrical activity were triggered at lower glucose concentrations (5.6 mmol/l) than in control islets (11 mmol/l), and maximal electrical activity was achieved at lower glucose concentrations (11 mmol/l) than in control islets (22 mmol/l). When membrane potential was plotted as a function of glucose concentration, the dose-response curve was shifted to the left. Compared with control islets, glucose-induced [Ca2+]i oscillations were broader in duration (22.3 +/- 0.6 s vs. 118.1 +/- 12.6 s; P < 0.01) and higher in amplitude (135 +/- 36 nmol/l vs. 352 +/- 36 nmol/l; P < 0.01). Glucose supersensitivity was attributed to a resting decrease in the fraction of blockable ATP-sensitive K+ (K+(ATP)) channels in transplanted islets that maintained normoglycemia with a limited beta-cell mass

The pancreatic β-cell in ageing: Implications in age-related diabetes

The prevalence of type 2 diabetes (T2D) and impaired glucose tolerance (IGT) increases with ageing. T2D generally results from progressive impairment of the pancreatic islets to adapt fi-cell mass and function in the setting of insulin resistance and increased insulin demand. Several studies have shown an age-related decline in peripheral insulin sensitivity. However, a precise understanding of the pancreatic fi-cell response in ageing is still lacking. In this review, we summarize the age-related alterations, adaptations and/or failures of fi-cells at the molecular, morphological and functional levels in mouse and human. Age-associated alterations include processes such as fi-cell proliferation, apoptosis and cell identity that can influence fi-cell mass. Age-related changes also affect fi-cell function at distinct steps including electrical activity, Ca2+ signaling and insulin secretion, among others. We will consider the potential impact of these alterations and those mediated by senescence pathways on fi-cells and their implications in age-related T2D. Finally, given the great diversity of results in the field of fi-cell ageing, we will discuss the sources of this heterogeneity. A better understanding of fi-cell biology during ageing, particularly at older ages, will improve our insight into the contribution of fi-cells to ageassociated T2D and may boost new therapeutic strategies

Limited Joint Mobility Progression In Type 1 Diabetes: A 15-year Follow-up Study

Objective. To assess the evolution of joint mobility over a period of 15 years in type 1 diabetic patients and healthy controls and to determine whether microalbuminuria is associated with a different evolution of joint mobility. Methods. Joint mobility of hand and wrist was determined in 63 patients with type 1 diabetes and 63 healthy subjects. Fifteen years later, 37 (58.7%) diabetic patients and 16 (25.4%) healthy subjects were studied again. Joint mobility was assessed with the Prayer sign and by measuring the angle of maximal flexion of the fifth and third metacarpophalangeal (MCP) joints and wrist. Patients with diabetes were visited 2-4 times every year with regular assessment of glycated hemoglobin (HbA1(c)), urinary albumin excretion (UAE), and ophthalmoscopy. Results. Fifteen years after the initial exam, diabetic patients showed reduced flexion of the fifth MCP joint (82.6 +/- 5.8 versus 76.0 +/- 6.4 degrees, p < 0 001) and wrist (75.9 +/- 8.1 versus 73.2 +/- 7.4 degrees, p = 0 015) compared to baseline examination. Joint mobility did not change significantly in healthy subjects. Patients with microalbuminuria showed greater reduction in hand joint mobility than diabetic patients with normal UAE or than healthy subjects (p < 0 001). Conclusions. In type 1 diabetic patients, the severity of LJM progresses with time, and the progression is enhanced in patients with microalbuminuria

Short term culture with the caspases inhibitor z-VAD fmk reduces beta cell apoptosis in transplanted islets and improves the metabolic outcome of the graft

In the initial days after transplantation islets are particularly vulnerable and show increased apoptosis and necrosis. We have studied the effects of caspase inhibition on this early beta cell death in syngeneically transplanted islets. Streptozotocin-diabetic C57BL/6 mice were transplanted with 150 syngeneic islets, an insufficient mass to restore normoglycemia, preincubated with or without the pan-caspase inhibitor z-VAD. fmk 2 h before transplantation. Beta cell apoptosis was increased in control islets on day 3 after transplantation (0.28 ± 0.02%) compared with freshly isolated islets (0.08 ± 0.02%, p< 0.001), and was partially reduced in transplanted islets preincubated with z-VAD.fmk 200 μM (0.14 ± 0.02%, p = 0.003) or with z-VAD.fmk 500 μM (0.17 ± 0.01%, p = 0.012), but not with a lower z-VAD.fmk (100 μM) concentration. Diabetic mice transplanted with islets preincubated with z-VAD.fmk 500 μM showed an improved metabolic evolution compared with control and z-VAD.fmk 200 μM groups. The z-VAD.fmk 500 μM group showed an overall lower blood glucose after transplantation (p = 0.02), and at the end of the study blood glucose values were reduced compared with transplantation day (15.7 ± 3.6 vs. 32.5 ± 0.5 mmol/L, p = 0.001). In contrast, blood glucose was not significantly changed in control and z-VAD.fmk 200 μM groups. Four weeks after transplantation beta cell mass was higher in z-VAD.fmk 500 μM group (0.15 ± 0.02 mg) than in the control group (0.10 ± 0.02 mg) (p = 0.043). In summary, the treatment of freshly isolated islets with the caspase inhibitor z-VAD.fmk reduced the subsequent apoptosis of the islets once they were transplanted and improved the outcome of the graft

Insuficiencia suprarrenal aguda por metástasis suprarrenales secundarias a melanoma cutáneo

Primary adrenal insufficiency is a vital emergency that requires a substitute treatment without delay. Clinically, it can present with nonspecific symptoms, so that its diagnostic suspicion can sometimes be a challenge. Bilateral adrenal metastases are well described but they uncommonly trigger adrenal insufficiency. We illustrate a case of malignant melanoma that required immediate emergency assistance with hydrocortisone.La insuficiencia suprarrenal primaria es una urgencia vital que requiere tratamiento sustitutivo sin demora. Clínicamente se puede presentar con síntomas inespecíficos, por lo que su sospecha diagnóstica en algunas ocasiones puede suponer un reto. Las metástasis suprarrenales bilaterales están bien descritas, pero raramente desencadenan insuficiencia suprarrenal. Ilustramos un caso de melanoma maligno que requirió asistencia inmediata en Urgencias con hidrocortisona