Search

In an effort to better understand the phenomenon of lipotoxicity in human beta-cells, we evaluated the effects of 48-h preculture with 1.0 or 2.0 mmol/l free fatty acid (FFA) (2:1 oleate to palmitate) on the function and survival of isolated human islets and investigated some of the possible mechanisms. Compared with control islets, triglyceride content was significantly increased and insulin content and glucose-stimulated insulin release were significantly reduced in islets precultured with increased FFA concentrations. These changes were accompanied by a significant reduction of glucose utilization and oxidation. By cell death detection techniques, it was observed that exposure to FFAs induced a significant increase of the amount of dead cells. Electron microscopy showed the involvement of beta-cells, with morphological appearance compatible with the presence of apoptotic phenomena. FFA-induced islet cell death was blocked by inhibition of upstream caspases and partially prevented by inhibiton of ceramide synthesis or serine protease activity, whereas inhibition of nitric oxide synthesis had no effect. RT-PCR studies revealed no major change of iNOS and Bax mRNA expression and a marked decrease of Bcl-2 mRNA expression in the islets cultured with FFA. Thus, prolonged exposure to FFAs has cytostatic and pro-apoptotic effects on human pancreatic beta-cells. The cytostatic action is likely to be due to the FFA-induced reduction of intraislet glucose metabolism, and the proapoptotic effects are mostly caspase mediated, partially dependent on ceramide pathway, and possibly Bcl-2 regulated. [ABSTRACT FROM AUTHOR]

Type 2 (non-insulin-dependent) diabetes results from decreased insulin action in peripheral target tissues (insulin resistance) and impaired pancreatic beta-cell function. These defects reflect both genetic components and environmental risk factors. Recently, the common Gly(972)-->Arg amino acid polymorphism of insulin receptor substrate 1 (Arg(972) IRS-1) has been associated with human type 2 diabetes. In this study, we report on some functional and morphological properties of isolated human islets carrying the Arg(972) IRS-1 polymorphism. Insulin content was lower in variant than control islets (94 +/- 47 vs. 133 +/- 56 microU/islet; P < 0.05). Stepwise glucose increase (1.7 to 16.7 mmol/l) significantly potentiated insulin secretion from control islets, but not Arg(972) IRS-1 islets, with the latter also showing a relatively lower response to glyburide and a significantly higher response to arginine. Proinsulin release mirrored insulin secretion, and the insulin-to-proinsulin ratio in response to arginine was significantly lower from Arg(972) IRS-1 islets than from control islets. Glucose utilization and oxidation did not differ in variant and wild-type islets at both low and high glucose levels. Electron microscopy showed that Arg(972) IRS-1 beta-cells had a severalfold greater number of immature secretory granules and a lower number of mature granules than control beta-cells. In conclusion, Arg(972) IRS-1 islets have reduced insulin content, impaired insulin secretion, and a lower amount of mature secretory granules. These alterations may account for the increased predisposition to type 2 diabetes in individuals carrying the Gly(972)-->Arg amino acid polymorphism of IRS-1. [ABSTRACT FROM AUTHOR]

Mixed dyslipidemia (MD; elevated triglycerides [TG], apolipoprotein B [ApoB], and non-high-density lipoprotein cholesterol [non-HDL-C], and decreased HDL-C) is often observed in type 2 diabetes (T2D) and may contribute to increased cardiovascular (CV) risk. Non-HDL-C rather than TG has been proposed as a more useful lipid predictor of CV risk and an important therapeutic target in those with MD. This post hoc subgroup analysis of the open label ODYSSEY DM-DYSLIPIDEMIA trial investigated effects of the PCSK9 inhibitor alirocumab (ALI; 75 mg every 2 weeks [Q2W] with possible increase to 150 mg Q2W at Week 12) vs. usual care (UC; ezetimibe [EZE], fenofibrate [FENO], or no additional lipid lowering therapy [LLT]) on non-HDL-C and other lipids in people with T2D and baseline non-HDL-C ≥100 mg/dL, TG ≥200 mg/dL, and HDL-C <40 mg/dL (men) or <50 mg/dL (women) on maximally tolerated statin dose. Compared with UC, ALI significantly reduced non-HDL-C, ApoB, Lp(a) and LDL particle number from baseline to Week 24 (Table 1); TG reductions were greater with FENO vs. ALI but were not significantly different for any comparison. Adverse events frequency was similar in ALI and UC groups (67.2% vs. 70.7%). ALI was generally well tolerated and effective at further reducing atherogenic lipids in this subgroup of people with T2D and high TG and low HDL-C levels already on maximally tolerated statin dose. Disclosure: H.M. Colhoun: Advisory Panel; Self; Eli Lilly and Company, Novartis Pharmaceuticals Corporation, Sanofi-Aventis. Research Support; Self; AstraZeneca, Boehringer Ingelheim International GmbH, Eli Lilly and Company, Novo Nordisk A/S, Pfizer Inc., Regeneron Pharmaceuticals, Roche Pharma, Sanofi-Aventis. Speaker's Bureau; Self; Eli Lilly and Company, Regeneron Pharmaceuticals, Sanofi. Stock/Shareholder; Self; Bayer AG, Roche Pharma. L.A. Leiter: Advisory Panel; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Eli Lilly and Company, Janssen Pharmaceuticals, Inc., Merck & Co., Inc., Novo Nordisk Inc., Sanofi, Servier. Research Support; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Eli Lilly and Company, GlaxoSmithKline plc., Janssen Pharmaceuticals, Inc., Novo Nordisk Inc., Sanofi. Speaker's Bureau; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Eli Lilly and Company, Janssen Pharmaceuticals, Inc., Merck & Co., Inc., Novo Nordisk Inc., Sanofi. D. Müller-Wieland: Advisory Panel; Self; Amgen Inc., AstraZeneca, Bayer AG, Boehringer Ingelheim Pharmaceuticals, Inc., Merck Sharp & Dohme Corp., Novo Nordisk Inc., Sanofi-Aventis Deutschland GmbH. Speaker's Bureau; Self; Amgen Inc., AstraZeneca, Bayer AG, Boehringer Ingelheim Pharmaceuticals, Inc., Merck Sharp & Dohme Corp., Novo Nordisk Inc., Sanofi-Aventis Deutschland GmbH. B. Cariou: Board Member; Self; Novo Nordisk A/S, Regeneron Pharmaceuticals. Consultant; Self; GENFIT, Sanofi-Aventis. Research Support; Self; Amgen Inc., Pfizer Inc. Speaker's Bureau; Self; Abbott, Akcea Therapeutics, Merck Sharp & Dohme Corp. K.K. Ray: Research Support; Self; Amgen Inc., MSD Pharma, Pfizer Inc., Regeneron Pharmaceuticals. Other Relationship; Self; AbbVie Inc., Akcea Therapeutics, Algorithm, Amgen Inc., Amgen Inc., AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Boehringer Ingelheim Pharmaceuticals, Inc., Cerenis, Cipla, Eli Lilly and Company, Esperion, Kowa Pharmaceutical Europe Co. Ltd., Medco, Pfizer Inc., Regeneron Pharmaceuticals, Resverlogix Corp., Sanofi, Sanofi, Takeda Pharmaceutical Company Limited, Takeda Pharmaceutical Company Limited. F. Tinahones: Advisory Panel; Self; Lilly Diabetes, Novo Nordisk A/S, Regeneron Pharmaceuticals, Sanofi-Aventis. Consultant; Self; AstraZeneca. Research Support; Self; AstraZeneca. Speaker's Bureau; Self; Danone Nutricia Research, Lilly Diabetes, Novo Nordisk A/S, Regenerative Medical Solutions, Sanofi-Aventis. C. Domenger: Employee; Self; Sanofi. A. Letierce: Employee; Self; Sanofi Research & Development. M. Israel: Employee; Self; Regeneron Pharmaceuticals. Stock/Shareholder; Self; Regeneron Pharmaceuticals. S. Del Prato: Advisory Panel; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Eli Lilly and Company, GlaxoSmithKline plc., Merck Sharp & Dohme Corp., Novartis Pharmaceuticals Corporation, Novo Nordisk A/S, Sanofi, Servier, Takeda Pharmaceutical Company Limited. Board Member; Self; AstraZeneca. Research Support; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc. Speaker's Bureau; Self; Boehringer Ingelheim Pharmaceuticals, Inc., Takeda Pharmaceutical Company Limited. Funding: Sanofi; Regeneron Pharmaceuticals, Inc. [ABSTRACT FROM AUTHOR]

Pre-existing and post-transplant diabetes in LTP can heavily impact on recipients' morbidity and mortality. However, little and inconsistent information is available on the prevalence and characteristics of PED in LTP. We studied 203 patients (age: 52±9 yrs, M/F: 157/46, BMI: 24.6±3 Kg/m²) who entered the waiting list for orthotopic liver transplantation in our institution. Based on clinical history and fasting plasma glucose (FPG) concentration, 73 patients (36%) had PED (age: 55±8 yrs, M/F: 60/13, BMI: 25±2.8 kg/m² diabetes duration 6±7 yrs), and 130 patients (64%) normal FPG (N, age: 51±9 yrs, M/F: 97/33, BMI: 24.4±3.1 kg/m²). Compared with N, patients with PED were older (p<0.01), with more frequent family history of diabetes (42% vs 28%, p<0.05) and higher BMI (p<0.03); in addition, the prevalence of hepatitis C virus (HCV) infection tended to be higher (52 vs 44%, p=0.32). Out of patients with PED, 37 (51%) were on insulin, 8 (11%) on oral agents and 28 (38%) on diet alone. PED average glycemic control was fair, with FPG and A1c of 157±66 mg/dl and 6.7±1.4%, respectively, which was associated (in patients on oral agents or diet alone) with lower insulin sensitivity (HOMA1-IR: Homeostatic Model Assessment-Insulin Resistance; 16±28.6 vs 4.85±3.37 p<0.001) and reduced beta-cell function (HOMA1-%B: Homeostatic Model Assessment-Beta-Cell Function; 167±112 vs 293±216 p<0.001). Diabetic retinopathy was found in 9 patients (12%), with previous laser treatment in 3 cases. In conclusion, the prevalence of PED in LTP is very high, and is associated with higher age, positive family history and increased BMI; insulin is the most common form of treatment, and non-insulin treated PED is characterized by lower insulin sensitivity and reduced beta-cell function; diabetic complications may be present in PED. [ABSTRACT FROM AUTHOR]

A proper use of immunosuppressive drugs is crucial for successful pancreas or islet transplantation. However, immunosuppressant agents can have deleterious side effects on the beta-cell. Such results have been obtained more frequently with no longer used molecules or with pharmacological, more than therapeutical, drug concentrations. We investigated the effects of prolonged exposure of isolated human islets to 10 ng/ml Tacrolimus (Tac) or 150 ng/ml Cyclosporin A (CsA). Islets were isolated from the pancreas of 12 multiorgan donors (age: 64±15 years; gender: 8M/4F; BMI: 26.6±3.0 kg/m²) by enzymatic digestion and density gradient centrifugation. Morphological, functional and survival studies were then performed after 96 hr incubation with Tac or CsA. Glucose-induced insulin secretion was significantly decreased in Tac (p<0.05), but not in CsA exposed islets, (stimulation index, i.e. the ratio of insulin release at 16.7 vs 3.3 mmol/1 glucose: Control islets, Ctrl, 2.3±0.5; Tac 1.6±0.2; CsA, 2.0±0.4). This was associated with unchanged mRNA expression of insulin and glut2, and increased expression of glucokinase in Tac exposed cells. Furthermore, electron microscopy (EM) showed a significant (p<0.05) decrease of insulin granules in Tac exposed beta-cells (volume density, ml%: 0.63±0.39), but not in the CsA group (1.55±0.01 ), as compared to Ctrl (1.54±0.09). Finally, by EM it was shown that the percentage of apoptotic beta-cells was higher in Tac (11.7±4.7) than Ctrl (2.7±1.2) (the respective value in CsA was 6.3±1.2). Therefore, Tac has more deleterious effects on the function and survival of human beta-cells compared to CsA, when both agents were used at therapeutical concentration. [ABSTRACT FROM AUTHOR]

Background: GPR40 is a membrane-bound receptor paired with medium and long-chain fatty acids (FFA) as endogenous ligands. Its activation potentiates insulin secretion from beta-cells and might contribute to the deleterious effects of prolonged exposure to FFA. Little information is available on the expression of GPR40 and its regulation in human islets (HI). Materials and Methods: HI were prepared by enzymatic digestion and gradient separation from the pancreas of 37 non-diabetic (Ctrl, age 52.6±16.8 yrs, gender 27M/10F, BMI 24.4±3.6 kg/m2) and 13 type 2 diabetic (T2DM, age 66.3±8.0 yrs, gender 6F/7M, BM1 27.4±3.1 kg/m2) multiorgan donors, and functional and molecular studies were then performed. Results: By qualitative and quantitative PCR experiments, mRNA expression was shown in HI. Expression was unaffected in Ctrl islets by 24h exposure to 16.7 mmol/1 glucose; however, preincubation for 24h with 1.0 mmol/1 FFA (palmitate:oleate, 2:1), significantly (p<0.01) decreased GPR40 expression (-88.9±5.4%), which was prevented by co-incubation with therapeutical concentration of metformin. T2DM islets showed a significantly (p<0.01) lower expression of GPR40 (-81.8±6.2%) as compared to Ctrl cells. A significant positive correlation (r: 0.935, p<0.01) was found between glucose-stimulated insulin secretion and GPR40 expression. Conclusion: These results show the expression of GPR40 in human pancreatic islets, which is regulated by FFA but not by glucose; the finding that T2DM islets have a lower GPR40 expression and the correlation of the expression with insulin secretion raises the possibility of an involvement of GPR40 in human diabetes. [ABSTRACT FROM AUTHOR]

Reduction of pancreatic beta-cell functional mass is central to T2DM, and increased apoptosis plays a major role. Other mechanisms may be involved. Autophagy, a physiological process which degrades suffering proteins and cellular organelles, may lead to a distinct form of programmed cell death. We studied beta-cells from the pancreata and/or isolated islets of 18 multiorgan donors (8 non-diabetic, Ctrl: age, 61±12 yrs; BMI, 25.2±2.5 Kg/m²; and 10 with T2DM: age, 67±8 yrs; BMI, 27.5±3.8 Kg/m²). Electron microscopy showed a slight, but significant decrease of beta-cells in T2DM islets (62.0±2.2 vs 71.0±3.3%, p<0.05). Dead cell percentage was 14±2.4% in T2DM and 5±1.4% in Ctrl (p<0.01), with apoptotic cells representing 7±4 and 2.0±1.3% (p<0.05). Seven±3% (T2DM) and 3±1% (Ctrl) (p<0.05) dead beta-cells showed autophagic cell death (membrane dysruption, extreme vacuole overload, no chromatin condensation, no inflammation). Density volume (ml%) of autophagic vacuoles (containing recognizable organelles, 0.6±0.1 vs 0.1±0.05) and autophagosomes (containing lysosome degraded material, 7.7±1.1 vs 4.7±0.3) was higher (p<0.05) in T2DM than Ctrl. When isolated Ctrl islets were incubated for 5 days in alternating high (16.7 mmol/l) glucose, apoptotic beta-cells (from 3.0±2.4 to 14±3.1%), but not beta-cells with autophagic death (from 8.0±3.0 to 6.0±2.5%), increased significantly. Pre-culture with high FFA concentration (48h, 1 mmol/l oleate:palmitate, 2:1) caused a significant increase of both apoptotic (from 5.3±4.0 to 19±8.2%) and autophagie (from 11±3.0 to 26±3%) beta-cell death, and a marked increase of autophagic vacuoles and autophagosomes. Finally, incubation of Ctrl islets with 0.1 mmol/1 chloroquine (a compound well known to induce autophagy) reproduced the alterations observed in T2DM and FFA-exposed Ctrl islets. Therefore, pancreatic beta-cells in human T2DM have signs of altered autophagy, which may contribute to the loss of functional mass; substrate (in particular FFA) overfueling is likely to play a role. [ABSTRACT FROM AUTHOR]

Subclinical inflammation and platelets activation are involved in the patho-physiology of cardiovascular disease (CVD). Recently both sCD40L (released by activated platelets) and hs-CRP (an inflammatory marker) have been reported higher among hypertensive patients, but factors affecting both parameters are poorly understood. We aimed to measure hs-CRP and sCD40L in 89 CVD event-free men with essential hypertension and normal renal function, and to explore CVD risk factors affecting circulating levels of hs-CRP and sCD40L with emphasis to Metabolic Syndrome (MS) components. We studied 89 patients (age 49±11 yrs, range of BMI: 18.4-34.5 kg/m2, creatinine below 1.2 mg/dl and cystatin-C ranging from 0.53 to 1.18 mg/1). All subjects underwent a 24 hour ABPM and an OGTT. Specific immnnoassay were used for sCD40L and hs-CRP. Among these subjects, 18% had IFG, 8% IGT, 5% both conditions. MS (NCEP/ATPIII criteria) was present in 30 out of 89 subjects (34%). Mean levels of hs-CRP were 2.4±2.3 (range 0.4 to 8.5) ng/ml; mean levels of sCD40L 8.8±5:0 (range 0.8 to 18.5) ng/ml. No correlation was observed between hs-CRP and sCD40L (p=0.97) and no differences in both parameters emerged in subjects with and without MS. Neither hs-CRP and sCD40L did correlate with age, BMI, systolic and diastolic 24h ABPM parameters, renal function indexes or urinary albumin excretion. Within lipid parameters, a weak positive regression was observed between total cholesterol and sCD40L (p=0.047). hs-CRP correlated with fibrinogen levels (p<0.0001). sCD40L positively correlated with fasting glucose (r=0.23, p=0.029), basal insulin levels (r=0.34, p=0.002), and HOMA-IR (r=0.37, p=0.001), while hs-CRP weakly correlated with the insulinogenic index (r=0.25, p=0.05) and the area under the OGTT curve for insulin (r=0.32, p=0.027). In a multiple regression model, HOMA-IR emerged as the main determinant of the variability of sCD40L levels, while AUC insulin was the main determinant of the variability of hs-CRP concentrations. In conclusion, minor changes in glucose and insulin metabolism might account at least in part for the inflammatory disorders related to essential hypertension. [ABSTRACT FROM AUTHOR]

Cumulative cardiovascular risk, and cardiovascular disease status alter number and function of circulating endothelial progenitor cells (EPCs). Furthermore, circulating EPCs levels predict cardiovascular events. Diabetes reduces circulating EPCs level. In this scenario, our purpose was to investigate whether pre-diabetes impacts on progenitor cells (CPCs) and EPCs levels. A group of 27 pre-diabetic subjects (8 subjects with impaired fasting glucose, IFG; 14 with impaired glucose tolerance, IGT; and 5 with both IFG and IGT) and a group 22 control individuals have been recruited. No subject was on pharmacological treatments. BMI, waist, OGTT, HbA1c, blood pressure, lipids and inflammatory parameters were measured. Metabolic syndrome (MS) was assessed by NCEP/ATPIII criteria. CPCs (CD34+ cells) and EPCs (CD34+KDR+ cells) were assessed by FACS analysis. Data are expressed as cells number for ml by ISHAGE protocol and as EPCs/CPCs ratio. No differences were observed in EPCs and in EPCs/CPCs in pre-diabetics compared with controls (EPCs: 532±424 vs 511±348/ml, p=0.86; EPCs/CPCs: 28.0±16.1 vs 22.9±17.2%, p--0.30). No correlations were observed between EPCs or EPCs/CPCs and clinical (age, BMI, waist, blood pressure) or laboratory parameters (fasting or 2-h glucose, HbA1c, glucose area trader OGTT, lipids, hs-CRP). In pre-diabetic subjects, EPCs (516±298 vs 542±494/ml, p=0.88) and EPCs/CPCs (27.6±16.4 vs 28.3±16.4%, p=0.93) were superimposable in those with (n. 11, 40%) and without MS. All control subjects were free of MS. A positive first-degree family history for diabetes was present in 10 controls (45%) and 19 pre-diabetic subjects (70%). EPCs (436±334 vs 651±-432/ml, p=0.067) and EPCs/CPCs (21.9±14.3 vs 31.1± 18.5%, p=0.069) were lower in subjects with a family history for diabetes. In conclusions, minor impairments of glucose regulation (pre-diabetes) do not affect circulating EPCs. In pre-diabetic subjectss, EPCs levels are not modified by the co-occurrence of the Metabolic Syndrome. [ABSTRACT FROM AUTHOR]