Loss-of-function mutation of the GPR40 gene associates with abnormal stimulated insulin secretion by acting on intracellular calcium mobilization.

BACKGROUND: Free fatty acids (FFAs) acutely stimulate but chronically impair glucose-stimulated insulin secretion from beta-cells. The G protein-coupled transmembrane receptor 40 (GPR40) mediates both acute and chronic effects of FFAs on insulin secretion and plays a role in glucose homeostasis. Limited information is available on the effect of GPR40 genetic abnormalities on insulin secretion and metabolic regulation in human subjects. STUDY DESIGN AND RESULTS: For in vivo studies, we screened 734 subjects for the coding region of GPR40 and identified a new single-nucleotide mutation (Gly180Ser). The mean allele frequency was 0.75%, which progressively increased (P < 0.05) from nonobese subjects (0.42%) to moderately obese (body mass index = 30-39.9 kg/m2, 1.07%) and severely obese patients (body mass index > or = 40 kg/m2, 2.60%). The relationship between the GPR40 mutation, insulin secretion, and metabolic alterations was studied in 11 Gly/Ser mutation carriers. In these subjects, insulin secretion (insulinogenic index derived from oral glucose tolerance test) was significantly lower than in 692 Gly/Gly carriers (86.0 +/- 48.2 vs. 183.7 +/- 134.4, P < 0.005). Moreover, a case-control study indicated that plasma insulin and C-peptide responses to a lipid load were significantly (P < 0.05) lower in six Gly/Ser than in 12 Gly/Gly carriers. In vitro experiments in HeLa cells cotransfected with aequorin and the mutated Gly/Ser GPR40 indicated that intracellular Ca2+ concentration increase after oleic acid was significantly lower than in Gly/Gly GPR40-transfected cells. This fact was confirmed using fura-2 acetoxymethyl ester. CONCLUSIONS: This newly identified GPR40 variant results in a loss of function that prevents the beta-cell ability to adequately sense lipids as an insulin secretory stimulus because of impaired intracellular Ca2+ concentration increase.

Adiponectin relationship with lipid metabolism is independent of body fat mass: evidence from both cross-sectional and intervention studies.

Adiponectin influences insulin sensitivity and lipid metabolism, but it is not clear whether these effects are correlated with fat mass or distribution. We studied the relationship between plasma adiponectin and leptin levels, insulin sensitivity, and serum lipids by a cross-sectional study (n = 242 subjects) and by an intervention study (95 of 242) to evaluate the effect of weight loss (WL). Considering all subjects both together and subdivided into nonobese (n = 107) and obese (n = 135) groups, plasma adiponectin, but not plasma leptin, was significantly (P < 0.01) correlated with insulin sensitivity [homeostasis model assessment of insulin-resistance index (HOMAIR), insulin sensitivity index (ISI) at oral glucose tolerance test, and clamp in 115 of 242 individuals], high-density lipoprotein cholesterol, and triglycerides. These relationships were still significant (P < 0.01) after adjusting for age, gender, body mass index (BMI), and ISI. After WL (-16.8 +/- 0.8%), plasma adiponectin increased, and plasma leptin decreased (P < 0.0001 for both). Their changes (Delta) were significantly correlated with Delta-BMI (P < 0.05 for both). Delta-Adiponectin, but not Delta-leptin, significantly (P < 0.001) correlated with Delta-high-density lipoprotein cholesterol and Delta-triglycerides; these correlations were independent of age, gender, Delta-BMI, and Delta-ISI (P < 0.005). In conclusion, both cross-sectional and intervention studies indicate that plasma adiponectin level correlates with serum lipids independently of fat mass. The intervention study also suggests that adiponectin increase after WL is correlated with serum lipid improvement independently of insulin sensitivity changes.

Optimization of the principal parameters for the ultrarapid electrophoretic separation of reduced and oxidized glutathione by capillary electrophoresis.

Several factors can influence the analytical efficiency and rapidity of the quantitative determination of erythrocyte glutathione by capillary zone electrophoresis (CZE). We optimized the time, efficiency and resolution of the electrophoretic separation of reduced (GSH) and oxidized (GSSG) glutathione by studying the influence of the most important factors affecting the separation, i.e. the pH and ionic strength of the electrolyte solution, the capillary length and temperature. Best results in the shortest time are obtained at 25 degrees C, using an uncoated 37 cm x 75 microm i.d. capillary and a 300 mmol/l borate buffer pH 7.8. These conditions give a good reproducibility of the corrected peak areas (R.S.D. 1.41 and 1.31%) and of the migration time (R.S.D. 0.22 and 0.26%) for GSH and GSSG, respectively. The high concentration buffer, besides permitting a good resolution of standard GSH and GSSG mix, allows also N-nitrosoglutathione detection. By shortening the capillary length to 27 cm, the separation time of GSH and GSSG can be further decreased to less than 60s. This shortened method, the most rapid described in literature, can detect and quantify GSH in red blood cells despite a loss of sensitivity. To compare the new method here described with the Beutler colorimetric method, the data relative to the GSH content of red blood cells from young normal subjects were analyzed by the Passing and Bablok regression and the Bland-Altman test.

Very severely obese patients have a high prevalence of type 2 diabetes mellitus and cardiovascular disease.

The prevalence of very severe obesity has increased progressively and faster than other classes of obesity over the last years. It is unclear whether the prevalence of obesity-related complications and health risks increases progressively or reaches a plateau above a certain degree of obesity. The aim of our study was to investigate whether the severity of obesity was correlated with the prevalence of type 2 diabetes mellitus (T2DM), impaired fasting glucose, impaired glucose tolerance (IGT), metabolic syndrome (MS), and cardiovascular diseases (CVDs) in a large cohort of patients with different degrees of obesity. A cross-sectional study was conducted in 938 obese patients without a previous diagnosis of diabetes. Patients were assigned to different categories of obesity: mild-moderate obesity (BMI 30-39.9 kg/m(2)), morbid obesity (BMI 40-49.9 kg/m(2)), and super-obesity (SO, BMI ≥50 kg/m(2)). The prevalence of IGF, IGT, screen-detected T2DM, MS, and CVD was higher in SO patients than in the other groups. Interestingly, the association between SO and either MS or CVD was independent of glucose tolerance status, indicating that factors other than glucose metabolism also favor cardio-metabolic complications in obese patients. In patients without screen-detected T2DM (n = 807), insulin sensitivity and secretion OGTT-derived indexes indicated that SO patients had the worst glucose homeostasis relative to the other categories of obesity, which was indicated by the most reduced disposition index in these patients, a predictor of future T2DM. In conclusion, SO patients have an extremely high prevalence of glucose metabolism deterioration, and cardio-metabolic complications are more prevalent in these patients compared to less obese patients.

Intragastric balloon in association with lifestyle and/or pharmacotherapy in the long-term management of obesity.

BACKGROUND: Intragastric balloon (BioEnterics Intragastric Balloon, BIB®) or pharmacotherapy are possible options for the treatment of obese patients when traditional approaches have failed. The aim of our study was to compare in obese patients the effect on weight loss and metabolic changes of lifestyle modifications associated with either BIB or pharmacotherapy or the two treatments in sequence as a maintenance strategy for weight loss. METHODS: Fifty obese patients were recruited and randomly assigned to lifestyle modifications combined with either BIB for 6 months (n = 30) or sibutramine (pharmacotherapy group) for 1 year (n = 20). After BIB removal, patients were randomly assigned to either correct lifestyle (BIB/lifestyle) or lifestyle plus pharmacotherapy (BIB/pharmacotherapy). RESULTS: At 6 months, patients treated with BIB lost significantly (P < 0.05) more weight (percent of initial weight lost, %IWL = 14.5 ± 1.2; percent of excess BMI lost, %EBL = 37.7 ± 3.2) than patients who received pharmacological treatment (%IWL = 9.1 ± 1.5, %EBL = 25.3 ± 4.1). At 1 year, the weight lost was significantly (P < 0.05) greater in patients treated with either BIB/pharmacotherapy (%IWL = 15.8 ± 2.3%, %EBL = 41.3 ± 6.7%) or BIB/lifestyle (%IWL = 14.3 ± 2.7, %EBL = 34.9 ± 6.5%) in respect to pharmacotherapy group (%IWL = 8.0 ± 1.4%, %EBL = 22.1 ± 3.9%). Moreover, patients treated sequentially with BIB/lifestyle or BIB/pharmacotherapy showed a significant (P < 0.05) improvement in insulin sensitivity and triglycerides levels. CONCLUSIONS: BIB represents an efficacious long-term obesity treatment when supplemental strategies, as lifestyle modifications or pharmacotherapy, are established for weight maintenance after its removal.

Role of the ENPP1 K121Q polymorphism in glucose homeostasis.

OBJECTIVE: To study the role of the ENPP1 Q121 variant on glucose homeostasis in whites from Italy. RESEARCH DESIGN AND METHODS: We conducted case-control studies in 764 adults (from two independent samples of 289 nonobese and 485 obese individuals) and 240 overweight/obese children undergoing oral glucose tolerance testing (OGTT). Early-phase insulin secretion and insulin sensitivity (the insulinogenic index and the insulin sensitivity index) and their interplay (the disposition index) were calculated. RESULTS: In adult subjects, glucose profiles during OGTT were significantly (P = 2 x 10(-2)) different across K121Q genotype groups and higher in QQ than KK individuals (P = 5 x 10(-2)). The insulinogenic index was significantly reduced in QQ (18.5 +/- 3.4) compared with both KK (31.6 +/- 1.0; P = 2.2 x 10(-7)) and KQ (30.5 +/- 1.5; P = 3.2 x 10(-6)) individuals. KQ individuals also showed a reduced insulin sensitivity index compared with KK subjects (P = 3.6 x 10(-2)). The disposition index was lower in QQ carriers than in KQ and KK individuals (P = 8 x 10(-3) and 4 x 10(-4), respectively) and lower in KQ than in KK individuals (P = 3 x 10(-2)). Data obtained in overweight/obese children were very similar to those observed in adults, with QQ individuals showing (compared with KQ and KK subjects) a reduced insulinogenic index (P = 7 x 10(-3) and 2 x 10(-2), respectively) and disposition index (P = 2 x 10(-2) and 7 x 10(-3), respectively). CONCLUSIONS: Homozygous carriers of the ENPP1 Q121 variant are characterized by an altered glucose homeostasis. Reduced early-phase insulin secretion and inefficient interplay between insulin secretion and sensitivity, which occur at early ages, are major determinants of this defect.