This investigation was made to assess the effect of iron overload on function of pancreatic islet cells in Wistar rats. Sixty-five male rats were randomly divided into four groups: Group A received repeated intraperitoneal (i. p.) injections of ferric nitrilotriacetate (FeNTA); Group B received the equivalent dose of Na2 NTA; Group C received i. p. injection of Diethylenetriaminepentaacetic acid in addition to FeNTA; and Group D rats were untreated controls. Glucose tolerance tests were performed at the beginning, 5th week, and 10th week. Serum iron(SI) and serum ferritin (SF) were measured. The pancreatic tissues were taken for immunohistochemical exam; the levels of Insulin, Glucagon, ss in islets were also evaluated. At the 10th week, the levels of plasma glucose at 2 hours after glucose load in groups A and C were higher than those in groups B and D (P = 0.043); the granules of insulin in beta cells of group A were decreased obviously, the area of islets of group A was smaller than those of other groups (P = 0. 000). Iron overload might influence glycometabolism. And the beta cells' capability to secrete insulin was decreased obviously. Therefore, by way of removing iron, it is possible to protect the rat's glycometabolism to some extent.
Animals ; Glucose Tolerance Test ; Insulin ; secretion ; Insulin-Secreting Cells ; physiology ; secretion ; Iron Overload ; complications ; Male ; Random Allocation ; Rats ; Rats, Wistar

MicroRNAs have been identified as a new class of regulatory molecules that affect many biological functions by interferring the target gene expressions. Latest studies demonstrate that microRNAs can influence many pivotal bio-processes and deeply involve in the metabolism of glucose, lipid and amino acid and biological oxidation. For glucose metabolism, microRNAs are related to insulin secretion, insulin sensitivity, glucose uptake, glycolysis, oxidation and mitochondrial function. For lipid matebolism, microRNAs can regulate the target genes related to lipid biosynthesis, catabolism and transportation. MicroRNAs can influence glutamine catabolism.
Animals ; Glucose ; metabolism ; Glutamine ; metabolism ; Humans ; Insulin ; metabolism ; Insulin Secretion ; Lipid Metabolism ; physiology ; Metabolism ; physiology ; MicroRNAs ; physiology

OBJECTIVETo study the effect of a new obesity-related gene NYGGF4 on the insulin sensitivity and secretory function of adipocytes.

METHODS3T3-L1 preadipocytes transfected with either an empty expression vector (pcDNA3.1; control group) or an NYGGF4 expression vector (NYGGF4-pcDNA3.1) were cultured in vitro and differentiated into the matured adipocytes with the standard insulin plus dexamethasone plus 3-isobutyl-methylxanthine (MDI) induction cocktail. 2-deoxy-D-[3H] glucose uptake was determined by liquid scintillation counting. Western blot was performed to detect the protein content and translocation of glucose transporter 4 (GLUT4). The supernatant concentrations of TNF-alpha, IL-6, adiponectin and resistin were measured using ELISA.

RESULTSNYGGF4 over-expression in 3T3-L1 adipocytes reduced insulin-stimulated glucose uptake. NYGGF4 over-expression impaired insulin-stimulated GLUT4 translocation without affecting the total protein content of GLUT4. The concentrations of TNF-alpha, IL-6, adiponectin and resistin in the culture medium of 3T3-L1 transfected with NYGGF4 were not significantly different from those in the control group.

CONCLUSIONSNYGGF4 over-expression impairs the insulin sensitivity of 3T3-L1 adipocytes through decreasing GLUT4 translocation and had no effects on the secretory function of adipocytes.

3T3-L1 Cells ; Adipocytes ; drug effects ; secretion ; Adiponectin ; secretion ; Animals ; Carrier Proteins ; genetics ; physiology ; Glucose ; metabolism ; Glucose Transporter Type 4 ; analysis ; metabolism ; Insulin ; pharmacology ; Interleukin-6 ; secretion ; Mice ; Resistin ; analysis ; Transfection ; Tumor Necrosis Factor-alpha ; secretion

BACKGROUNDType 2 diabetes is a chronic disease characterized by a progressive loss of beta cell functions. However, the evaluation of beta cell functions is either expensive or inconvenient for clinical practice. We aimed to elucidate the association between the changes of insulin responsiveness and the fasting plasma glucose (FPG) during the development of diabetes.

METHODSA total of 1192 Chinese individuals with normal blood glucose or hyperglycemia were enrolled for the analysis. The early insulinogenic index (DeltaI30/DeltaG30), the area under the curve of insulin (AUC-I), and homeostasis model assessment were applied to evaluate the early phase secretion, total insulin secretion, and insulin resistance respectively. Polynomial regression analysis was performed to estimate the fluctuation of beta cell functions.

RESULTSThe DeltaI30/DeltaG30 decreased much more rapidly than the AUC-I accompanying with the elevation of FPG. At the FPG of 110 mg/dl (a pre-diabetic stage), the DeltaI30/DeltaG30 lost 50% of its maximum while the AUC-I was still at a compensated normal level. The AUC-I exhibited abnormal and decreased gradually at the FPG of from 130 mg/dl to higher (overt diabetes), while the DeltaI30/DeltaG30 almost remained at 25% of its maximum value. When hyperglycemia continuously existed at > 180 mg/dl, both the DeltaI30/DeltaG30 and AUC-I were totally lost.

CONCLUSIONThe increased fasting plasma glucose reflects progressive decompensation of beta cell functions, and could be used to guide the strategy of clinical treatments.

Adult ; Aged ; Aged, 80 and over ; Blood Glucose ; analysis ; Diabetes Mellitus ; blood ; physiopathology ; Fasting ; blood ; Female ; Humans ; Insulin ; secretion ; Insulin Resistance ; Insulin-Secreting Cells ; physiology ; Male ; Middle Aged

OBJECTIVETo elucidate GPR40/FFA1 and its downstream signaling pathways in regulating insulin secretion.

METHODSGPR40/FFA1 expression was detected by immunofluorescence imaging. We employed linoleic acid (LA), a free fatty acid that has a high affinity to the rat GPR40, and examined its effect on cytosolic free calcium concentration ([Ca2+]i) in primary rat beta-cells by Fluo-3 intensity under confocal microscopy recording. Downregulation of GPR40/FFA1 expression by antisense oligonucleotides was performed in pancreatic beta-cells, and insulin secretion was assessed by enzyme-linked immunosorbent assay.

RESULTSLA acutely stimulated insulin secretion from primary cultured rat pancreatic islets. LA induced significant increase of [Ca2+]i in the presence of 5.6 mmol/L and 11.1 mmol/L glucose, which was reflected by increased Fluo-3 intensity under confocal microscopy recording. LA-stimulated increase in [Ca2+]i and insulin secretion were blocked by inhibition of GPR40/FFA1 expression in beta-cells after GPR40/FFA1-specific antisense treatment. In addition, the inhibition of phospholipase C (PLC) activity by U73122, PLC inhibitor, also markedly inhibited the LA-induced [Ca2+]i increase.

CONCLUSIONLA activates GPR40/FFA1 and PLC to stimulate Ca2+ release, resulting in an increase in [Ca2+]i and insulin secretion in rat islet beta-cells.

Animals ; Calcium ; metabolism ; Enzyme Activation ; Insulin ; secretion ; Insulin-Secreting Cells ; drug effects ; metabolism ; secretion ; Linoleic Acid ; pharmacology ; Male ; Rats ; Rats, Sprague-Dawley ; Receptors, G-Protein-Coupled ; physiology ; Type C Phospholipases ; physiology

BACKGROUNDAbnormal insulin secretion of pancreatic beta cells is now regarded as the more primary defect than the insulin function in the etiology of type 2 diabetes. Previous studies found impaired mitochondrial function and impaired Ca(2+) influx in beta cells in diabetic patients and animal models, suggesting a role for these processes in proper insulin secretion. The aim of this study was to investigate the detailed relationship of mitochondrial function, Ca(2+) influx, and defective insulin secretion.

METHODSWe investigated mitochondrial function and morphology in pancreatic beta cell of diabetic KK-Ay mice and C57BL/6J mice. Two types of Ca(2+) channel activities, L-type and store-operated Ca(2+) (SOC), were evaluated using whole-cell patch-clamp recording. The glucose induced Ca(2+) influx was measured by a non-invasive micro-test technique (NMT).

RESULTSMitochondria in KK-Ay mice pancreatic beta cells were swollen with disordered cristae, and mitochondrial function decreased compared with C57BL/6J mice. Ca(2+) channel activity was increased and glucose induced Ca(2+) influx was impaired, but could be recovered by genipin.

CONCLUSIONDefective mitochondrial function in diabetic mice pancreatic beta cells is a key cause of abnormal insulin secretion by altering Ca(2+) influx, but not via Ca(2+) channel activity.

Animals ; Calcium ; metabolism ; Diabetes Mellitus ; metabolism ; physiopathology ; Electrophysiology ; Insulin ; secretion ; Insulin-Secreting Cells ; metabolism ; Male ; Membrane Potential, Mitochondrial ; physiology ; Mice ; Mice, Inbred C57BL ; Mitochondria ; metabolism

The purpose of the present study is to investigate the effects of urotensin II (UII) on insulin secretion in islet beta cells and the underlying mechanism. Glucose tolerance test was performed in Wistar rats to evaluate the effect of UII on the levels of plasma glucose and insulin. Static incubation experiment was employed to investigate the effect of UII on glucose-induced insulin secretion (GIIS) in betaTC-6 cells. After the incubation, insulin content and mRNA level in betaTC-6 cells were analyzed. Finally, Western blot was used to find out if UII could change the expression levels of pancreatic duodenal homeobox-1 (PDX-1) and glucokinase (GCK). It was observed that intravenous administration of UII (30, 300 nmol/kg) resulted in a significant decrease in insulin level 15 min after glucose load, and induced an obvious increase in plasma glucose 90 min after the load. In vitro, two hours of UII incubation inhibited GIIS in betaTC-6 cells without affecting insulin content and mRNA levels. The inhibitory effect of UII was blocked by UII receptor antagonist (urantide), and partially blunted by protein kinase C (PKC) inhibitor (chelerythrine) and somatostatin receptor antagonist (cyclosomatostatin). Moreover, we found that GCK protein level was significantly reduced by UII, while PDX-1, a key regulator of insulin gene transcription in beta cells, was not affected. These results suggest that UII-induced inhibition of GIIS in betaTC-6 cells are mediated by UII receptor and PKC pathway, as well as somatostatin receptor which could be activated by high dose of UII. The inhibitory effect of UII on insulin secretion is rather associated with a suppression of GCK expression than a regulation on PDX-1 expression.
Animals ; Blood Glucose ; metabolism ; Cell Line ; Glucokinase ; metabolism ; Homeodomain Proteins ; metabolism ; Insulin ; secretion ; Insulin-Secreting Cells ; metabolism ; physiology ; secretion ; Male ; Mice ; Rats ; Rats, Wistar ; Trans-Activators ; metabolism ; Urotensins ; pharmacology

Animals ; Glucose ; pharmacology ; Homeodomain Proteins ; analysis ; genetics ; physiology ; Immunohistochemistry ; Insulin ; secretion ; Male ; RNA, Messenger ; analysis ; Rats ; Rats, Sprague-Dawley ; Trans-Activators ; analysis ; genetics ; physiology

BACKGROUNDIslet transplantation represents an ideal therapeutic approach for treatment of type 1 diabetes but islet function and regeneration may be influenced by necrosis or apoptosis induced by oxidative stress and other insults. Heme oxygenase-1 (HO-1) is the rate-limiting enzyme in the catabolism of heme into biliverdin, releasing free iron and carbon monoxide. It has also been reported to be an antioxidant enzyme which can improve the function of grafted islets by cytoprotection via free radical scavenging and apoptosis prevention. In the present study, we investigated whether transduction of HO-1 genes into human islets with an adenovirus vector has cytoprotective action on islets cultured in vitro and discuss this method of gene therapy for clinical islet transplantation.

METHODSCadaveric pancreatic islets were isolated and purified in vitro. Transduction efficiency of islets was determined by infecting islets with adenovirus vector containing the enhanced green fluorescent protein gene (Ad-EGFP) at multiplicities of infection (MOI) of 2, 5, 10, or 20. Newly isolated islets were divided into three groups: EGFP group, islets transduced with Ad-EGFP using MOI = 20; HO-1 group, transduced with adenovirus vectors containing the human HO-1 gene using MOI = 20; and control group, mock transduced islets. Insulin release after glucose stimulation of the cell lines was determined by a radioimmunoassay kit and the stimulation index was calculated. Flow cytometry was used to detect apoptotic cells in the HO-1 group and in the control group after induction by recombinant human tumor necrosis factor-alpha (rTNFalpha) and cycloheximide (CHX) for 48 hours.

RESULTSAdenovirus vectors have a high efficiency of gene transduction into adult islet cells. Transduction of islets with the Ad-EGFP was most successful at MOI 20, at which MOI fluorescence was very intense on day 7 after transduction and EGFP was expressed in cultured islet cells for more than four weeks in vitro. The insulin release in the control group was (182.36 +/- 58.96) mIU/L after stimulation by high glucose media (16.7 mmol/L), while insulin release from the HO-1 group and the EGFP group were (270.09 +/- 89.37) mIU/L and (175.95 +/- 75.05) mIU/L respectively. Compared to the control group and the EGFP group, insulin release in the HO-1 group increased significantly (P < 0.05). After treatment with rTNFalpha and CHX the apoptotic ratio of islet cells was (63.09 +/- 10.86)% in the HO-1 group, significantly lower than (90.86 +/- 11.25)% in the control group (P < 0.05).

CONCLUSIONSTransduction of human islets with Ad-HO-1 can protect against TNF-alpha and CHX mediated cytotoxicity. The HO-1 gene also appears to facilitate insulin release from human islets. Transduction of donor islets with the adenovirus vector containing an HO-1 gene might have potential value in clinical islet transplantation.

Adenoviridae ; genetics ; Apoptosis ; drug effects ; Cycloheximide ; pharmacology ; Cytoprotection ; Genetic Therapy ; Heme Oxygenase-1 ; genetics ; physiology ; Humans ; Insulin ; secretion ; Islets of Langerhans ; physiology ; Transduction, Genetic ; Tumor Necrosis Factor-alpha ; pharmacology

OBJECTIVETo investigate the relationship of insulin resistance and the polymorphisms of insulin receptor-related genes in essential hypertension patients of two different kinds of TCM constitution.

METHODSOral glucose tolerance test (OGTT) and insulin release test (InRT) were conducted in 217 essential hypertensive patients of either sluggish meticulous (SM) constitution (139 cases) or prosperous impetuous (PI) constitution (78 cases), and the polymorphism of three genes, including insulin-like growth factor-1 receptor (IGF-1R), insulin receptor substrate-1 (IRS-1) and 2 (IRS-2) genes were detected.

RESULTS(1) OGTT, InRT and insulin resistance index (Homa-IR) were higher and insulin sensitive index (ISI) was lower in the patients of SM constitution than those in patients of PI constitution. (2) Significant difference of ISI and Homa-IR was shown in patients of both constitutions with genotype G of the 3 genes.

CONCLUSIONDecrease of insulin sensitivity and increase of insulin resistance are more obvious in hypertensive patients with genotype G of the 3 genes of SM constitution than in those of PI constitution. Therefore, the difference in constitution might be one of the genetic characteristics for insulin resistance in hypertensive patients.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Blood Glucose ; Body Constitution ; physiology ; Female ; Glucose Tolerance Test ; Humans ; Hypertension ; genetics ; Insulin ; secretion ; Insulin Resistance ; physiology ; Male ; Middle Aged ; Phenotype ; Polymorphism, Genetic