Quantitative analysis of pancreatic glucokinase gene expression in cultured beta cells by competitive polymerase chain reaction

Regulation of glucokinase (GK) gene expression in pancreatic beta cells has been poorly investigated, both due to low abundance of the gene and to difficulties in cells isolation. The present study describes the establishment of a competitive RT-PCR method for quantitative analysis of GK gene. The method has been applied to the analysis of GK mRNA expression RIN 1046-38 cells. We have monitored modifications of GK mRNA expression after different periods of time in culture and we have studied the effect induced by dexamethasone (DEX) treatment. We show that the method is very sensitive and requires very low amount of RNA. Data demonstrate that GK mRNA expression in RIN cells is reduced as a function of passages in culture and that the reduction is positively correlated with the decrease of insulin responsiveness observed in high passages cells. DEX treatment inhibits GK mRNA expression in RIN cells in a dose-dependent and time-dependent manner

DBI mRNA is expressed in endocrine pancreas and its post-translational product DBI 33-50 inhibits insulin release

It has been previously demonstrated that DBI is present in endocrine pancreas and it is able to inhibit insulin release in isolated rat islets. Its mechanism of action has been investigated, demonstrating the possible involvement of cAMP and ATP-dependent K+ channels. DB133-50, a post-translational product of DBI, is also able to inhibit insulin release, but its action has not been characterized. In the present study, we have investigated the presence of DBI mRNA in pancreas, islets and cultured ß cells. The possible mechanism of action of DBI33-50 and the involvement of BZ/GABAA receptors has been studie

Dose-dependent effect of octreotide on insulin secretion after OGTT in obesity

Objective: The present study aimed at evaluating the acute effect of increasing doses of octreotide (OCT), a long-acting somatostatin analogue, on glucose tolerance and insulin secretion. Methods: A standard and two other oral glucose tolerance tests 30 min after subcutaneous administration of OCT were performed in randomized order in each subject. Obese subjects received 10, 25, or 50 mu g of OCT; control subjects received only 10 and 25 mu g. Fifteen obese and 10 control subjects were studied; all of them had a normal glucose tolerance. Plasma glucose and insulin levels were measured at times -30, 0, 30, 60, 90, 120, 150, and 180 min after the glucose tolerance test. Results: The results demonstrated that, following OCT administration, both control and obese subjects developed a reduced glucose tolerance, a delayed glycemic peak, and an increase of late plasma glucose values. Fasting as well as stimulated insulin secretions were higher in obese subjects as compared with controls, and insulin secretion was inhibited in a dose-dependent manner by OCT. Conclusions: These data indicate that the action of OCT might be due to at least two different cooperative mechanisms: (1) a delayed glucose absorption, as suggested by the delay of glycemic peak, and (2) a direct or vagal-mediated effect on p-cells, as suggested by the reduction of the area under the curve values in spite of the elevated late glycemic levels. It is noteworthy that doses of OCT as low as 10 and 25 mu g are sufficient to inhibit insulin secretion both in normal and obese subjects

Dose-dependent effect of octreotide on insulin secretion after OGTT in obesity

The present study aimed at evaluating the acute effect of increasing doses of octreotide (OCT), a long-acting somatostatin analogue, on glucose tolerance and insulin secretion

Effect of biotin on glucokinase activity, mRNA expression and insulin release in cultured beta-cells

Biotin is known to influence hepatic glucokinase (GK) expression both at a transcriptional and at a translational level. The aim of the present paper was to investigate the effect of biotin on pancreatic GK. For this purpose, RIN1046-38 cells were cultured in the presence of different biotin concentrations for different times; thereafter, GK mRNA expression, GK activity and insulin release were studied. Results demonstrated that biotin has a biphasic effect on GK mRNA expression, being stimulatory after short-term treatment and inhibitory after longterm treatment. GK activity was increased after long-term treatment. Insulin release was not affected by biotin treat ment. These data suggest that biotin may influence glucose metabolism also by acting directly at the level of beta-cells

Increased superoxide anion production by platelets in hypercholesterolemic patients

Objectives: The purpose of this study was to investigate the relationship between hypercholesterolemia and Superoxide anion production.Background: Experimental Studies demonstrated that hypercholesterolemia is associated with enhanced cellular Superoxide anion (O-2(-)) production. Aim of the study was to assess whether the same phenomenon occurs in humans.Methods: Lipid profile and platelet O-2(-) production Acre measured in 28 patients with hypercholesterolemia, compared with 25 age- and sex-matched healthy subjects. and in 21 out of the 28 patients after 8-week treatment with 10 mg/day atorvastatin (a HMGCoA reductase inhibitor). In order to assess the mechanism by which LDL cholesterol interferes kith platelet production of O-2(-), human platelets were incubated with LDL cholesterol in the presence of either an inhibitor of the phospholipaseA2 enzyme, AACOCF3 or an inhibitor of NADH/NADPH oxidases. DPI.Results: O-2(-) platelet generation was significantly higher (p<0.001) and significantly related to LDL cholesterol (p<0.001) in patient,, as Compared to controls. 8-keek treatment with 10 mg/day atorvastatin significantly reduced both LDL cholesterol and O-2(-) platelet production. This effect was partially related to the cholesterol-lowering, in that three days of treatment with atorvastatin significantly decreased platelet O-2(-) production, While no significant change in LDL-cholesterol levels was observed. Platelets incubated kith LDL cholesterol showed an increased O-2(-) production, which as significantly inhibited by, AACOCF3 (-78%) and DPI (-56%).Conclusions: LDL cholesterol increases platelet O-2(-) production by activating PLA2 and NADH/NADPH enzymes. Inhibition of platelet O-2(-) release by atorvastatin is partially related to cholesterol lowering effect, suggesting that other mechanisms could he responsible for the antioxidant activity of the drug

DBI mRNA is expressed in endocrine pancreas and its post-translational product DBI(33-50) inhibits insulin release

It has been previously demonstrated that DBI is present in endocrine pancreas and it is able to inhibit insulin release in isolated rat islets. Its mechanism of action has been investigated, demonstrating the possible involvement of cAMP and ATP-dependent K(+) channels. DB1(33-50), a post-translational product of DBI, is also able to inhibit insulin release, but its action has not been characterized. In the present study, we have investigated the presence of DBI mRNA in pancreas, islets and cultured ß cells. The possible mechanism of action of DBI(33-50) and the involvement of BZ/GABA(A) receptors has been studied