Islets Of Obese-hyperglycemic Mice Research Articles

Effect of rosiglitazone on the differential expression of diabetes-associated proteins in pancreatic islets of C57Bl/6 lep/lep mice.

The insulin sensitizer drug, rosiglitazone, has been shown to have a protective effect on pancreatic islet cell structure and function in animal models of type 2 diabetes. The identification of new molecular targets associated both with islet cell dysfunction and protection is a crucial research goal. In the present study, a proteomics approach has been used to identify such targets. Obese C57Bl/6J lep/lep mice and lean littermates were given the insulin sensitizer drug BRL49653, rosiglitazone. It normalized the impaired glucose tolerance in lep/lep mice but had no significant effect on glucose tolerance in the lean mice. Pancreatic islet polypeptides were arrayed by a two-dimensional gel electrophoresis system that separated more than 2500 individual spots. Three overexpressed and six underexpressed proteins were significant (p < 0.05) between lep/lep and lean mice, and four were modulated significantly (p < 0.05) by the rosiglitazone treatment of the obese mice. The identity of these differentially expressed proteins was made using mass spectrometric analysis and provided evidence that differential expression of actin-binding proteins may be an important aspect of defective islet function. Rosiglitazone increased carboxypeptidase B expression in both lep/lep and normal mice suggesting that this might be an independent effect of rosiglitazone that contributes to improved insulin processing.

Glucose Cycling Is Markedly Enhanced in Pancreatic Islets of Obese Hyperglycemic Mice*

Pancreatic islets from fed 7-month old lean and obese hyperglycemic mice (ob/ob) were incubated with 3H2O and 5.5 mM or 16.7 mM glucose. Incorporation of 3H into the medium glucose was taken as the measure of glucose-6-P hydrolysis to glucose. Glucose utilization was measured from the yield of 3H2O from [5-3H]glucose. Only 3-4% of the glucose phosphorylated was dephosphorylated by the lean mouse islets irrespective of the glucose concentration. In contrast, the ob/ob mouse islets at 5.5 mM glucose dephosphorylated 18% of the glucose phosphorylated and 30% at 16.7 mM. Thus, the islets of hyperglycemic mice demonstrate increased glucose cycling as compared to the islets of normoglycemic lean mice.

Studies on the role of β-cell metabolism in the insulinotropic effect of α-ketoisocaproic acid

α-Ketoisocaproic acid has been shown to be apotent insulin secretagogue but the mechanism has not been elucidated. To define the role of β-cell metabolism in the insulinotropic activity of α-ketoisocaproic acid the utilization of glucose and the oxidation of α-ketoisocaproic and isovaleric acid by incubated islets of obese hyperglycemic mice were measured. Glucose metabolism was never enhanced by α-ketoisocaproic acid. The same 14CO 2 amounts were released from the non-secretagogue [1- 14C]isovaleric acid (10 mM) or from α-keto [2- 14C]isocaproic acid (5–20 mM). Pyruvate (20 mM) did not inhibit α-ketoisocaproic acid-induced insulin secretion in spite of reduction of decarboxylation of α-ketoisocaproic acid by more than 40%. The results indicate that stimulated insulin release in response to α-ketoisocaproic acid is not mediated by an indirect increase in glucose metabolism and further suggest that isovaleryl-CoA and following CoA-esters in α-ketoisocaproic acid degradation are not likely recognized as signals. The possibility, however, remains that enhanced intramitochondrial production of reducing equivalents elicits insulin secretion.

Somatostatin content and release of isolated pancreatic islets from obese-hyperglycemic mice

The somatostatin content in pancreatic islets of obese-hyperglycemic mice was much lower than in the islets of normal mice. Also the release of somatostatin was decreased from the islets obtained from the obese-hyperglycemic mice. Tissue culture for 1 week changed neither the content of, nor the amount of somatostatin released from, the pancreatic islets.

Hypothesis: Physiologic role of pancreatic somatostatin and the contribution of D-cell disorders to diabetes mellitus

The demonstration of direct inhibitory actions of pharmacologic doses of somatostatin upon gastrointestinal function suggests the possibility that somatostatin may be a physiologic regulator of the rate of nutrient entry from the gut and thus provide the A-, B-, and D-cells with an influence over all avenues of nutrient flux. If so, the relative reduction in somatostatin-containing cells in the pancreatic islets of obese hyperglycemic mice might be responsible for some or all of the metabolic derangements of the obese hyperglycemic syndrome of mice, and conceivably its human counterpart as well. Hyposomatostatinemia would permit accelerated gastrointestinal function with more rapid entry of nutrients; higher postprandial nutrient levels would cause hyperinsulinemia and reduce the insulin sensitivity of glucoregulatory tissues. As hyperglycemia increased, the B-cell response to hyperglycemia would wane, further escalating the metabolic derangement. By contrast with the obese hyperglycemic syndrome, diabetes characterized by marked dimunution in B-cells is associated with an apparent augmentation of D-cells, perhaps reflecting a futile compensatory effort of D-cells to correct the hyperglycemia of insulin deficiency by inhibiting the entry rate of exogenous nutrients.

The effect of fatty acids and ketone bodies on the biosynthesis of insulin in isolated pancreatic islets of obese hyperglycemic mice.

Collagenase-isolated islets from obese hyperglycemic mice were used to study the effects of fatty acids and ketone bodies on insulin biosynthesis. The biosynthetic rate was quantitated as the incorporation of 3H-leucine into proinsulin and insulin, which were separated from the crude islet homogenates with anti-insulin serum coupled to cyanogen bromide activated Sepharose 4B. Palmitate and oleate did not stimulate the incorporation into proinsulin and insulin while 16.7 mM glucose caused a nine-fold increase of 3H-leucine incorporation during a 3 h incubation as compared to the basal buffer alone. A weak stimulatory effect was seen with 5 mM octanoate. The ketone bodies had a weak stimulating effect on islet protein biosynthesis, but no preferential stimulus to the insulin biosynthesis was found.

Melatonin inhibition of insulin secretion in the rat and mouse.

The effect of melatonin (100 µg/ml) on basal and glucose-induced insulin secretion was investigated using micro-dissected pancreatic islets of obese hyper-glycaemic mice and their normal lean littermates, and pieces of rat pancreas. Melatonin had no significant effect on basal insulin secretion from both normal and obese mouse islets, but significantly reduced basal insulin secretion from pieces of rat pancreas. However, glucose-induced insulin secretion was significantly inhibited by melatonin in both preparations. A constant infusion of melatonin into rats undergoing intravenous glucose tolerance tests did not alter blood sugar levels and produced only a marginal decrease in plasma insulin levels. It is suggested that melatonin may be involved in the monoaminergic regulation of insulin secretion.

Factors affecting the uptake of glibenclamide in microdissected pancreatic islets rich in beta-cells.

The uptake of radioactively labelled glibenclamide was studied in microdissected pancreatic islets of obese-hyperglycemic mice. The steady-state concentration reached was linearly related to the medium concentration of glibenclamide. This is in contrast to the short-term uptake, for which there was evidence for saturation. The amounts of glibenclamide incorporated increased by more than 100 % when the plasma membranes were made permeable to sucrose by reducing the osmolality of the incubation medium. No binding of glibenclamide in excess of sucrose could be detected after increasing the albumin concentration to 10 mg/ml. The steady-state concentrations reached were approximately the same in the presence of different monovalent cations. The observation of a reduced uptake of glibenclamide in the presence of cysteine raises the question whether sulfonylureas stimulate insulin release by reacting with sulf-hydryl groups in the /3-cell membrane.

Transport and oxidation of glycine in mammalian pancreatic islets with reference to the mechanism of amino acid-induced insulin release.

Microdissected pancreatic islets of obese-hyperglycemic mice were used for studying the uptake, oxidation and insulin-releasing ability of glycine. Glycine did not influence insulin release either in the presence or absence of glucose. It was rapidly taken up by the islet cells, reaching distribution ratios much higher than unity. This concentrative uptake was depressed by the omission of sodium from the medium. L-Alanine, L-leucine, D-glucose, or glibenclamide also reduced the glycine uptake, whereas diazoxide had no effect. The production of radioactive CO&lt;sub&gt;2&lt;/sub&gt; from uniformly &lt;sup&gt;14&lt;/sup&gt;C-labeled glycine remained insignificant even when tested at concentrations as high as 20 mM. It is suggested that the &lt;i&gt;β&lt;/i&gt;-cells transport glycine by the alanine-preferring A-system. The data are consistent with our previous concept that this system does not represent a trigger site for insulin release from mature mouse-&lt;i&gt;β&lt;/i&gt;-cells. Glycine appears to be of minor importance as a fuel for the &lt;i&gt;β&lt;/i&gt;-cells.

The pancreatic β-cell recognition of insulin secretagogues. II. Site of action of tolbutamide

The distribution of 35S-labelled tolbutamide was studied in microdissected pancreatic islets of obese-hyperglycemic mice. These islets contain more than 90 % β-cells. A comparison with the uptake of 3H-labelled sucrose, mannitol, or 3-O-methyl-D-glucose revealed that tolbutamide did not enter the β-cells but was restricted to the extracellular space. It is suggested that the β-cell plasma membrane contains a tolbutamide receptor, which is responsible for the recognition of sulfonylureas as insulin secretagogues.

Accumulation of citrate in pancreatic islets of obese hyperglycemic mice

Accumulation of citrate in pancreatic islets of obese hyperglycemic mice

Quantitative studies on isolated pancreatic islets of mammals. Activity and heterogeneity of lactate dehydrogenase in obese-hyporglycemic mice.

Histochemical staining, microchemical assays and disc electrophoresis were employed in a study of the lactate dehydrogenase activity present in the pancreatic islets of obesehyperglycemic mice. The following observationswere made: 1. By means of a histochemical substrate-film technique, a weaker staining for LDH activity was noted in the islet β cells than in the acinar tissue. Addition of 0.08 mM phenazine methosulfate did not apparently influence the degree of staining, either in the endocrine or in the exocrine pancreas. 2. Quantitative microchemical assays on crude tissue homogenates revealed a significantly lower LDH activity in the pancreatic islets than in the exocrine parenchyma. A considerably lower ratio between the dehydrogenase activities toward 2-oxobutyrate and pyruvate was observed in both the endocrine and exocrine pancreas, as compared with the heart. 3. Five LDH isoenzymes were separated electrophoretically, 65 % of the total enzyme activity being represented by LDH-5. A theoretical calc...