Estrogen replacement therapy reduces the incidence of type 2 diabetes in postmenopausal women; however, the mechanism is unknown. Therefore, the aim of this study was to evaluate the metabolic effects of estrogen replacement therapy in an experimental model of menopause. At 8 weeks of age, female mice were ovariectomized (OVX) or sham (SHAM) operated, and OVX mice were treated with vehicle (OVX) or estradiol (E2) (OVX+E2). After 4 weeks of high-fat diet feeding, OVX mice had increased body weight and fat mass compared with SHAM and OVX+E2 mice. OVX mice displayed reduced whole-body energy expenditure, as well as impaired glucose tolerance and whole-body insulin resistance. Differences in whole-body insulin sensitivity in OVX compared with SHAM mice were accounted for by impaired muscle insulin sensitivity, whereas both hepatic and muscle insulin sensitivity were impaired in OVX compared with OVX+E2 mice. Muscle diacylglycerol (DAG), content in OVX mice was increased relative to SHAM and OVX+E2 mice. In contrast, E2 treatment prevented the increase in hepatic DAG content observed in both SHAM and OVX mice. Increases in tissue DAG content were associated with increased protein kinase Cϵ activation in liver of SHAM and OVX mice compared with OVX+E2 and protein kinase Cθ activation in skeletal muscle of OVX mice compared with SHAM and OVX+E2. Taken together, these data demonstrate that E2 plays a pivotal role in the regulation of whole-body energy homeostasis, increasing O₂ consumption and energy expenditure in OVX mice, and in turn preventing diet-induced ectopic lipid (DAG) deposition and hepatic and muscle insulin resistance.

The presence of a phagocyte-like NAD(P)H oxidase in pancreatic β-cells was investigated. Three NAD(P)H oxidase components were found in pancreatic islets by RT-PCR: gp91PHOX, p22PHOX, and p47PHOX. The components p67PHOX and p47PHOX were also demonstrated by Western blotting. Through immunohistochemistry, p47PHOX was mainly found in the central area of the islet, confirming the expression of this component by insulin-producing cells. Activation of NAD(P)H oxidase complex in the β-cells was also examined by immunohistochemistry. The pancreatic islets presented slower kinetics of superoxide production than HIT-T15 cells, neutrophils, and macrophages, but they reached 66% that of the neutrophil nitroblue tetrazolium (NBT) reduction after 2 h of incubation. Glucose (5.6 mmol/l) increased NBT reduction by 75% when compared with control. The involvement of protein kinase C (PKC) in the stimulatory effect of glucose was confirmed by incubation of islets with phorbol myristate acetate (a PKC activator) and bysindoylmaleimide (GF109203X) (a PKC-specific inhibitor). Diphenylene iodonium [an NAD(P)H oxidase inhibitor] abolished the increase of NBT reduction induced by glucose, confirming the NAD(P)H oxidase activity in pancreatic islets. Because reactive oxygen species are involved in intracellular signaling, the phagocyte-like NAD(P)H oxidase activation by glucose may play an important role for β-cell functioning.

Hyperlipidemia is frequently associated with insulin resistance states as found in type 2 diabetes and obesity. Effects of free fatty acids (FFA) on pancreatic β‐cells have long been recognized. Acute exposure of the pancreatic β‐cell to FFA results in an increase of insulin release, whereas a chronic exposure results in desensitization and suppression of secretion. We recently showed that palmitate augments insulin release in the presence of non‐stimulatory concentrations of glucose. Reduction of plasma FFA levels in fasted rats or humans severely impairs glucose‐induced insulin release. These results imply that physiological plasma levels of FFA are important for β‐cell function. Although, it has been accepted that fatty acid oxidation is necessary for its stimulation of insulin secretion, the possible mechanisms by which fatty acids (FA) affect insulin secretion are discussed in this review. Long‐chain acyl‐CoA (LC‐CoA) controls several aspects of the β‐cell function including activation of certain types of protein kinase C (PKC), modulation of ion channels, protein acylation, ceramide‐ and/or nitric oxide (NO)‐mediated apoptosis, and binding to nuclear transcriptional factors. The present review also describes the possible effects of FA on insulin signaling. We showed for the first time that acute exposure of islets to palmitate upregulates the intracellular insulin‐signaling pathway in pancreatic islets. Another aspect considered in this review is the source of FA for pancreatic islets. In addition to be exported to the medium, lipids can be transferred from leukocytes (macrophages) to pancreatic islets in co‐culture. This process consists an additional source of FA that may plays a significant role to regulate insulin secretion.

A high fructose diet induces insulin resistance in rats, although the exact molecular mechanism involved is unknown. In this study, we used immunoprecipitation and immunoblotting to examine the levels and phosphorylation status of the insulin receptor (IR) and insulin receptor substrate-1 (IRS-1), as well as the association of the IRS-1 with phosphatidylinositol 3-kinase (PI 3-kinase), and phosphotyrosine phosphatase (SHP2) in the liver and muscle of rats fed a control or high fructose diet for 28 d. There were no differences in IR and the IRS-1 protein levels in the liver and muscle of rats fed the control and high fructose diets. However, tyrosine-phosphorylation of the insulin receptor after insulin stimulation was reduced to 71 ± 2% (P < 0.05) of control in the liver of the fructose-fed rats. In samples previously immunoprecipitated with anti-IRS-1 antibody and blotted with antiphosphotyrosine antibody, the insulin-stimulated IRS-1 phosphorylation levels in the liver and muscle of the fructose-fed group were only 70 ± 6% (P < 0.05) and 76 ± 5% (P < 0.05) of those of control rats, respectively. The insulin-stimulated IRS-1 association with PI 3-kinase was reduced to 84 ± 3% (P < 0.05) in the liver and to 84 ± 4% (P < 0.05) in the muscle of the fructose-fed group compared with control rats. Insulin-stimulated IRS-1 association with SHP2 was reduced to 79 ± 5% (P < 0.05) in liver of the fructose-fed rats. These data suggest that changes in the early steps of insulin signal transduction may have an important role in the insulin resistance observed in these rats.

Objective We investigated the effects of high-fat diet-induced obesity on vascular proinflammatory factors and oxidative stress on endothelium-dependent relaxation of the aorta. Methods Female Swiss mice were submitted to a high-fat diet for 16 weeks. At the end of the experimental period, we evaluated blood pressure, relaxation in response to acetylcholine in aortic rings in the absence and the presence of the superoxide anion scavenger, superoxide dismutase (SOD, 150 U/ml), and the nuclear factor (NF)-κB inhibitor, sodium salicylate (5 mmol/l). Aortic protein expression of endothelial nitric oxide synthase, Cu/Zn-SOD, NF-κB, IκB-α, and proinflammatory cytokines were also evaluated. Results Obese mice presented higher systolic and diastolic blood pressure than control mice (P < 0.05). The relaxation of aortas to acetylcholine, but not to sodium nitroprusside, was significantly decreased in obese mice and was corrected by both SOD and sodium salicylate (P < 0.05). The protein expression of endothelial nitric oxide synthase and Cu/Zn-SOD was significantly decreased in aorta from obese mice (P < 0.05). Total p65 NF-κB subunit protein expression was not affected by obesity, but the protein expression of NF-κB inhibitor IκB-α was lower in aorta from obese mice (P < 0.05). There were no significant differences in the interleukin (IL)-1β and IL-6 protein expression between groups. In contrast, the expression of TNF-α was significantly increased in aortas from obese mice. Conclusion Our results suggest that the reduced antioxidant defense and the local NF-κB pathway play an important role in the impairment of endothelium-dependent relaxation in aorta from obese mice.