Increased Hypothalamic Anti-Inflammatory Mediators in Non-Diabetic Insulin Receptor Substrate 2-Deficient Mice.

Insulin receptor substrate (IRS) 2 is a key mediator of insulin signaling and IRS-2 knockout (IRS2-/-) mice are a preclinical model to study the development of diabetes, as they develop peripheral insulin resistance and beta-cell failure. The differential inflammatory profile and insulin signaling in the hypothalamus of non-diabetic (ND) and diabetic (D) IRS2-/- mice might be implicated in the onset of diabetes. Because the lipid profile is related to changes in inflammation and insulin sensitivity, we analyzed whether ND IRS2-/- mice presented a different hypothalamic fatty acid metabolism and lipid pattern than D IRS2-/- mice and the relationship with inflammation and markers of insulin sensitivity. ND IRS2-/- mice showed elevated hypothalamic anti-inflammatory cytokines, while D IRS2-/- mice displayed a proinflammatory profile. The increased activity of enzymes related to the pentose-phosphate route and lipid anabolism and elevated polyunsaturated fatty acid levels were found in the hypothalamus of ND IRS2-/- mice. Conversely, D IRS2-/- mice have no changes in fatty acid composition, but hypothalamic energy balance and markers related to anti-inflammatory and insulin-sensitizing properties were reduced. The data suggest that the concurrence of an anti-inflammatory profile, increased insulin sensitivity and polyunsaturated fatty acids content in the hypothalamus may slow down or delay the onset of diabetes.

Cerebral Insulin Bolus Revokes the Changes in Hepatic Lipid Metabolism Induced by Chronic Central Leptin Infusion.

Central actions of leptin and insulin on hepatic lipid metabolism can be opposing and the mechanism underlying this phenomenon remains unclear. Both hormones can modulate the central somatostatinergic system that has an inhibitory effect on growth hormone (GH) expression, which plays an important role in hepatic metabolism. Using a model of chronic central leptin infusion, we evaluated whether an increase in central leptin bioavailability modifies the serum lipid pattern through changes in hepatic lipid metabolism in male rats in response to an increase in central insulin and the possible involvement of the GH axis in these effects. We found a rise in serum GH in leptin plus insulin-treated rats, due to an increase in pituitary GH mRNA levels associated with lower hypothalamic somatostatin and pituitary somatostatin receptor-2 mRNA levels. An augment in hepatic lipolysis and a reduction in serum levels of non-esterified fatty acids (NEFA) and triglycerides were found in leptin-treated rats. These rats experienced a rise in lipogenic-related factors and normalization of serum levels of NEFA and triglycerides after insulin treatment. These results suggest that an increase in insulin in leptin-treated rats can act on the hepatic lipid metabolism through activation of the GH axis.

Leptin Modulates the Response of Brown Adipose Tissue to Negative Energy Balance: Implication of the GH/IGF-I Axis.

The growth hormone (GH)/insulin-like growth factor I (IGF-I) axis is involved in metabolic control. Malnutrition reduces IGF-I and modifies the thermogenic capacity of brown adipose tissue (BAT). Leptin has effects on the GH/IGF-I axis and the function of BAT, but its interaction with IGF-I and the mechanisms involved in the regulation of thermogenesis remains unknown. We studied the GH/IGF-I axis and activation of IGF-I-related signaling and metabolism related to BAT thermogenesis in chronic central leptin infused (L), pair-fed (PF), and control rats. Hypothalamic somatostatin mRNA levels were increased in PF and decreased in L, while pituitary GH mRNA was reduced in PF. Serum GH and IGF-I concentrations were decreased only in PF. In BAT, the association between suppressor of cytokine signaling 3 and the IGF-I receptor was reduced, and phosphorylation of the IGF-I receptor increased in the L group. Phosphorylation of Akt and cyclic AMP response element binding protein and glucose transporter 4 mRNA levels were increased in L and mRNA levels of uncoupling protein-1 (UCP-1) and enzymes involved in lipid anabolism reduced in PF. These results suggest that modifications in UCP-1 in BAT and changes in the GH/IGF-I axis induced by negative energy balance are dependent upon leptin levels.

Sex Differences in Metabolic Recuperation After Weight Loss in High Fat Diet-Induced Obese Mice.

Dietary intervention is a common tactic employed to curtail the current obesity epidemic. Changes in nutritional status alter metabolic hormones such as insulin or leptin, as well as the insulin-like growth factor (IGF) system, but little is known about restoration of these parameters after weight loss in obese subjects and if this differs between the sexes, especially regarding the IGF system. Here male and female mice received a high fat diet (HFD) or chow for 8 weeks, then half of the HFD mice were changed to chow (HFDCH) for 4 weeks. Both sexes gained weight (p < 0.001) and increased their energy intake (p < 0.001) and basal glycemia (p < 0.5) on the HFD, with these parameters normalizing after switching to chow but at different rates in males and females. In both sexes HFD decreased hypothalamic NPY and AgRP (p < 0.001) and increased POMC (p < 0.001) mRNA levels, with all normalizing in HFDCH mice, whereas the HFD-induced decrease in ObR did not normalize (p < 0.05). All HFD mice had abnormal glucose tolerance tests (p < 0.001), with males clearly more affected, that normalized when returned to chow. HFD increased insulin levels and HOMA index (p < 0.01) in both sexes, but only HFDCH males normalized this parameter. Returning to chow normalized the HFD-induced increase in circulating leptin (p < 0.001), total IGF1 (p < 0.001), IGF2 (p < 0.001, only in females) and IGFBP3 (p < 0.001), whereas free IGF1 levels remained elevated (p < 0.01). In males IGFBP2 decreased with HFD and normalized with chow (p < 0.001), with no changes in females. Although returning to a healthy diet improved of most metabolic parameters analyzed, fIGF1 levels remained elevated and hypothalamic ObR decreased in both sexes. Moreover, there was sex differences in both the response to HFD and the switch to chow including circulating levels of IGF2 and IGFBP2, factors previously reported to be involved in glucose metabolism. Indeed, glucose metabolism was also differentially modified in males and females, suggesting that these observations could be related.

Increased oxidative stress and apoptosis in the hypothalamus of diabetic male mice in the insulin receptor substrate-2 knockout model.

Insulin receptor substrate-2-deficient (IRS2(-/-)) mice are considered a good model to study the development of diabetes because IRS proteins mediate the pleiotropic effects of insulin-like growth factor-I (IGF-I) and insulin on metabolism, mitogenesis and cell survival. The hypothalamus might play a key role in the early onset of diabetes, owing to its involvement in the control of glucose homeostasis and energy balance. Because some inflammatory markers are elevated in the hypothalamus of diabetic IRS2(-/-) mice, our aim was to analyze whether the diabetes associated with the absence of IRS2 results in hypothalamic injury and to analyze the intracellular mechanisms involved. Only diabetic IRS2(-/-) mice showed increased cell death and activation of caspase-8 and -3 in the hypothalamus. Regulators of apoptosis such as FADD, Bcl-2, Bcl-xL and p53 were also increased, whereas p-IκB and c-FLIPL were decreased. This was accompanied by increased levels of Nox-4 and catalase, enzymes involved in oxidative stress. In summary, the hypothalamus of diabetic IRS2(-/-) mice showed an increase in oxidative stress and inflammatory markers that finally resulted in cell death via substantial activation of the extrinsic apoptotic pathway. Conversely, non-diabetic IRS2(-/-) mice did not show cell death in the hypothalamus, possibly owing to an increase in the levels of circulating IGF-I and in the enhanced hypothalamic IGF-IR phosphorylation that would lead to the stimulation of survival pathways. In conclusion, diabetes in IRS2-deficient male mice is associated with increased oxidative stress and apoptosis in the hypothalamus.

Differential insulin receptor substrate-1 (IRS1)-related modulation of neuropeptide Y and proopiomelanocortin expression in nondiabetic and diabetic IRS2-/- mice.

Insulin resistance and type 2 diabetes correlate with impaired leptin and insulin signaling. Insulin receptor substrate-2 deficient (IRS2(-/-)) mice are an accepted model for the exploration of alterations in these signaling pathways and their relationship with diabetes; however, disturbances in hypothalamic signaling and the effect on neuropeptides controlling food intake remain unclear. Our aim was to analyze how leptin and insulin signaling may differentially affect the expression of hypothalamic neuropeptides regulating food intake and hypothalamic inflammation in diabetic (D) and nondiabetic (ND) IRS2(-/-) mice. We analyzed the activation of leptin and insulin targets by Western blotting and their association by immunoprecipitation, as well as the mRNA levels of neuropeptide Y (NPY), proopiomelanocortin, and inflammatory markers by real-time PCR and colocalization of forkhead box protein O1 (FOXO1) and NPY by double immunohistochemistry in the hypothalamus. Serum leptin and insulin levels and hypothalamic Janus kinase 2 and signal transducer and activator of transcription factor 3 activation were increased in ND IRS2(-/-) mice. IRS1 levels and its association with Janus kinase 2 and p85 and protein kinase B activation were increased in ND IRS2(-/-). Increased FOXO1 positively correlated with NPY mRNA levels in D IRS2(-/-) mice, with FOXO1 showing mainly nuclear localization in D IRS2(-/-) and cytoplasmic in ND IRS2(-/-) mice. D IRS2(-/-) mice exhibited higher hypothalamic inflammation markers than ND IRS2(-/-) mice. In conclusion, differential activation of these pathways and changes in the expression of NPY and inflammation may exert a protective effect against hypothalamic deregulation of appetite, suggesting that manipulation of these targets could be of interest in the treatment of insulin resistance and type 2 diabetes.