EZRMI-13K MilliporeRat/Mouse Insulin ELISA

The AIN-93 diet was proposed by the American Institute of Nutrition with the objective of standardising studies in experimental nutrition. Our objective was to analyze the effects of AIN-93 diet after myocardial infarction in rats.

Diabetes mellitus causes cerebral microvasculature deterioration and cognitive decline. The specialized endothelial cells of cerebral microvasculature comprise the blood-brain barrier, and the pericytes (PC) that are in immediate contact with these endothelial cells are vital for blood-brain barrier integrity. In diabetes, increased mitochondrial oxidative stress is implicated as a mechanism for hyperglycemia-induced PC loss as a prerequisite leading to blood-brain barrier disruption. Mitochondrial carbonic anhydrases (CA) regulate the oxidative metabolism of glucose and thus play an important role in the generation of reactive oxygen species and oxidative stress. We hypothesize that the inhibition of mitochondrial CA would reduce mitochondrial oxidative stress, rescue cerebral PC loss caused by diabetes-induced oxidative stress, and preserve blood-brain barrier integrity. We studied the effects of pharmacological inhibition of mitochondrial CA activity on streptozotocin-diabetes-induced oxidative stress and PC loss in the mouse brain. At 3 wk of diabetes, there was significant oxidative stress; the levels of reduced glutathione were lower and those of 3-nitrotyrosine, 4-hydroxy-2-trans-nonenal, and superoxide dismutase were higher. Treatment of diabetic mice with topiramate, a potent mitochondrial CA inhibitor, prevented the oxidative stress caused by 3 wk of diabetes. A significant decline in cerebral PC numbers, at 12 wk of diabetes, was also rescued by topiramate treatment. These results provide the first evidence that inhibition of mitochondrial CA activity reduces diabetes-induced oxidative stress in the mouse brain and rescues cerebral PC dropout. Thus, mitochondrial CA may provide a new therapeutic target for oxidative stress related illnesses of the central nervous system.

Hypothalamic inflammation has been demonstrated to be an important mechanism in the pathogenesis of obesity-induced type 2 diabetes mellitus. Feeding pregnant and lactating rodents a diet rich in saturated fatty acids has consistently been shown to predispose the offspring for the development of obesity and impaired glucose metabolism. However, hypothalamic inflammation in the offspring has not been addressed as a potential underlying mechanism. In this study, virgin female C57BL/6 mice received high-fat feeding starting at conception until weaning of the offspring at postnatal d 21. The offspring developed increased body weight, body fat content, and serum leptin concentrations during the nursing period. Analysis of hypothalamic tissue of the offspring at postnatal d 21 showed up-regulation of several members of the toll-like receptor 4 signaling cascade and subsequent activation of c-Jun N-terminal kinase 1 and I?B kinase-? inflammatory pathways. Interestingly, glucose tolerance testing in the offspring revealed signs of impaired glucose tolerance along with increased hepatic expression of the key gluconeogenic enzyme phosphoenolpyruvate carboxykinase. In addition, significantly increased hepatic and pancreatic PGC1? expression suggests a role for sympathetic innervation in mediating the effects of hypothalamic inflammation to the periphery. Taken together, our data indicate an important role for hypothalamic inflammation in the early pathogenesis of glucose intolerance after maternal perinatal high-fat feeding.

Fenretinide is a synthetic retinoid that is being tested in clinical trials for the treatment of breast cancer and insulin resistance, but its mechanism of action has been elusive. Recent in vitro data indicate that fenretinide inhibits dihydroceramide desaturase, an enzyme involved in the biosynthesis of lipotoxic ceramides that antagonize insulin action. Because of this finding, we assessed whether fenretinide could improve insulin sensitivity and glucose homeostasis in vitro and in vivo by controlling ceramide production. The effect of fenretinide on insulin action and the cellular lipidome was assessed in a number of lipid-challenged models including cultured myotubes and isolated muscles strips incubated with exogenous fatty acids and mice fed a high-fat diet. Insulin action was evaluated in the various models by measuring glucose uptake or disposal and the activation of Akt/PKB, a serine/threonine kinase that is obligate for insulin-stimulated anabolism. The effects of fenretinide on cellular lipid levels were assessed by LC-MS/MS. Fenretinide negated lipid-induced insulin resistance in each of the model systems assayed. Simultaneously, the drug depleted cells of ceramide, while promoting the accumulation of the precursor dihydroceramide, a substrate for the reaction catalyzed by Des1. These data suggest that fenretinide improves insulin sensitivity, at least in part, by inhibiting Des1 and suggest that therapeutics targeting this enzyme may be a viable therapeutic means for normalizing glucose homeostasis in the overweight and diabetic.

Non-alcoholic fatty liver disease (NAFLD) is one of the most common manifestations of chronic liver disease worldwide. The aim of the present study was to assess the effect of resveratrol on liver fat accumulation, as well as on the activity of those enzymes involved in lipogenesis and fatty acid oxidation in fa/fa Zucker rats. A total of thirty rats were assigned to three experimental groups and orally treated with resveratrol for 6 weeks, or without resveratrol (C: control group; RSV15 group: 15 mg/kg body weight per d; RSV45 group: 45 mg/kg body weight per d). Liver histological analysis was performed by microscopy. Levels of hepatic carnitine palmitoyltransferase-Ia (CPT-Ia), acyl-coenzyme A oxidase (ACO), fatty acid synthase, glucose-6-phosphate dehydrogenase and malic enzyme were assessed by spectrophotometry, and acetyl-CoA carboxylase was assessed by radiometry. Commercial kits were used to determine serum TAG, NEFA, total HDL and non-HDL-cholesterol, glycerol, ketonic bodies, glucose, insulin, adiponectin, aspartate aminotransferase (AST), alanine aminotransferase (ALT) and alkaline phosphatase (ALP), hepatic TAG, thiobarbituric acid reactive substrates, GSH (GSSG) and superoxide dismutase. Resveratrol reduced liver weight and TAG content. It did not modify the activity of lipogenic enzymes but it did increase CPT-Ia and ACO activities. NEFA and ALP were reduced in both resveratrol-treated groups. AST/GOT was reduced only by the lowest dose. ALT/GPT, TAG and adiponectin remained unchanged. Resveratrol reduced liver oxidative stress. This study demonstrates that resveratrol can protect the liver from NAFLD by reducing fatty acid availability. Moreover, resveratrol also protects liver from oxidative stress.

The serine/threonine kinase Akt is frequently activated in human cancers and is considered an attractive therapeutic target. However, the relative contributions of the different Akt isoforms to tumorigenesis, and the effect of their deficiencies on cancer development are not well understood. We had previously shown that Akt1 deficiency is sufficient to markedly reduce the incidence of tumors in Pten(+/-) mice. Particularly, Akt1 deficiency inhibits endometrial carcinoma and prostate neoplasia in Pten(+/-) mice. Here, we analyzed the effect of Akt2 deficiency on the incidence of tumors in Pten(+/-) mice. Relative to Akt1, Akt2 deficiency had little-to-no effect on the incidence of prostate neoplasia, endometrial carcinoma, intestinal polyps and adrenal lesions in Pten(+/-) mice. However, Akt2 deficiency significantly decreased the incidence of thyroid tumors in Pten(+/-), which correlates with the relatively high level of Akt2 expression in the thyroid. Thus, unlike Akt1 deletion, Akt2 deletion is not sufficient to markedly inhibit tumorigenesis in Pten(+/-) mice in most tested tissues. The relatively small effect of Akt2 deletion on the inhibition of tumorigenesis in Pten(+/-) mice could be explained, in part, by an insufficient decrease in total Akt activity, due to the relatively lower Akt2 versus Akt1 expression, and relatively high blood insulin levels in Pten(+/-)Akt2(-/-) mice. The relatively high blood insulin levels in Pten(+/-)Akt2(-/-) mice may elevate the activity of Akt1, and possibly Akt3, thus, limiting the reduction of total Akt activity and preventing this activity from dropping to a threshold level required to inhibit tumorigenesis.

Accelerated atherosclerosis is the leading cause of death in type 1 diabetes, but the mechanism of type 1 diabetes-accelerated atherosclerosis is not well understood, in part due to the lack of a good animal model for the long-term studies required. In an attempt to create a model for studying diabetic macrovascular disease, we have generated type 1 diabetic Akita mice lacking the low density lipoprotein receptor (Ins2(Akita)Ldlr⁻/⁻). Ins2(Akita)Ldlr⁻/⁻ mice were severely hyperglycemic with impaired glucose tolerance. Compared with Ldlr⁻/⁻ mice, 20-week-old Ins2(Akita)Ldlr⁻/⁻ mice fed a 0.02% cholesterol AIN76a diet showed increased plasma triglyceride and cholesterol levels, and increased aortic root cross-sectional atherosclerotic lesion area [224% (P < 0.001) in males and 30% (P < 0.05) in females]. Microarray and quantitative PCR analyses of livers from Ins2(Akita)Ldlr⁻/⁻ mice revealed altered expression of lipid homeostatic genes, including sterol-regulatory element binding protein (Srebp)1, liver X receptor (Lxr)α, Abca1, Cyp7b1, Cyp27a1, and Lpl, along with increased expression of pro-inflammatory cytokine genes, including interleukin (Il)1α, Il1β, Il2, tumor necrosis factor (Tnf)α, and Mcp1. Immunofluorescence staining showed that the expression levels of Mcp1, Tnfα, and Il1β were also increased in the atherosclerotic lesions and artery walls of Ins2(Akita)Ldlr⁻/⁻ mice. Thus, the Ins2(Akita)Ldlr⁻/⁻ mouse appears to be a promising model for mechanistic studies of type 1 diabetes-accelerated atherosclerosis.

Intrauterine growth restriction (IUGR), along with postnatal growth trajectory, is closely linked with metabolic diseases and obesity at adulthood. The present study reports the time-dependent metabolomic response of male offspring of rat dams exposed to maternal adequate protein diet during pregnancy and lactation (CC) or protein deprivation during pregnancy only (IUGR with rapid catch-up growth, RC) or through pregnancy and lactation (IUGR with slow postnatal growth, RR). Plasma LC-HRMS metabolomic fingerprints for 8 male rats per group, combined with multivariate statistical analysis (PLS-DA and HCA), were used to study the impact of IUGR and postnatal growth velocity on the offspring metabolism in early life (until weaning) and once they reached adulthood (8 months). Compared with CC rats, RR pups had clear-cut alterations in plasma metabolome during suckling, but none at adulthood; in contrast, in RC pups, alterations in metabolome were minimal in early life but more pronounced in the long run. In particular, our results pinpoint transient alterations in proline, arginine, and histidine in RR rats, compared to CC rats, and persistent differences in tyrosine and carnitine, compared to RC rats at adulthood. These findings suggest that the long-term deregulation in feeding behavior and fatty acid metabolism in IUGR rats depends on postnatal growth velocity.

Dang Gui Bu Xue Tang (DBT), a Chinese medicinal decoction contains Radix Angelicae sinensis (Danggui) and Radix Astragali (Huangqi) at a ratio of 1 : 5, is used commonly for treating women's ailments. This study was conducted to explore the effects of this preparation on insulin resistance in rats fed with 6-week diet containing 60% fructose. Similar to the action of rosiglitazone (4 mg kg(-1) per day by an oral administration), repeated oral administration of DBT (2.5 g kg(-1) per day) for 14 days was found to significantly alleviate the hyperglycemia but made no influence on plasma lipid profiles nor weight gain in fructose chow-fed rats. Also, the higher degree of insulin resistance as measured by homeostasis model assessment of basal insulin resistance in fructose chow-fed rats was significantly decreased by repeated DBT treatment. DBT displays the characteristic of rosiglitazone by increasing the whole-body insulin sensitivity in fructose chow-fed rats after 2-week treatment, as evidenced by the marked elevation of composite whole-body insulin sensitivity index during the oral glucose tolerance test. DBT improves insulin sensitivity through increased post-receptor insulin signaling mediated by enhancements in insulin receptor substrate-1-associated phosphatidylinositol 3-kinase step and glucose transporter subtype 4 translocation in soleus muscles of animals exhibiting insulin resistance. DBT is therefore proposed as potentially useful adjuvant therapy for patients with insulin resistance and/or the patients who wish to increase insulin sensitivity.

The Akt signalling pathway plays vital roles in controlling cellular responses to insulin as well as in proliferation and survival. Inhibition of Akt signalling leads to insulin resistance and type 2 diabetes, whereas hyperactivation of Akt promotes tumorigenesis. In this study, we investigate how modest changes in the activity of the Akt signalling pathway, to an extent that might be achieved by drug treatment, would impact on insulin resistance and tumorigenesis. Using insulin-resistant PDK1(K465E/K465E) PH domain knock-in mice, we found that introducing the PTEN(+/-) mutation to slightly stimulate Akt restored normal insulin sensitivity. Introducing the PDK1(K465E/K465E) PH domain knock-in mutation into cancer-prone PTEN(+/-) mice, lowered Akt activity only by about 50%, but led to a delay in tumour onset of ∼4 months in a broad range of tumours. This was also accompanied by slower growth of B cell follicular lymphomas, as monitored by magnetic resonance imaging. Our findings imply that signal transduction inhibitors that lead to a modest reduction in Akt activity would not only delay onset of tumours possessing elevated phosphoinositide 3-kinase pathway activity but would also reduce the growth rate of developed tumours.