A genistein-enriched diet neither improves skeletal muscle oxidative capacity nor prevents the transition towards advanced insulin resistance in ZDF rats.
ABSTRACT: Genistein, a natural food compound mainly present in soybeans, is considered a potent antioxidant and to improve glucose homeostasis. However, its mechanism of action remains poorly understood. Here, we analyzed whether genistein could antagonize the progression of the hyperinsulinemic normoglycemic state (pre-diabetes) toward full-blown T2DM in Zucker Diabetic Fatty (ZDF) rats by decreasing mitochondrial oxidative stress and improving skeletal muscle oxidative capacity. Rats were assigned to three groups: (1) lean control (CNTL), (2) fa/fa CNTL, and (3) fa/fa genistein (GEN). GEN animals were subjected to a 0.02% (w/w) genistein-enriched diet for 8 weeks, whereas CNTL rats received a standard diet. We show that genistein did not affect the overall response to a glucose challenge in ZDF rats. In fact, genistein may exacerbate glucose intolerance as fasting glucose levels were significantly higher in fa/fa GEN (17.6 ± 0.7 mM) compared with fa/fa CNTL animals (14.9 ± 1.4 mM). Oxidative stress, established by electron spin resonance (ESR) spectroscopy, carbonylated protein content and UCP3 levels, remained unchanged upon dietary genistein supplementation. Furthermore, respirometry measurements revealed no effects of genistein on mitochondrial function. In conclusion, dietary genistein supplementation did not improve glucose homeostasis, alleviate oxidative stress, or augment skeletal muscle metabolism in ZDF rats.
Project description:Hyperglycemia associated with inflammation and oxidative stress is a major cause of vascular dysfunction and cardiovascular disease in diabetes. Recent data reports that a selective sodium-glucose co-transporter 2 inhibitor (SGLT2i), empagliflozin (Jardiance®), ameliorates glucotoxicity via excretion of excess glucose in urine (glucosuria) and significantly improves cardiovascular mortality in type 2 diabetes mellitus (T2DM). The overarching hypothesis is that hyperglycemia and glucotoxicity are upstream of all other complications seen in diabetes. The aim of this study was to investigate effects of empagliflozin on glucotoxicity, β-cell function, inflammation, oxidative stress and endothelial dysfunction in Zucker diabetic fatty (ZDF) rats. Male ZDF rats were used as a model of T2DM (35 diabetic ZDF-Leprfa/fa and 16 ZDF-Lepr+/+ controls). Empagliflozin (10 and 30mg/kg/d) was administered via drinking water for 6 weeks. Treatment with empagliflozin restored glycemic control. Empagliflozin improved endothelial function (thoracic aorta) and reduced oxidative stress in the aorta and in blood of diabetic rats. Inflammation and glucotoxicity (AGE/RAGE signaling) were epigenetically prevented by SGLT2i treatment (ChIP). Linear regression analysis revealed a significant inverse correlation of endothelial function with HbA1c, whereas leukocyte-dependent oxidative burst and C-reactive protein (CRP) were positively correlated with HbA1c. Viability of hyperglycemic endothelial cells was pleiotropically improved by SGLT2i. Empagliflozin reduces glucotoxicity and thereby prevents the development of endothelial dysfunction, reduces oxidative stress and exhibits anti-inflammatory effects in ZDF rats, despite persisting hyperlipidemia and hyperinsulinemia. Our preclinical observations provide insights into the mechanisms by which empagliflozin reduces cardiovascular mortality in humans (EMPA-REG trial).
Project description:The aim of this study was to characterize the early alterations of the liver mitochondrial function in ZDF (fa/fa) rats that develop diabetes compared to that of their lean counterparts ZDF (fa/+). Liver mitochondrial function was examined in 11- and 14-week-old ZDF (fa/fa) and ZDF lean (fa/+) rats. Oxygen consumption, H2O2 release, calcium retention capacity (CRC), membrane potential, membrane fluidity, and fatty acid composition were analyzed. State 3 oxygen consumption with palmitoyl-carnitine increases between 11 and 14 weeks of age in lean but not in diabetic animals. This response was not seen with other substrates, suggesting that the use of fatty acids is impaired in diabetic rats. H2O2 release was lower in 14-week-old ZDF (fa/fa) rats as compared to ZDF lean (fa/+). These changes were not associated with differences in enzymatic activities of the respiratory complexes, suggesting regulatory mechanisms independent of their expression levels. Membrane fluidity and composition analyses show only slight effects linked to diabetes progression. The most salient feature was a reduction in CRC in the presence of CsA, an effect reflecting PTP dysregulation. Our data suggest few changes of mitochondrial function in ZDF fa/fa rats. At the age of 11 weeks, liver mitochondria have mainly a reduced effect of CsA on CRC.
Project description:Type 2 diabetes (T2D) is a complex metabolic disease, which involves a maintained hyperglycemia due to the development of an insulin resistance process. Among multiple risk factors, host intestinal microbiota has received increasing attention in T2D etiology and progression. In the present study, we have explored the effect of long-term supplementation with a non-dairy fermented food product (FFP) in Zucker Diabetic and Fatty (ZDF) rats T2D model. The supplementation with FFP induced an improvement in glucose homeostasis according to the results obtained from fasting blood glucose levels, glucose tolerance test, and pancreatic function. Importantly, a significantly reduced intestinal glucose absorption was found in the FFP-treated rats. Supplemented animals also showed a greater survival suggesting a better health status as a result of the FFP intake. Some dissimilarities have been observed in the gut microbiota population between control and FFP-treated rats, and interestingly a tendency for better cardiometabolic markers values was appreciated in this group. However, no significant differences were observed in body weight, body composition, or food intake between groups. These findings suggest that FFP induced gut microbiota modifications in ZDF rats that improved glucose metabolism and protected from T2D development.
Project description:Background:Early diagnosis of type 2 diabetes mellitus (T2DM) is still difficult. Screening of plasma biomarkers has great significance of optimizing diagnosis and predicting the complications of T2DM. Methods:We used a special diet, Purina #5008, to induce diabetes in Zucker leptin receptor gene-deficient rats (fa/fa) to establish Zucker diabetic fatty (ZDF) rats, simulating the early stage of T2DM. The differentially expressed proteins (DEP) and lipids (DEL), as potential biomarkers, were screened to compare the plasma expression levels in ZDF rats and their basic diet-fed wild-type controls (fa/+) by Tandem Mass Tags (TMT) and liquid chromatography-tandem mass spectrometry. Results:These two groups had different plasma proteins and lipids profiles consisting of 84 DEPs and, 179 DELs identified in the positive ion mode and 178 DELs in the negative ion mode, respectively. Enrichment analysis of these different indicators showed that oxidative stress, insulin resistance and metabolic disorders of glycan and lipid played an important role in generating the difference. Some markers can be used as candidate biomarkers in prediction and treatments of T2DM, such as ceruloplasmin, apolipoprotein C-I, apolipoprotein C-II and apolipoprotein C-IV. Conclusion:These plasma differences help to optimize the diagnosis and predict the complications of T2DM, although this remains to be verified in the crowd. Trace elements related-metalloproteins, such as ceruloplasmin, and lipid metabolism and transport-related apolipoprotein C are expected to be candidate biomarkers of T2DM and should be given more attention.
Project description:Gene expression changes have been associated with type 2 diabetes mellitus (T2DM); however, the alterations are not fully understood. We investigated the effects of anti-diabetic drugs on gene expression in Zucker diabetic fatty (ZDF) rats using oligonucleotide microarray technology to identify gene expression changes occurring in T2DM. Global gene expression in the pancreas, adipose tissue, skeletal muscle, and liver was profiled from Zucker lean control (ZLC) and anti-diabetic drug treated ZDF rats compared with those in ZDF rats. We showed that anti-diabetic drugs regulate the expression of a large number of genes. We provided a more integrated view of the diabetic changes by examining the gene expression networks. The resulting sub-networks allowed us to identify several biological processes that were significantly enriched by the anti-diabetic drug treatment, including oxidative phosphorylation (OXPHOS), systemic lupus erythematous, and the chemokine signaling pathway. Among them, we found that white adipose tissue from ZDF rats showed decreased expression of a set of OXPHOS genes that were normalized by rosiglitazone treatment accompanied by rescued blood glucose levels. In conclusion, we suggest that alterations in OXPHOS gene expression in white adipose tissue may play a role in the pathogenesis and drug mediated recovery of T2DM through a comprehensive gene expression network study after multi-drug treatment of ZDF rats.
Project description:Brown adipose tissue (BAT) is an attractive therapeutic target to combat diabetes and obesity due to its ability to increase glucose expenditure. In a genetic rat model (ZDF fa/fa) of type-2 diabetes and obesity, we aimed to investigate glucose utilization of BAT by 18F-FDG PET imaging. Male Zucker diabetic fatty (ZDF) and Male Zucker lean (ZL) control rats were studied at 13 weeks. Three weeks prior to imaging, ZDF rats were randomized into a no-restriction (ZDF-ND) and a mild calorie restriction (ZDF-CR) group. Dynamic 18F-FDG PET using a dedicated small animal PET system was performed under hyperinsulinemic-euglycemic clamp. 18F-FDG PET identified intense inter-scapular BAT glucose uptake in all ZL control rats, while no focally increased 18F-FDG uptake was detected in all ZDF-ND rats. Mild but significant improved BAT tracer uptake was identified after calorie restriction in diabetic rats (ZDF-CR). The weight of BAT tissue and fat deposits were significantly increased in ZDF-CR and ZDF-ND rats as compared to ZL controls, while UCP-1 and mitochondrial concentrations were significantly decreased. Whitening and severely impaired insulin-stimulated glucose uptake in BAT was confirmed in a rat model of type-2 diabetes. Additionally, calorie restriction partially restored the impaired BAT glucose uptake.
Project description:The ZDF rat, with spontaneous homozygous mutation of the leptin receptor gene (fa/fa), is one of the widely used animal model for studying the human type 2 diabetes mellitus (T2DM). Male ZDF rats have the symptoms of obesity and insulin resistance at a young age, accompanying with impaired islet function. However, their hepatic pathogenesis is still unclear. Based on the successive observations and the transcriptomic analyses of the liver tissue at 22 weeks old, we detected the typical clinical indications of T2DM, severe hepatic metabolic remodeling and the inflammatory liver injury in the ZDF rats. The integrin linked kinase signaling, as well as the endoplasmic reticulum stress and its downstream p38 MAPK signaling, seemed to play crucial roles in it. We have proved the ZDF rats could better simulate the pathogenesis of the human T2DM associated nonalcoholic fatty liver disease (NAFLD), and provided targets and reference for future T2DM studies. Overall design: Examined two samples and each has two repeats
Project description:To correlate changes between VEGF expression with systemic and retinal oxidative stress and inflammation in rodent models of obesity induced insulin resistance and diabetes.Retinal VEGF mRNA and protein levels were assessed by RT-PCR and VEGF ELISA, respectively. Urinary 8-hydroxydeoxyguanosine (8-OHdG), blood levels of C-reactive protein (CRP), malondialdehyde (MDA), and CD11b/c positive cell ratio were used as systemic inflammatory markers. Retinal expression of Nox2, Nox4, and p47phox mRNA levels were measured as oxidative stress markers. TNF-?, inter-cellular adhesion molecule-1 (ICAM-1), IL1?, and activation of nuclear factor ?B (NF-?B) were used as retinal inflammatory markers.Retinal VEGF mRNA and protein expression increased in Zucker diabetic fatty (ZDF(fa/fa)) rats and streptozotosin (STZ) induced diabetic Sprague-Dawley rats, after two months of disease, but not in Zucker fatty (ZF) rats. Systemic markers of oxidative stress and inflammation were elevated in insulin resistant and diabetic rats. Some oxidative stress and inflammatory markers (TNF-?, IL-6, ICAM-1, and IL1-?) were upregulated in the retina of ZDF(fa/fa) and STZ diabetic rats after 4 months of disease. In contrast, activation of NF-?B in the retina was observed in high fat fed nondiabetic and diabetic cis-NF-?B(EGFP) mice, ZF, ZDF(fa/fa), and STZ-induced diabetic rats.Only persistent hyperglycemia and diabetes increased retinal VEGF expression. Some markers of inflammation and oxidative stress were elevated in the retina and systemic circulation of obese and insulin resistant rodents with and without diabetes. Induction of VEGF and its associated retinal pathologies by diabetes requires chronic hyperglycemia and factors in addition to inflammation and oxidative stress.
Project description:BACKGROUND: Metabolic syndrome (coexisting visceral obesity, dyslipidemia, hyperglycemia, and hypertension) is a prominent risk factor for cardiovascular morbidity and mortality, however, its effect on cardiac gene expression pattern is unclear. Therefore, we examined the possible alterations in cardiac gene expression pattern in male Zucker Diabetic Fatty (ZDF) rats, a model of metabolic syndrome. METHODS: Fasting blood glucose, serum insulin, cholesterol and triglyceride levels were measured at 6, 16, and 25 wk of age in male ZDF and lean control rats. Oral glucose tolerance test was performed at 16 and 25 wk of age. At week 25, total RNA was isolated from the myocardium and assayed by rat oligonucleotide microarray for 14921 genes. Expression of selected genes was confirmed by qRT-PCR. RESULTS: Fasting blood glucose, serum insulin, cholesterol and triglyceride levels were significantly increased, glucose tolerance and insulin sensitivity were impaired in ZDF rats compared to leans. In hearts of ZDF rats, 36 genes showed significant up-regulation and 49 genes showed down-regulation as compared to lean controls. Genes with significantly altered expression in the heart due to metabolic syndrome includes functional clusters of metabolism (e.g. 3-hydroxy-3-methylglutaryl-Coenzyme A synthase 2; argininosuccinate synthetase; 2-amino-3-ketobutyrate-coenzyme A ligase), structural proteins (e.g. myosin IXA; aggrecan1), signal transduction (e.g. activating transcription factor 3; phospholipase A2; insulin responsive sequence DNA binding protein-1) stress response (e.g. heat shock 70kD protein 1A; heat shock protein 60; glutathione S-transferase Yc2 subunit), ion channels and receptors (e.g. ATPase, (Na+)/K+ transporting, beta 4 polypeptide; ATPase, H+/K+ transporting, nongastric, alpha polypeptide). Moreover some other genes with no definite functional clusters were also changed such as e.g. S100 calcium binding protein A3; ubiquitin carboxy-terminal hydrolase L1; interleukin 18. Gene ontology analysis revealed several significantly enriched functional inter-relationships between genes influenced by metabolic syndrome. CONCLUSIONS: Metabolic syndrome significantly alters cardiac gene expression profile which may be involved in development of cardiac pathologies in the presence of metabolic syndrome.
Project description:Antagonists of angiotensin AT(1) receptors elicit beneficial vascular effects in diabetes mellitus. We hypothesized that diabetes induces sustained availability of AT(1) receptors, causing enhanced arterial constriction to angiotensin II.To assess functional availability of AT(1) receptors, constrictions to successive applications of angiotensin II were measured in isolated skeletal muscle resistance arteries (?150 µm) of Zucker diabetic fatty (ZDF) rats and of their controls (+/Fa), exposed acutely to high glucose concentrations (HG, 25 mM, 1 h). AT(1) receptors on cell membrane surface were measured by immunofluorescence.Angiotensin II-induced constrictions to first applications were greater in arteries of ZDF rats (maximum: 82 ± 3% original diameter) than in those from +/Fa rats (61 ± 5%). Constrictions to repeated angiotensin II administration were decreased in +/Fa arteries (20 ± 6%), but were maintained in ZDF arteries (67 ± 4%) and in +/Fa arteries vessels exposed to HG (65 ± 6%). In ZDF arteries and in HG-exposed +/Fa arteries, Rho-kinase activities were enhanced. The Rho-kinase inhibitor, Y27632 inhibited sustained constrictions to angiotensin II in ZDF arteries and in +/Fa arteries exposed to HG. Levels of surface AT(1) receptors on cultured vascular smooth muscle cells (VSMCs) were decreased by angiotensin II but were maintained in VSMCs exposed to HG. In VSMCs exposed to HG and treated with Y27632, angiotensin II decreased surface AT(1) receptors.In diabetes, elevated glucose concentrations activate Rho-kinase which inhibits internalization or facilitates recycling of AT(1) receptors, leading to increased functional availability of AT(1) receptors and sustained angiotensin II-induced arterial constriction.