Effects Of Long-term Supranutritional Supplementation With Selenate In Type2Diabetic Db/db Mice

Diabetes is one of the top three diseases seriously threatening worldwide human health, and its prevalence has grown markedly year by year. The pathogenesis, diagnosis and treatment of diabetes are therefore the focus of intense worldwide research. Selenium is one of the necessary trace nutrient elements for human and animals. Many studies have showed that selenium has an insulin-mimetic property and acts as an antioxidant nutrient, can be used as dietary supplements for the prevention or treatment of diabetes. However, in recent decades, findings from large-scale human studies and animal experiments have suggested that there are two sides in the role of selenium plays on diabetes. On the one hand, selenium could decrease the high concentration of blood glucose and enhance insulin sensitivity; on the other hand, both low and high selenium status could increase the risk of type2diabetes, and long-term selenium supplementations actually increase the incidence of insulin resistance and type2diabetes.In order to investigate the possible role and mechanisms of selenium as a "double-edged sword" in diabetes, our studies were designed to determine the effects of a supranutritional dose of selenate daily oral supplementation in type2diabetic db/db mice for two months. The fasting blood glucose concentrations were monitored during the supplement stage, and then histopathological observation, cDNA microarray profiling and the two-dimensional differential gel electrophoresis (2D-DIGE) coupled with mass spectrometry (MS) were used to determine the changes of pathologies and mRNA, protein expression profiling in pancreatic and liver tissues after selenate simultaneously decresed the hyperglycemia in db/db mice, and suppose the possible mechanisms of selenate. We have made systematical and comprehensive analysis on the therapeutic efficacy and its possible side effects of oral selenate supplementation for type2diabetes. The main findings are summarized as follows: 1. Firstly we determined a moderate dose of0.8mg Na2SeO4/kg BW (about18fold of the recommended dietary amount) for supplementation by a dosage gradient study. Fasting blood glucose levels increased continuously in db/db mice administered placebo (DMCtrl) but decreased gradually in selenate-supplemented db/db mice (DMSe) and almost approached normal values when the experiment ended. The plasma insulin concentration of DMSe mice were increased nearly to double and impaired glucose tolerance decreased also in DMSe mice because of the suppression of hyperglycemic growth. However, the serum lipid parameters of total cholesterol, high density lipoprotein and low density lipoprotein significantly increased in DMSe mice compared with that in DMCtrl mice, indicating aggravating hyperlipidemia.2. The islets in diabetic db/db mice increased in size by a compensatory mechanism, with occasional mild vacuolation and stromal fibrous proliferation, showing cell damage and decreased insulin secretion. After selenate treatment, the area of islet enlarged more, as well as the number of β cell, and the insulin secretion increased obviously by30%. suggesting that selenate suppressed the cell damage and β cell functional destruction caused by compensatory secrete excess. Moreover, by cDNA microarray and real-time fluorescent quantitative polymerase chain reaction analysis, a wide range of differentially expressed genes either closely associated with key pancreatic β-cell functions or the development of diabetes were detected to have benefit change in pancreatic tissue of DMSe mice compared with that of DMCtrl mice, such as Pax6、Neurodl、Ins1、Ins2、Scg3、Scgn、Bace2、Acvrlc、G6pc2、Fbp2、 Abcc8、Slc2a5, indicating increased insulin secretion and protective P-cell function integrity. It is the first time to show that selenate also could stimulate the key genes and proteins to express in islets, promote insulin synthesis and secretion and protect islet β-cell structure and funtion integrity.3. Unlike expected, hepatopathy observed in the DMSe mice was more severe than that in the DMCtrl mice and was accompanied by increased hepatocyte lipid vacuolation and fat accumulation. Hepatic cord congestion, inflammatory cell infiltration and occasional necrosis and apoptosis were observed in DMSe, and hepatic glycogen storage capacity was reduced accompanied by decreased serum glucose. cDNA microarray analysis of the liver samples showed that there were582transcripts had changed by a factor of≥1.5in expression levels responded to selenate supplementation, with301upregulated and281downregulated. Functional annotation and pathway analysis showed that a large part of these differentially expressed genes with redox activity is involved in regulating lipid metabolism, such as those involved in fatty acid synthesis through arachidonic acid metabolism and the PPARa signaling pathway. The other genes differentially expressed are mainly associated with detoxification, inflammatory responses, and cell cycle regulation. Proteomic analysis of liver using2D-DIGE and MS identified five proteins with slightly but significantly expressed difference, which involve in carbohydrates energy metabolism and endoplasmic reticulum calcium flow regulation, indicating promotion of hepatic fat accumulation and fibrosis by selenate supplementation. In addition, the hepatic total antioxidant defense capacity also performed a decreasing trend.In summary, our findings suggest that supranutritional supplementation (0.8mg/kg BW) of db/db mice with selenate does not improve or prevent the symptoms of type2diabetes, although hyperglycemia and impaired glucose tolerance decreased by stimulating key genes and proteins expressed in pancreatic islets and increasing β-cell number and insulin secretion, enhancing islet function in some degree; however, long-term excess secretion by islet β-cell and hyperinsulinemia would adversely affect functional and structural integrity of pancreatic islets and liver tissue, and eventually led to the antioxidant defense capacity reduced and liver fatty degeneration exacerbated, even non-alcoholic fatty liver complications development.