| In recent years, the metabolic syndrome represented by the obesity and type 2 diabetes increased sharply in China, China has become the first power of diabetes in the world. Insulin resistance is the main pathogenesis of the metabolic syndrome,therefore, the effective intervention insulin resistance has great significance.Hippocrates who is the father of western medicine raised in the BC "let food be your drugs, drugs should be your food" with Chinese traditional medicine "medicine food homology", explain the dialectic relationship of food and medicine to explore food intervention insulin resistance research laid the foundation. Barley is planting in the first four crops in China. Barley is one of the full nutrition foods, which provides health needs for us by having the necessary nutrients. ?-glucan and polyphenol of barley possess antioxidant activity, regulating lipid metabolism and glucose homeostasis and improving insulin resistance. But, barley fermentations showed significant potential in improvement and design of the nutritional quality and health effects of foods and ingredients. Therefore, the way can provide the efficient utilization of barley resources in new ways.In this paper, barley was used as raw materials, Lactobacillus plantarum dy-1 fermented barley preparing fermented barley extract (LFBE), and we used insulin resistant model of HepG2 cell and obese rat model to study the effect and mechanism of LFBE on insulin resistance. The main contents and conclusions are presented as the following:1. The content of main components of RBE were compared and analyzed. The results showed that:(1) Raw barley extract(RBE) contained 13.94 % crude protein, 8.96 mg/g total phenols, 4.83% (P-glucan and 64.94% total sugar. Compared to the RBE, the protein in LFBE significant increased to 34.94%, the total phenols content of LFBE increased to 13.61 mg/g, the concentration of ?-glucan significantly increased to 13.44 % and the sugar significant decreased to 35.44%. The phenolics compounds of LFBE increased significantly, the gallic acid, coumaric acid, ferulic acid and vanillic acid content were significantly higher than that of RBE.2. The obese rat model was selected to study the effect of LFBE on insulin resistance. The results showed that:(1) LFBE could reduce the body weight, Lee's index and the fat content of obese rats. Meanwhile, the serum insulin levels was reduced and the glucose tolerance was improved when treated with LFBE. So, LFBE could lower rate of increase of obese rats' body weights, reverse the disorders of glucose metabolism and ameliorate the symptoms of insulin resistance in obese rats.(2) LFBE could reduce the serum and liver TG and TC contents of obese rats.Moreover, the steatosis of liver was inhibited in obese rats after treatment with LFBE.Therefore, LFBE could improve lipid metabolism in obese rats and prevent the formation of fatty liver.(3) LFBE might increase the antioxidant capacity and inhibit the NF-?B and the JNK/p38 mediated MAPK signaling pathway in obese rats, and then ameliorate the symptoms of insulin resistance.(4) There were similarities and differences between LFBE and pioglitazone on the effects of insulin resistance in obese rats. LFBE and pioglitazone could both ameliorate the symptoms of insulin resistance and up-regulate the protein expression of I?B-? and down-regulate the protein expression of NF-?B in obese rats; while pioglitazone showed no significant effect on p-JNK and p-p38 signaling pathway body weight and the body fat content in obese rats, but LFBE could inhibit p-JNK and p-p38 signaling pathway, lower rate of increase of obese rats' body weights and body fat content.3. The palmitic acid induced insulin resistant model of HepG2 cells was selected to study the main active component of intervention insulin resistance in LFBE. The results showed that: Protein, ?-glucan, polyphenols, ferulic acid and vanillic acid in LFBE can increase glucose consumption in certain degree, vanillic acid showed the higher enhancement activity of glucose consumption in HepG2 cells. LFBE and vanillic acid can significantly reduce lipid content, the levels of TNF-a, IL-1? and IL-6 in HepG2 cells. Its mechanism is associated with LFBE and vanillic acid can inhibit phosphorylation of JNK and p38, regulate the MAPK signaling pathways.4. Using microRNA microarray, qRT-PCR method and bioinformatics technology, study LFBE effects on microRNA expression in liver of obesity rats,explore its mechanism. The results showed that: A total of 18 miRNAs were detected differentially expressed between HFD group and NC group by the miRNA microarray.A total of 15 miRNAs were detected differentially expressed between HFD group and LFBE group by the miRNA microarray. The analysis with KEGG database found that targets of miRNAs were associated with insulin resistance signaling pathways, mainly including MAPK signaling pathway and Insulin secretion. Speculate that LFBE can regulat the expression of miR-34a-5p and miR-212-3p to intervention of insulin resistance.5. Using luciferase reporter gene assay, in vitro transfection experiment and the palmitic acid induced insulin resistant model of HepG2 cells to Validate the molecular mechanisms of LFBE and vanillic acid on insulin resistance by regulating expression of miR-212. The results showed that:(1) Luciferase reporter gene assay showed that HepG2 cells were transfected with luciferase reporter vector containing WT-DUSP9 3'-UTR, cell fluorescence luciferase activity was significantly decreased, significantly lower than the control group; HepG2 cells were transfected with luciferase reporter vector containing MUT-DUSP9 3'-UTR, cell fluorescence luciferase activity was no significant effect,show that DUSP9 is target gene of miR-212.(2) In the HepG2 cells were transfected miR-212 mimics, DUSP9 mRNA and protein levels were significantly decreased; but in the HepG2 cells were transfected miR-212 inhibitor, DUSP9 mRNA and protein levels were significantly increased,show that miR-212 can negative regulat the expression of DUSP9 mRNA and protein.(3) miR-212 mimics could reduce glucose consumption, increased lipid accumulation and decreased DUSP9 mRNA and protein levels in HepG2 cells;miR-212 inhibitor showed the enhancement activity of glucose consumption and inhibition of lipid accumulation and increased DUSP9 mRNA and protein levels in HepG2 cells.(4) LFBE and vanillic acid could ameliorate insulin resistance by down-regulate the expression of miR-212 and up-regulate the expression of DUSP9 in HepG2 cells. |
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