Effects Of Zinc On Glucose Consumption And Its Mechanismin Insulin Resistance HepG2 Cells

ObjectiveTo investigate the effect of zinc on glucose metabolism and the key targets of PI3K-AKT signaling pathway in insulin resistance hepatocyte,to provide the experimental evidence of relation between zinc and insulin resistance.MethodsInsulin resistance model was established in human hepatoma cell line HepG2 cells with different concentrations of insulin(10-7,5×10-7,10-6,5×10-6,10-5,5×10-5 mol/L)treatment for 24 hours and the established model was evaluated.To establish insulin resistance model with the proper insulin concentration,then deal with different concentrations of zinc.Consumptions of glucose were tested by glucose oxidase reaction,glycogen contents were detected by glycogen kit after insulin and zinc action,to investigate the effect of zinc in normal cells and insulin resistance cells on glucose metabolism.The protein expression of phosphorylated insulin receptor ? subunit(pY-IR?)?phosphorylated protein kinase B(Akt/PKB)?phosphorylated glycogen synthase kinase(GSK)were tested by western blot.ResultsThe glucose consumptions of HepG2 cells decreased after different concentrations(10-7,5×10-7,10-6,5×10-6,10-5,5×10-5mol/L)of insulin for 24h while glucose consumptions was the least with 10-6mol/L insulin(P<0.05).Therefore concentration of 10-6mol/L was chosen to build resistance model.The effects of zinc on glucose uptake:The glucose consumptions of normal and insulin resistance HepG2 cells showed dose-response relationship with different concentrations(20?50?100?200?mol/L)of zinc.While the concentrations of zinc were 100?200?mol/L,it showed insulin-like effects,but this effects were significantly lower than the insulin.When the zinc action with insulin together,it can enhance the insulin action.20 and 50?mol/L zinc groups didn't increase the consumption of glucose in insulin resistance cells,but with the 50?mol/L of zinc and insulin's stimulate can increase the consumption of glucose in normal cells.The effects of zinc on glycogen contents:Without insulin stimulation,compared with normal cells and insulin resistance cells,glycogen content of the 100?200?mol/L zinc groups increased significantly.200?mol/L zinc action with insulin,showed more increased compared with insulin respectively and zinc respectively.The effects of zinc on key targets of PI3K-AKT signaling pathway:Insulin stimulation increased the IR??AKT Ser473?GSK 3?Ser9 phosphorylation level significantly in normal cells and the model cells but to a lesser extent,indicating decreased sensitivity to insulin stimulation in model cells.Protein expressions of pY-IR??pAKT Ser473?pGSK 3 ?Ser9 increased in 100?200?mol/L zinc treatment respectively groups.200?mol/L zinc action with insulin,showed more increased compared with insulin respectively and zinc respectively.The effects of zinc on key targets of PI3K-AKT signaling pathway after inhibit by wortmannin:Compared with non wortmannin groups,insulin stimulation increased the IRP?AKT Ser473?GSK 3?Ser9 phosphorylation level.After treated with wortmannin,the protein expressions of p AKT Ser473?pGSK 3?Ser9 were inhibited significantly.It showed that the effects of zinc on Akt and GSK were achieved through the PI3K-AKT signaling pathway.ConclusionsWe used a high concentration of insulin(10-6mol/L)to build HepG2 insulin resistance model cells with 24 hours treatment.100?200?mol/L zinc can promote glycogen synthesis of insulin resistant cells.Zinc showed a synergistic effect with insulin,which enhanced insulin action and improved glucose metabolism in resistance cells.100?200?mol/L zinc showed insulin-like effects.They can increased the protein expressions of pY-IR??p AKT Ser473?pGSK 3?Ser9.Therefore,this may be one of the molecular mechanisms of zinc to improve glucose metabolism in insulin resistance HepG2 model cells.The effects of zinc on AKT and GSK were achieved through the PI3K-AKT signaling pathway,rather than the direct effect on AKT and GSK.