Effect Of Valsartan On P38MAPK And TGF-β1 Of High Glucose Cultured Rat Masangium Cells

Objective:Explore the effect of Valsartan on proliferation, oxidative stress and interaction with p38 mitogen-activated protein kinase (p38MAPK) and transforming growth factor-β1 (TGF-β1) Clarify the kidney protection mechanisms of Vasartan on rat glomerular mesangial cells (HBZY-1) cultured in high glucose.Method:1.Cultured the HBZY-1 cells in 37℃,5%CO2 box. The culture medium contained 10%fetal bovine serum and low glucose based DMEM medium. The first 3 to 5 generation were used in the experiment.2.The HBZY-1 cells in vitro were divided into normal group, high glucose group,mannitol group,high glucose+valsartan group,high glucose+ p38MAPK blocker group.3.Inverted the cells by microscope after cultured for 24 hours,48hours. Observe the cytological morphology and measure the cell viability using tetrazolium blue method (MTT).4.Measured the superoxide dismutase (SOD),nitrogen monoxidum (NO) and malondialdehyde (MDA) in the content of supernatant of these groups using biochemistry method.5.Detected the mRNA level expression of phosphorylated p38MAPK and TGF-β1 in these groups using the method of RT-PCR.6.Detected the protein level expression of phosphorylated p38MAPK and TGF-β1 in these groups using the method of immunocyto-chemistry and Western blot. Results:1.Morphology in the groups:After cultured for 24 hours,the N group cells are normal in morphology,the structure is clear.The H group cells,most are disorganized and flat obviously.The M group cells are similar to the N group cells in morphology.The cell number slightly increased,the cell bodies are slightly stronger.The V group and the B group cells are similar in morphological changes,the cell gaps are larger,the cell structures are not very clear,the cells are flat and show an apoptosis state.After cultured for 48 hours,the H group cells have all been suspended from the sides of the bottle and float in the cultured supernatant.2.The cell proliferation effect:Compared with the N group,the H group cells increased in proliferation,the OD value and the cell survival rate increase.Compared with the H group,the proliferation of the V group and the B group cells was inhibited.the OD value and the cell survival rate reduced (P<0.01).3.The oxidative stress effect:Compared with the N group,the SOD activity in the supernatant of the H group cells decreased,the MDA content increased and the NO content decreased significantly (P<0.01).Compared with the H group the SOD activity in the supernatant of the V group cells increased,the MDA content decreased and the NO content increased significantly (P<0.01).Compared with the H group the SOD activity in the supernatant of the V group cells increased,the MDA content decreased (P<0.01) and the NO content increased significantly(P<0.05).4.The mRNA expression of p38MAPK and TGFβ1:Compared with the N group,the mRNA expression of p38MAPK and TGFβ1 in the H group increased.Compared with the H group cells,the mRNA expression of p38MAPK and TGFβ1 in the V group and B group decreased (P<0.01).5.The protein expression of p38MAPK and TGFβ1:Compared with the N group cells,the protein expression of p38MAPK and TGFβ1 in H group increased.Compared with the H group cells,the protein expression of p38MAPK and TGFβ1 in the V group and B group decreased (P<0.01).6.The p38MAPK protein mainly located in cytoplasm and nucleus.there is hardly any p38MAPK protein in cell membrane and cells stromal.The TGFβ1 protein mainly located in cytoplasm,the protein can be found in membrane in a small number of cells,the protein can be found in nucleus only in a very small part of the cells.There is hardly any TGFβ1 protein in the cells stromal.Conclusion:1.High glucose have a proliferation effect on the rat mesangial cells in a certain period of time.There is nothing to do with the osmotic pressure. Valsatan can reduce the proliferation effect caused by high glucose and protect the rat mesangial cells by counteracting oxidative stress.2.p38MAPK is actived in the mesangical cells of diabetic nephropathy. Valsartan can inhibit the p38MAPK activation in the mesangial cells of diabetic nephropathy and play a role in kidney protection by reducing the expression of TGF-β1.