| Background and objective:Diabetic nephropathy(DN)is one of the common and serious complications of diabetic mellitus(DM).It is the primary cause of end stage renal disease(ESRD)need renal replacement therapy in developed countries,and the number of patients with end-stage kidney disease caused by diabetic nephropathy in China is also increasing year by year.The exact causes and pathophysiological mechanisms of diabetic nephropathy have not been fully elucidated,so it is very important to find effective treatment measures to prevent the occurrence of diabetic nephropathy and delay the progression of renal function.In recent years,a large number of studies have shown that inflammation plays an important role in the pathogenesis of diabetic nephropathy.Monocyte chemoattractant protein-1(MCP-1),tumor necrosis factor-α(TNF-α),interleukin-6(IL-6)and other inflammatory cytokines participate in and promote the progress of DN.MicroRNAs(miRNAs)are single-stranded RNAs widely existed in the human body that can negatively regulate the expression of relative target genes,which play an important role in cell proliferation,differentiation and energy metabolism.Recently,evidence of mi RNAs involved in DN development has been widely reported.Among them,MicroRNA-374a(miRNA-374a)has been proved to inhibit the proliferation and metastasis of breast cancer cells through its target gene MCP-1.A clinical trial on lupus nephritis has also described the targeted regulatory relationship between miRNA-374 a and MCP-1.Whether miRNA-374 a could regulate the expression of MCP-1 and other proinflammatory cytokines,and thus participate in the progression of DN is still unclear.Simulation of diabetes mellitus with high glucose environment,this experiment study on the human tubular epithelial cells(HK2).To research the effect of miRNA-374 a on the expression of MCP-1 and related inflammatory factors in human tubular epithelial cells(HK2),and to explore the possible mechanisms of miRNA-374 a regulating the expression of proinflammatory factors and mediating inflammation participated the progress of DN.In order to find new treatment ideas for the intervention of early diabetic nephropathy and the delay of its progress.Methods:HK2 were cultured in DMEM medium which contain 10% fetal bovine serum.Cells were divided into following groups:(1)Normal Control Group(D-glucose 5.6mmol/L),Mannitol group(D-glucose 5.6 mmol/L+D-mannitol 24.4mmol/L),High glucose group(D-glucose 30mmol/L);(2)Regulation of miRNA-374 a expression by transfection of miRNA-374 a mimics and miRNA-374 a inhibitor.Divide into: Blank control group,Transfection control group,miRNA-374 a mimics group,miRNA-374 a inhibitor group.The above four groups were co-cultured with normal glucose,mannitol and high glucose.Western blotting was used to test the protein expression of MCP-1.The supematant level of mRNA expression of MCP-1 and mi RNA-374 a was dectected by Real Time-PCR.ELISA was used to assess the levels of protein expression of interleukin-6(IL-6),interleukin-18(IL-18)and tumor necrosis factor-α(TNF-α).Results:Compared with the control group,the expression of mi RNA-374 a in the high glucose group were decreased(P <0.05),the protein and mRNA expression of MCP-1 were increased,and the protein expression of IL-6,IL-18,TNF-α were all increased(P <0.05).Compared with the blank control group,the mRNA expression of MCP-1 were decreased in miRNA-374 a mimics group,the protein expression of IL-6,IL-18,TNF-α were also decreased(P <0.05).The mRNA expression of MCP-1,the protein expression of IL-6,IL-18,TNF-α were increased in miRNA-374 a inhibitor group.Conclusions:Micro RNA-374 a may be involved in the process of inflammatory injury of diabetic nephropathy by regulating the expression of cytokines such as MCP-1、IL-6、IL-18 and TNF-α in human renal tubular epithelial cells under high glucose condition. |
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