Impact of STAT1-siRNA transfection on the expressions of STAT3 and transforming growth factor β1 in human glomerulur mesangial cells induced by high glucose |
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Authors: | ZHENG Xiao-yu JIAO Su-min WANG Li-ning CHEN Ying YANG Xue ZHANG Jin YANG Shuang GAO Xin-ran FAN Qiu-ling |
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Institution: | Department of Nephrology, The First Affiliated Hospital, China Medical University, Shenyang 110001, China;
Corresponding author:WANG Li-ning, Email: lnwang56@medmail.com.cn |
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Abstract: | Objective To observe the cell proliferation and the protein expression of STAT1,phosphorylation of STAT1 (p-STAT1), STAT3, p-STAT3 and transforming growth factor β1 (TGF-β1) in human glomerulur mesangial cells (HMCs) induced by high glucose after STAT1-siRNA transfection. Methods Three STAT1-siRNA sequences were designed and synthetized. HMCs in 6-well plate were transiently transfected with STAT1-siRNA using Lipofectamine 2000. After transfection for 48 h or 72 h, STAT1 mRNA and protein expression were detected by real-time PCR and Western blotting, respectively, to choose the effective sequence in later experiments. After transfection for 24 h and stimulated with 25 mmol/L glucose for 24 h, 48 h, 72 h, cell proliferation was measured by MTT assays, the protein expressions of STAT1, p-STAT1, STAT3 and p-STAT3 were detected by Western blotting, the expression of TGF-β1 was detected by ELISA in each group. Results High glucose could stimulate HMCs proliferation. The protein expressions of p-STAT1, p-STAT3 and TGF-β1 were increased in the group stimulated by high glucose (P<0.05). The protein expressions of p-STAT3 and TGF-β1 were further increased in HMCs induced by high glucose after STAT1-siRNA transfection (P<0.05). Conclusions Under high glucose conditions, JAK-STAT signal transduction pathway of HMCs can be activated, then it is far greater when HMCs are induced by high glucose after STAT1-siRNA transfection. The secretion of TGF-β1 is increased in HMCs under the state of high glucose, and it is further increased after STAT1-siRNA transfection, which is related to the kidney fibrosis. |
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