【摘要】 目的探讨细胞因子信号负调控因子(SOCS)3基因对高糖培养下人肾小球系膜细胞(HMCs)增殖、凋亡的影响。方法以脂质体为载体将PCR3.1 SOCS3转染至体外培养的HMCs,应用反转录聚合酶链反应(RT-PCR)和免疫细胞化学法鉴定转染前后细胞中SOCS3 mRNA和蛋白表达;高糖培养HMCs 12、24、48 h,采用蛋白印迹法分别检测正常对照组(CG)、高糖组(HG)、高糖+空质粒转染(HG+PV)组、高糖+SOCS3基因转染(HG+PS)组蛋白酪氨酸磷酸激酶(JAK)2、信号转导及转录活化因子(STAT)1蛋白酪氨酸磷酸化水平变化;免疫细胞化学法、末端脱氧核苷酸转移酶介导的dUTP缺口末端标记技术(TUNEL)法检测转染前后细胞增殖指数(PI)和凋亡指数(AI)。结果经鉴定,转染后HMCs中有SOCS3 mRNA和蛋白稳定表达;与CG组和HG+PV组相比,HG+PS组细胞中JAK2、STAT1蛋白酪氨酸磷酸化水平显著下降(P<0.05),PI显著减少(P<0.01),AI无明显差异。结论 SOCS3蛋白可能通过降低JAK2、STAT1酪氨酸磷酸化水平抑制HMCs增殖。更多还原

【Abstract】 Objective To investigate the effects of SOCS3 gene on proliferation and apoptosis of human glomerular mesangial cells(HMCs) upon high glucose(HG) stimulation. Methods HMCs were transfected with PCR3.1 SOCS3 by using liposomes as carriers. The expressions of SOCS3 mRNA and protein were detected by reverse transcriptase polymerase chain reaction and immunocytochemistry assay, respectively. Following incubation under HG conditions, the cells were divided into blank control group(CG), HG group(HG), HG +PCR3.1 SOCS3 transfection group(HG+PS) and HG+PCR 3.1-vector transfection group(HG+PV) for the determination of the levels of tyrosinephosphorylated JAK2 and STAT1 level by using Western blotting at 12, 24 48 hours after HG stimulation. Immunocy-tochemistry assay and TUNEL assay were adopted to analyze the SOCS3 tranfected HMCs proliferation activity index(PI) and cell apoptosis index(AI) 48 hours after HG stimulation. Results The SOCS3 mRNA and protein expression was confirmed following transfection with SOCS3 gene. The CG group and HG+PV group yielded markedly reduced levels of tyrosine-phosphorylated JAK2 and STAT1(P<0.05) and PI(P<0.01), but not AI(P>0.05). Conclusion SOCS3 protein may inhibit the proliferation of HMCs by down-regulating the tyrosine-phosphorylated levels of JAK2 and STAT1 protein.更多还原