摘要
目的探讨细胞因子信号转导抑制因子3(SOCS3)对IgA肾病(IgAN)患者血清IgA1刺激诱导人肾小球系膜细胞(HMC)增殖的作用及其可能机制。方法采用Jacalin亲和层析联合丙烯葡聚糖凝胶S-200凝胶过滤法纯化聚合IgAN患者血清热聚合IgA1(aIgA1)。以腺病毒为载体转染SOCS3至体外培养的HMC,再以aIgA1刺激培养12~48h,分为空白对照组、IgA1组、IgA1+空载腺病毒组、IgA1+SOCS3腺病毒组。采用噻唑蓝(MTT)法检测系膜细胞增殖情况,Western印迹法和实时定量PCR法检测各组SOCS3、Toll样受体4(TLR4)、转化生长因子β1(TGF-β1)蛋白和mRNA表达水平。结果algA1刺激HMC24h后开始表现出促增殖作用,与空白对照组相比,aIgA1刺激后SOCS3、TLR4、TGF-β1蛋白和mRNA表达均有显著升高(P〈0.05)。与IgA1组、IgA1+空载腺病毒组相比,IgA1+SOCS3腺病毒组TLR4、TGF-β1蛋白和mRNA表达均显著下降(P〈0.05),HMC增殖程度显著降低(P〈0.05)。结论上调SOCS3表达可降低aIgA1刺激后HMC的TLR4和TGF-β1表达水平,并抑制aIgA1诱导的HMC细胞增殖。
Objective To investigate the effect of suppressor of cytokinc signaling 3 (SOCS3) on the proliferation of human mesangial ceils stimulated by aggregated IgA1 (aIgA1) from patients with IgA nephropathy(IgAN), and explore its possible mechanism. Methods Serum monomeric IgA1 was isolated with jaealin affinity and Sephacryl S- 200 HR chromatography from IgAN patients, and then heated to aggregated form (algA1). Human glomerular mesangial cells(HMC) were transfected with Adv- SOCS3-IRES2-EGFP for 48 hours, and incubated with aIgA1 for 12-48 h. The cells were divided into blank control group, IgA1 group, IgA1+Adv-EGFP group and IgA1+Adv-SOCS3-IRES2-ECFP group. The mesangial cell proliferation was observed through MTT, and the levels of SOCS3, TLR4, TGF-β1 protein and mRNA were detected through Western blotting and real- time PCR. Results HMC proliferation was promoted significantly after IgA1 stimulated at 24 h. Compared with control group, the protein and mRNA expression of SOCS3, TLR4, TGF- β1 were significantly increased in IgA1 group (P 〈 0.05). Compared with IgA1 group and IgA1 +Adv-EGFP group, MTT absorbency was obviously reduced after incubation with aIgA1 for 24 h and 48 h in IgA+Adv-SOCS3-IRES2-EGFP group, and the protein and mRNA expression of TLR4 and TGF-β1 were significantly decreased in IgA1 +Adv- SOCS3- EGFP group (P 〈 0.05). Conclusion Over- expression of SOCS3 may inhibit the proliferation of HMC stimulated by aIgA1, partly through down-regulating the expression of TLRg and TGF-β1.
出处
《中华肾脏病杂志》
CSCD
北大核心
2017年第8期616-622,共7页
Chinese Journal of Nephrology
基金
国家自然科学基金(81160091)
江西省自然科学基金(2010GQY0324)