中文摘要：

目的：初步探讨HMGB1是否通过Toll like receptor-2 （TLR-2）促进小鼠系膜细胞增殖.方法：选取小鼠系膜细胞MMC为研究对象,为了确定HMGB1对小鼠系膜细胞增殖的影响,将小鼠系膜细胞随机分为正常对照组及50、100和200μg/L HMGB1刺激组,分别培养2、4、8和12 h,以BrdU掺人法检测小鼠系膜细胞的增殖水平.为了检测HMGB1对小鼠系膜细胞TLR2、PCNA、Cyclin D1、CDK4、p16表达的影响,将常规培养的MMC细胞随机分为正常对照组及100 μg/L HMGB1刺激组,分别于培养2、4、6、8和12 h收集细胞,采用Real-time PCR及Western blot检测小鼠系膜细胞TLR2、PCNA、Cyclin D1、CDK4、p16的表达变化.为了明确TLR2表达与HMGB1介导的MMC增殖中的作用,将MMC细胞随机分为正常对照组、HMGB1刺激组（100μg/L）、HMGB1刺激＋pYr-3.1-shTLR2组（转染组）、HMGB1刺激＋空白质粒组（空白质粒组）,刺激8h收集细胞,Western blot检测pYr-3.1-shTLR2的沉默效果及细胞周期蛋白PCNA、Cyclin D1、CDK4、p16蛋白的表达变化.结果：HMGB1上调小鼠系膜细胞增殖水平;HMGB1时间依赖性上调小鼠系膜细胞TLR2的表达;HMGB1刺激能够上调小鼠系膜细胞中PCNA、Cyclin D1、CDK4mRNA及蛋白的表达,时间依赖性地降低小鼠系膜细胞中p16的表达;RNA干扰沉默TLR2表达,能够有效地下调HMGB1诱导的小鼠系膜细胞中细胞周期蛋白的表达,降低系膜细胞的增殖水平.结论：HMGB1可能通过与小鼠系膜细胞膜表面受体TLR2结合后,通过某种信号途径上调细胞周期调控蛋白Cyclin D1、CDK4的表达,并下调细胞周期依赖性激酶抑制剂p16的表达,推动细胞从G0/G1期进入S期,促使细胞增殖水平提高.

英文摘要：

Objective：To explore the effect and possible mechanism of TLR2 on cell proliferation of MMC cells induced by HMGB1.Methods：Mouse renal mesangial cells line （MMC） were grown as described previously.Cells were made quiescent by culturing in serum-free medium for 24 h.In order to determine the effect of HMGB1 on cell proliferation of MMC cells,cells were randomly divided into normal control group,50,100 and 200 μg/L HMGB1 stimulated group,and cells were collected at 2,4,8 and 12h,BrdU assay was used to detect cell proliferation level.To determine the time-dependent effect of HMGB1 on the expression of TLR2,PCNA,Cyclin D1,CDK4 and pl6,MMC cells were randomly divided into normal control group,100 μg/L HMGB1 stimulated group,and were collected at 2,4,6,8 and 12 h respectively.Real-time PCR and Western Blot were used for the detection of TLR2,PCNA,Cyclin D1,CDK4 and p16 expressions.RNAi experiment was performed to assess the effect of TLR2 on HMGB1-stimulated cell proliferation.Cells were divided into four groups：control,untransfected HMGB1-treated,control vector sh-Scramble-transfected HMGB1-treated and shTLR2-transfected HMGB1-treated.Cells were collected after 8 h,Western blot were used to detect the silence effect of pYr-3.1-shTLR2 and the expression of PCNA,Cyclin D1,CDK4 and p16 protein.Results：HMGB1 upregulated the level of mouse mesangial cell proliferation; HMGB1 induced TLR2 upregulation of MMC cells in time-dependent manner; HMGB1 upregulated the expression of PCNA,Cyclin D1 and CDK4 protein in mouse mesangial cells; HMGB1 down-regulated the p16 expression time-dependently in mouse mesangial cells ;the sh-TLR2 vector effectively prevented HMGBl-induced cell proliferation,decreased Cyclin D1/CDK4/p16 pathway and PCNA expression.Conclusion：HMGB1 might combined with its surface receptor TLR2,and then up-regulated the expression of cell cycle regulatory protein Cyclin D1,CDK4 and down-regulated the expression of the cyclin-dependent kinase inhibitor p16 through activating signaling pathways to ind