CAJ | 학술논문

目的探讨高迁移率族蛋白(high mobility group box,HMGB)1对大鼠滑膜细胞STAT信号通路的调控作用.方法将常规培养大鼠滑膜细胞株RSC-364细胞随机分为正常对照组和10μg/L HMGB1刺激组,分别培养6 h、12 h和24 h,RT-PCR检测STAT 1 / 3 mRNA的表达,Western印迹法和流式细胞术(flow cytometric analysis,FCM)检测STAT 1、STAT3、磷酸化STAT 1(phospho-STAT1,p- STAT1)和磷酸化 3(phospho-STAT3,p- STAT3)蛋白的表达,免疫细胞化学(ICC)检测PCNA蛋白的表达.结果 HMGB1 刺激6 h~24 h 组STAT 1 mRNA 和p-STAT1蛋白的相对表达量呈时间依赖性上调,24 h表达最高,与正常对照组相比差异均具有显著统计学意义(P＜0.05,P＜0.01).PCNA蛋白阳性信号表达于细胞核内,呈棕黄色颗粒,且随着刺激时间延长阳性信号逐渐增强.RT-PCR、ICC染色和FCM均显示STAT3 mRNA和p-STAT3蛋白的相对表达量各组间相比差异均无统计学意义.结论 HMGB1可能通过激活STAT1信号转导通路促进滑膜细胞的增殖和分化.
목적 탐토고천이솔족단백(high mobility group box,HMGB)1대대서활막세포STAT신호통로적조공작용.방법 장상규배양대서활막세포주RSC-364세포수궤분위정상대조조화10μg/L HMGB1자격조,분별배양6 h、12 h화24 h,RT-PCR검측STAT 1 / 3 mRNA적표체,Western인적법화류식세포술(flow cytometric analysis,FCM)검측STAT 1、STAT3、린산화STAT 1(phospho-STAT1,p- STAT1)화린산화 3(phospho-STAT3,p- STAT3)단백적표체,면역세포화학(ICC)검측PCNA단백적표체.결과 HMGB1 자격6 h~24 h 조STAT 1 mRNA 화p-STAT1단백적상대표체량정시간의뢰성상조,24 h표체최고,여정상대조조상비차이균구유현저통계학의의(P＜0.05,P＜0.01).PCNA단백양성신호표체우세포핵내,정종황색과립,차수착자격시간연장양성신호축점증강.RT-PCR、ICC염색화FCM균현시STAT3 mRNA화p-STAT3단백적상대표체량각조간상비차이균무통계학의의.결론 HMGB1가능통과격활STAT1신호전도통로촉진활막세포적증식화분화.
Objective To investigate the effect signal transducer and activator of transcription (STAT) on proliferation process of rat class fibroblast-like synovial cells induced by high mobility group box (HMGB) 1.Methods RSC-364 cells induced by 10μg/L recombinant HMGB1 were collected in 6 h、12 h、24 h respectively,as well as normal control group cells in vitro.The expression of STAT 1 / 3 mRNA were analyzed by RT-PCR technique.Western blotting and flow cytometric analysis (FCM) were adopted to detect the expression of STAT1,STAT3,p-STAT1 and p-STAT3.Immunocytochemistry(ICC) staining was adopted to detect the expression of PCNA protein.Results The relative expression of STAT1 mRNA and p-STAT1 proterin was significantly up-regulated in time-dependent manner from 6 h group to 24 h group,the highest expression of them discovered at 24 h.Compared with normal control group,there were significant statistic differences (P＜0.05 or 0.01).The positive signal expression of PCNA proteins were localized in nucleus as buffy clumping,and gradually increased in time-dependent manner too.But the changes of STAT3 mRNA and p-STAT3 protein expression indicated no significant statistical differences among normal control group and 6 h~24 h groups by RT-PCR,ICC staining and FCM.Conclusion HMGB1 could activate STAT1 signal transduct process to promote cell proliferation of rat class fibroblast-like synovial cells.