CAJ | 학술논문

目的研究RhoA/Rho激酶(ROCK)信号通路对Toll样受体2(TLR-2)配体诱导的类风湿关节炎(RA)成纤维样滑膜细胞(FLS)分泌趋化因子的影响.方法 RA FLS来源于活动性RA患者滑膜组织;肽聚糖被用于TLR-2配体;RhoA活性检测采用pull down方法;ROCK活性用磷酸化肌球蛋白结合亚单位1(MYPT1)蛋白表达来表示,磷酸化MYPT1蛋白检测采用免疫印迹法;细胞活性检测采用四甲基偶氮唑蓝比色(MTT)法,趋化因子水平采用酶联免疫吸附测定法(ELISA)检测.结果肽聚糖(5 mg/L)刺激使体外培养的RA FLS分泌白细胞介素8(IL-8)、单核细胞趋化蛋白-2(MCP-2)和受激活调节正常T细胞表达和分泌因子(RANTES)明显增高,对RhoA和ROCK活化也有显著的刺激作用,肽聚糖诱导的上述作用能被TLR-2单克隆抗体所阻抑.RhoA抑制剂C3转化酶和ROCK抑制剂Y27632对PG诱导的IL-8、MCP-2和RANTES等趋化因子分泌有显著的抑制作用.结论 RhoA/ROCK信号通路对TLR2介导的RA FLS分泌趋化因子具有调控作用,通过抑制该通路活化可能有利于RA的治疗.
목적 연구RhoA/Rho격매(ROCK)신호통로대Toll양수체2(TLR-2)배체유도적류풍습관절염(RA)성섬유양활막세포(FLS)분비추화인자적영향.방법 RA FLS래원우활동성RA환자활막조직;태취당피용우TLR-2배체;RhoA활성검측채용pull down방법;ROCK활성용린산화기구단백결합아단위1(MYPT1)단백표체래표시,린산화MYPT1단백검측채용면역인적법;세포활성검측채용사갑기우담서람비색(MTT)법,추화인자수평채용매련면역흡부측정법(ELISA)검측.결과 태취당(5 mg/L)자격사체외배양적RA FLS분비백세포개소8(IL-8)、단핵세포추화단백-2(MCP-2)화수격활조절정상T세포표체화분비인자(RANTES)명현증고,대RhoA화ROCK활화야유현저적자격작용,태취당유도적상술작용능피TLR-2단극륭항체소조억.RhoA억제제C3전화매화ROCK억제제Y27632대PG유도적IL-8、MCP-2화RANTES등추화인자분비유현저적억제작용.결론 RhoA/ROCK신호통로대TLR2개도적RA FLS분비추화인자구유조공작용,통과억제해통로활화가능유리우RA적치료.
Objective To evaluate the modulation of RhoA/Rho kinase (ROCK) signaling pathway,a small Rho GTPase that is considered as an important modulator in inflammatory responses,on Toll-like receptor-2 mediated chemokine secretion in fibroblast-like synoviocytes (FLS)from rheumatoid arthritis (RA) patients.Methods The RhoA activity was measured by a pull-down assay.And the ROCK activity was assessed by Western blot.The secretion of chemokines was measured by ELISA (enzyme-linked immunosorbent assay).MTT test was used to detect the cellular viability.Results The stimulation of peptidoglycan(PG,5 mg/L) increased the levels of IL-8 (interleukin-8),RANTES (regulated upon activation normal T cell expressed & secreted) and MCP-2 (monocyte chemotactic protein-2)and boosted the activities of RhoA and ROCK versus the unstimulated RA FLS.And these effects of PG were suppressed by anti-TLR-2 monoclonal antibody.Inhibition of RhoA and ROCK with a specific inhibitor inhibited the secretion of IL-8,RANTES and MCP-2 in PG-induced RA FLS.Conclusion The present study provides novel evidence that the RhoA/ROCK signal pathway modulates the TLR-2-mediated secretion of chemokines in RA FLS.It suggests that the inhibition of RhoA/ROCK may be a new therapeutic approach for RA.