活血通络方对类风湿关节炎滑膜成纤维细胞miR-155及相关细胞因子表达的影响
|
摘要
目的:观察活血通络方对白细胞介素-1β(IL-1β)刺激类风湿关节炎(RA)滑膜成纤维细胞(HFLS)增殖和miR-155以及相关细胞因子分泌的影响。方法:HFLS-RA培养到第4代至第8代。经IL-1β诱导后给予含药血清,采用MTT法检测空白组、模型组、对照组、低剂量组、中剂量组、高剂量组含药血清对HFLS-RA增殖的影响,荧光定量PCR法检测miR-155及相关细胞因子表达。结果:MTT法测定HFLS-RA增殖活性的结果显示,模型组比正常组细胞数量明显增多(P <0.01),对照组和高、中剂量组增殖活性明显降低(P <0.01),低剂量组未表现出对HFLS-RA增殖有显著影响。对照组及低、中、高剂量组miR-155、IL-2、IFN-γ和RANKL相对表达水平均低于模型组,IL-4、IL-10相对表达水平高于模型组(P <0.05)。高剂量组miR-155、IL-2、IFN-γ和RANKL相对表达水平均低于对照组,IL-4、IL-10相对表达水平高于对照组(P <0.05),中剂量组miR-155、IL-2、IL-4、IL-10、IFN-γ和RANKL相对表达水平与对照组比较,差异无统计学意义(P> 0.05)。结论:活血通络方抑制HFLS-RA增殖,其治疗RA可能通过下调miR-155表达抑制RA滑膜成纤维细胞中IL-2、IFN-γ和RANKL等细胞因子表达,提高IL-4、IL-10等细胞因子表达相关。
Objective:To observe the effects of Huoxue Tongluo Fang(活血通络方)on the proliferation of rheumatoid arthritis(RA)synovial fibroblasts(HFLS)stimulated by interleukin-1β(IL-1β)and the secretion of miR-155 and related cytokines.Methods:HFLS-RA was cultured to the fourth to eighth generations.After induction by IL-1β,drug-containing serum was given.MTT assay was used to detect the effect of blank group,model group,control group,low-dose group,medium-dose group and high-dose group of drug-containing serum on the proliferation of HFLS-RA.Fluorescence quantitative PCR was used to detect the expression of miR-155 and related cytokines.Results:MTT assay showed that the number of cells in the model group was significantly higher than that in the normal group(P < 0.01).The proliferative activity of the control group and the highdose and medium-dose groups was significantly lower(P < 0.01).The low-dose group did not show significant effect on the proliferation of HFLS-RA.The relative expression levels of miR-155,IL-2,IFN-γ and RANKL in the control group,the low-dose group,the medium-dose group and the high-dose group were lower than those in the model group,while the relative expression levels of IL-4 and IL-10 were higher than those in the model group(P < 0.05).The relative expression levels of miR-155,IL-2,IFN-γ and RANKL in the high dose group were lower than those in the control group,while the relative expression levels of IL-4 and IL-10 were higher than those in the control group(P < 0.05).There was no significant difference in the relative expression levels of miR-155,IL-2,IL-4,IL-10,IFN-γ and RANKL between the middle dose group and the control group(P > 0.05).Conclusion:Huoxue Tongluo Fang can inhibit the proliferation of HFLS-RA,and its treatment for RA may inhibit the expression of IL-2,IFN-γ and RANKL in RA synovial fibroblasts by down-regulating the expression of miR-155,and increase the expression of IL-4,IL-10 and other cytokines.
Objective:To observe the effects of Huoxue Tongluo Fang(活血通络方)on the proliferation of rheumatoid arthritis(RA)synovial fibroblasts(HFLS)stimulated by interleukin-1β(IL-1β)and the secretion of miR-155 and related cytokines.Methods:HFLS-RA was cultured to the fourth to eighth generations.After induction by IL-1β,drug-containing serum was given.MTT assay was used to detect the effect of blank group,model group,control group,low-dose group,medium-dose group and high-dose group of drug-containing serum on the proliferation of HFLS-RA.Fluorescence quantitative PCR was used to detect the expression of miR-155 and related cytokines.Results:MTT assay showed that the number of cells in the model group was significantly higher than that in the normal group(P < 0.01).The proliferative activity of the control group and the highdose and medium-dose groups was significantly lower(P < 0.01).The low-dose group did not show significant effect on the proliferation of HFLS-RA.The relative expression levels of miR-155,IL-2,IFN-γ and RANKL in the control group,the low-dose group,the medium-dose group and the high-dose group were lower than those in the model group,while the relative expression levels of IL-4 and IL-10 were higher than those in the model group(P < 0.05).The relative expression levels of miR-155,IL-2,IFN-γ and RANKL in the high dose group were lower than those in the control group,while the relative expression levels of IL-4 and IL-10 were higher than those in the control group(P < 0.05).There was no significant difference in the relative expression levels of miR-155,IL-2,IL-4,IL-10,IFN-γ and RANKL between the middle dose group and the control group(P > 0.05).Conclusion:Huoxue Tongluo Fang can inhibit the proliferation of HFLS-RA,and its treatment for RA may inhibit the expression of IL-2,IFN-γ and RANKL in RA synovial fibroblasts by down-regulating the expression of miR-155,and increase the expression of IL-4,IL-10 and other cytokines.
引文
[1]何宏军,寇静鑫,王玲,等.联合云克治疗类风湿关节炎的中远期疗效观察[J].风湿病与关节炎,2016,5(11):15-18.
[2]冯佳,夏燕,陈安平,等.类风湿关节炎患者外周血miR-146a及miR-155的表达及临床意义[J].风湿病与关节炎,2016,5(10):8-11.
[3]SPOERL D,DUROUX-RICHARD I,LOUIS-PLENCEP,et al.The role of miR-155 in regulatory T cells and rheumatoid arthritis[J].Clin Immunol,2013,148(1):56-65.
[4]ALIVERNINI S,GREMESE E,MCSHARRY C,et al.MicroRNA-155-at the critical interface of innate and adaptive immunity in arthritis[J].Front Immunol,2017(8):1932.
[5]丁德经,周学平.辨病与辨证相结合治疗类风湿性关节炎[J].中华中医药杂志,2016,31(12):5058-5061.
[6]于萍,龙丽,王世瑶,等.MiR-155在类风湿关节炎滑膜成纤维细胞中的表达及功能研究[J].中华风湿病学杂志,2010,14(7):460-463.
[7]靖卫霞,朱跃兰.浅析活络效灵丹治疗类风湿关节炎瘀证[J].湖南中医药大学学报,2017,37(4):395-397.
[8]GEUSENS PP,LANDEWéRB,GARNERO P,et al.The ratio of circulating osteoprotegerin to RANKL in early rheumatoid arthritis predicts later joint destruction[J].Arthritis Rheum,2006,54(6):1772-1777.
[9]张良,白人骁.细胞因子与类风湿性关节炎治疗新进展[J].中国医药导报,2008,5(11):27-29.
[10]施晓柯.类风湿性关节炎相关细胞因子的研究进展[J].中山大学研究生学刊:自然科学与医学版,2012,33(4):1-5.
[11]王玉明,张秦.雷公藤多甙片在类风湿关节炎中的应用体会[J].风湿病与关节炎,2013,2(7):60-61.
[12]彭桉平,黄宪章,刘瑞萍,等.雷公藤内酯醇调控miR-155抑制类风湿性关节炎患者单核细胞促炎反应[J].细胞与分子免疫学杂志,2014,30(6):635-638.
[2]冯佳,夏燕,陈安平,等.类风湿关节炎患者外周血miR-146a及miR-155的表达及临床意义[J].风湿病与关节炎,2016,5(10):8-11.
[3]SPOERL D,DUROUX-RICHARD I,LOUIS-PLENCEP,et al.The role of miR-155 in regulatory T cells and rheumatoid arthritis[J].Clin Immunol,2013,148(1):56-65.
[4]ALIVERNINI S,GREMESE E,MCSHARRY C,et al.MicroRNA-155-at the critical interface of innate and adaptive immunity in arthritis[J].Front Immunol,2017(8):1932.
[5]丁德经,周学平.辨病与辨证相结合治疗类风湿性关节炎[J].中华中医药杂志,2016,31(12):5058-5061.
[6]于萍,龙丽,王世瑶,等.MiR-155在类风湿关节炎滑膜成纤维细胞中的表达及功能研究[J].中华风湿病学杂志,2010,14(7):460-463.
[7]靖卫霞,朱跃兰.浅析活络效灵丹治疗类风湿关节炎瘀证[J].湖南中医药大学学报,2017,37(4):395-397.
[8]GEUSENS PP,LANDEWéRB,GARNERO P,et al.The ratio of circulating osteoprotegerin to RANKL in early rheumatoid arthritis predicts later joint destruction[J].Arthritis Rheum,2006,54(6):1772-1777.
[9]张良,白人骁.细胞因子与类风湿性关节炎治疗新进展[J].中国医药导报,2008,5(11):27-29.
[10]施晓柯.类风湿性关节炎相关细胞因子的研究进展[J].中山大学研究生学刊:自然科学与医学版,2012,33(4):1-5.
[11]王玉明,张秦.雷公藤多甙片在类风湿关节炎中的应用体会[J].风湿病与关节炎,2013,2(7):60-61.
[12]彭桉平,黄宪章,刘瑞萍,等.雷公藤内酯醇调控miR-155抑制类风湿性关节炎患者单核细胞促炎反应[J].细胞与分子免疫学杂志,2014,30(6):635-638.