白细胞介素-22促进类风湿关节炎成纤维化膜细胞的增殖

目的 通过白细胞介素-22(IL-22)干预类风湿关节炎(RA)成纤维滑膜细胞(FLS)以期明确IL-22促进其增殖的机制.方法 无菌条件下获取的RA滑膜组织,应用组织块分离法培养RA-FLS,含10% FBS的DMEM培养基进行细胞传代培养,并对第4~5代FLS应用vimentin/CD68进行免疫组化法鉴定;不同浓度的IL-22干预FLS培养24、48、72 h后,MTT法检测细胞增殖程度;联合IL-22和/或AG490干预FLS,于冰面上用含蛋白酶抑制剂的细胞裂解液获取细胞总蛋白,再经超声、离心提取、蛋白定量后,Western blot法检测STAT3、ERK1/2、P38蛋白及其磷酸化蛋白的表达,统计分析采用SPSS20.0软件进行,P<0.05为差异有统计学意义.结果 不同浓度IL-22干预FLS的增殖均较对照组呈梯度增高变化,差异具有统计学意义(P<0.05).随着IL-22干预时间的延长,STAT3总蛋白表达相对灰度值的整体水平差异无统计学意义(P=0.68),但是其磷酸化蛋白表达相对灰度值整体水平差异具有统计学意义(P<0.001),且在干预的不同时间点STAT3磷酸化蛋白相对表达量较基线0 h时均显著升高,且差异具有统计学意义(P<0.001).IL-22干预后的ERK1/2和P38总蛋白及其磷酸化蛋白表达相对灰度值差异均无统计学意义(P>0.05).联合50 ng/mL IL-22和100 μmol/L AG490干预FLS不同时间后,细胞的增殖程度较单用IL-22干预均显著降低(P<0.01).结论 IL-22以浓度依赖方式促进RA成纤维化膜细胞STAT3蛋白磷酸化所介导的细胞增殖,而非ERK1/2和P38蛋白所介导的信号通路.

Interleukin-22 promotes proliferation of fibroblast-like synoviocytes from patients with rheumatoid arthritis by inducing STAT3 phosphorylation

Objective To clarify the mechanism by which interleukin-22 (IL-22) promotes the proliferation of fibroblast-like synoviocytes (FLS) from patients with rheumatoid arthritis (RA). Methods FLS were isolated from the synovial tissues of patients with RA and identified by immunohistochemistry for vimentin/CD68.The cells were subcultured and incubated with different concentrations of IL-22 for 24,48,or 72 h,and their proliferation was examined using MTT assay.After treatment of the cells with IL-22 and AG490,alone or in combination,the expressions of the total and phosphorylated proteins of STAT3, ERK1/2 and P38 were detected with Western blotting. Results IL-22 significantly increased the proliferation of FLS in a dose-dependent manner(P<0.05).The total protein of STAT3 in the cells showed no significant changes with extended time of IL-22 treatment(P=0.68),but the expression of phosphorylated STAT3 protein increased significantly(P<0.001).The total and phosphorylated proteins of ERK1/2 and P38 underwent no significant changes after IL-22 treatment (P>0.05). A combined treatment with 50 ng/mL IL-22 and 100 μmol/L AG490 resulted in a significant decrease in the proliferation of FLS as compared with IL-22 treatment alone(P<0.01).Conclusion IL-22 can dose-dependently promote the proliferation of FLS from patients with RA by inducing phosphorylation of STAT3 protein but not through ERK1/2 or P38 signal pathway.