天津医药

天津医药 ›› 2018, Vol. 46 ›› Issue (2): 113-117.doi: 10.11958/20171299

• 细胞与分子生物学 •    下一篇

类风湿关节炎成纤维样滑膜细胞中钙网织蛋白经NF-κB 通路上调c-FLIP的表达

焦亚冲1 ， 黄珊珊2 ， 万春友3 ， 刘宜昕1 ， 王阳1 ， 白英玉1 ， 马骏4 ， 郑芳1△   

  1. 1天津医科大学医学检验学院免疫学教研室 （邮编300203）； 2河北省儿童医院； 3天津市天津医院； 4天津医科大学公共卫生学院
  • 收稿日期:2017-11-17 修回日期:2017-12-25 出版日期:2018-02-15 发布日期:2018-02-15
  • 通讯作者: 焦亚冲 E-mail:1490662738@qq.com
  • 基金资助:
    天津市应用基础与前沿技术研究计划项目;天津医科大学“十三五”综合投资学科建设项目

CRT promoted c-FLIP expression via NF-κB pathway in rheumatoid arthritis fibroblast-like synoviocytes

CRT promoted c-FLIP expression via NF-κB pathway in rheumatoid arthritis fibroblast-like synoviocytes   

  • Received:2017-11-17 Revised:2017-12-25 Published:2018-02-15 Online:2018-02-15

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摘要： 摘要： 目的 探究类风湿关节炎 （RA） 患者成纤维样滑膜细胞 （FLS） 中钙网织蛋白 （CRT） 上调细胞型 Fas 相关死亡域样白细胞介素 1β 转换酶抑制蛋白 （c-FLIP） 表达的分子机制。方法 免疫组织化学染色分析 c-FLIP 在 RA 和骨关节炎 （OA） 患者膝关节滑膜中的表达与定位。酶解法分离 RA/OA FLS， 作为体外细胞培养模型。细胞免疫荧光和 Western blot 检测 c-FLIP 在 RA/OA FLS 中的表达。Western blot 探究 CRT 对 RA FLS 中 c-FLIP 表达的影响及其分子机制。结果 c-FLIP 在 RA 滑膜中呈高表达， 主要集中在衬里层和衬里下层的 FLS 和血管内皮细胞等， 而在 OA 滑膜罕见阳性； RA 滑膜中 c-FLIP 表达显著高于 OA （t=11.717， P＜0.001）。细胞免疫荧光和 Western blot 结果显示， c-FLIP 主要定位于胞质， 且 RA FLS 中 c-FLIP 表达高于 OA FLS。FLS 与 CRT （0、 10、 50、 100 μg/L） 共培养， RA FLS 中 c-FLIP 表达上调且核因子 （NF） -κB 磷酸化增加， 具有浓度依赖性。NF-κB 抑制剂 BAY 11-7082 预干预后， CRT 上调 RA FLS 中 c-FLIP 表达的作用受到明显抑制。结论 RA FLS 中的 CRT 可能部分经 NF-κB 通路上调抗凋亡蛋白c-FLIP 的表达， 为今后RA 的临床治疗提供新的思路。

关键词: 关键词： 钙网织蛋白, 关节炎, 类风湿, NF-κB, c-FLIP

Abstract: Abstract: Objective To investigate the molecular mechanisms of upregulated expression of cellular Fas-associated death domain-like interleukin-1 beta converting enzyme inhibitory protein (c-FLIP) by calreticulin (CRT) in patients with rheumatoid arthritis (RA). Methods The semi-quantitative analysis and localization of c-FLIP in RA and osteoarthritis (OA) synovium were detected by immunohistochemistry. The fibroblast-like synoviocytes (FLS) were isolated by enzymatic digestion of synovial tissue specimens obtained from RA and OA patients, and cultured as an in vitro experiment model. The expressions of c-FLIP in RA and OA synovial fibroblasts were detected by immunofluorescence and Western blot assay. Whether CRT influenced c-FLIP expression and its molecular mechanism were explored by Western blot assay. Results The high expression of c-FLIP was found in RA synovium, mainly in the lining and sublining areas of FLS and vascular endothelial cells detected by immunohistochemistry. Meanwhile, weak staining of c-FLIP was observed in OA synovium. The expression of c-FLIP was significantly higher in RA synovium than that of OA synovium (t=11.717, P＜0.001). Results of immunofluorescence and Western blot assay showed that c-FLIP was mainly located in cytoplasm, and which was higher expressed in FLS of RA than that of OA. The increased c-FLIP expression and phosphorylation of NF- κB were detected after being co-incubated with exogenous CRT (0, 10, 50, 100 μg / L), in dose-dependent manner. The effect of CRT upregulating c-FLIP expression was blocked by NF-κB inhibitor BAY 11-7082. Conclusion CRT can increase c-FLIP expression at least partly through NF-κB pathway in RA, which may provide therapeutic target for the treatment of RA.

Key words: Key words: calreticulin, arthritis, rheumatoid, NF-kappa B, c-FLIP