类风湿关节炎患者外周血单个核细胞中c-FLIP与外源性凋亡途径的相关性分析

目的: 分析类风湿关节炎患者外周血单个核细胞（PBMC）中Fas相关死亡区样白介素1转换酶抑制蛋白（c-FLIP）与外源性凋亡途径相关蛋白死亡结构域蛋白（FADD）、caspase-8表达水平变化的相关性。方法: 选择2014年1月至2015年6月福建医科大学附属漳州市医院类风湿关节炎患者60例，按照类风湿关节炎疾病活动指数将患者分为低、中、高活动度组（分别为22、20和18例）。同时选择健康体检者25名作为健康对照组。采用实时定量RT-PCR和蛋白质印迹法分别检测PBMC中c-FLIP、FADD和caspase-8的mRNA和蛋白表达水平，并采用Pearson检验对c-FLIP与FADD、caspase-8表达的相关性进行分析。结果: 类风湿关节炎患者PBMC中c-FLIP、FADD和caspase-8的mRNA表达量均高于健康对照组（均P < 0.05）。其中，中活动度组PBMC中FADD和caspase-8的mRNA表达量高于低活动度组（均P < 0.05），c-FLIP的mRNA表达量与低活动度组相近（P>0.05）；高活动度组PBMC中c-FLIP的mRNA表达量高于中、低活动度组（均P < 0.05），caspase-8的mRNA表达量低于中、低活动度组（均P < 0.05），FADD的mRNA表达量高于低活动度组（P < 0.05）。类风湿关节炎患者PBMC中c-FLIP mRNA水平与FADD mRNA水平呈正相关（r=0.323，P < 0.05），与caspase-8 mRNA水平呈负相关（r=-1.104，P < 0.05）。中活动度组c-FLIP和FADD蛋白表达量高于健康对照组、低活动度组和高活动度组（P < 0.05或P < 0.01）；中、高活动度组caspase-8蛋白表达量高于健康对照组和低活动度组（P < 0.05或P < 0.01），且高活动度组caspase-8蛋白表达量高于中活动度组（P < 0.05）。结论: 类风湿关节炎患者PBMC中凋亡相关蛋白c-FLIP可能参与了外源性凋亡途径，并可为类风湿关节炎病情评估提供参考。

Expression of c-FLIP in peripheral blood mononuclear cells of patients with rheumatoid arthritis and its relation with extrinsic apoptotic pathway

Objective: To investigate the expression of apoptosis related protein cellular Fas associated death domain like interleukin 1 converting enzyme inhibitory protein (c-FLIP) in peripheral blood mononuclear cells (PBMCs) of patients with rheumatoid arthritis and its relation with extrinsic apoptotic pathway. Methods: Sixty patients with rheumatoid arthritis were collected from Zhangzhou Affiliated Hospital of Fujian Medical University during January 2014 and June 2015, including 22 patients with low activities (DAS28 < 3.2), 20 patients with middle activities (3.2 ≤ DAS28 ≤ 5.1), and 18 patients with high activities (DAS28>5.1). And 25 healthy controls were also collected. The mRNA and protein expression levels of c-FLIP and the extrinsic apoptotic pathway related proteins Fas-associated protein with death domain (FADD), caspase-8 in PBMCs were detected by real-time RT-PCR and Western blot, respectively. Correlations between c-FLIP and FADD, caspase-8 in PBMCs were analyzed by pearson test. Results: mRNA expression levels of c-FLIP, FADD and caspase-8 in PBMCs of patients with rheumatoid arthritis were all higher than those of healthy controls (all P < 0.05). mRNA expression levels of FADD and caspase-8 in patients with middle activities were significantly higher than those in patients with low activities (all P < 0.05), but the mRNA expression level of c-FLIP was not significantly higher than that in patients with low activities. mRNA expression level of c-FLIP in patients with high activities was higher than those in patients with middle or low activities (all P < 0.05), while the mRNA expression level of caspase-8 was lower than those in patients with middle or low activities (all P < 0.05). mRNA expression level of FADD in patients with high activities was higher than those in patients with low activities (P < 0.05). Pearson analysis showed that there was a positive correlation between c-FLIP and FADD mRNA expression (r=0.323, P < 0.05), and negative correlation between c-FLIP and caspase-8 mRNA expression (r=-1.104, P < 0.05). The protein expression levels of c-FLIP and FADD in patients with middle activities were significantly higher than those in control group and patients with low or high activities (P < 0.05 or 0.01). The protein expression levels of caspase-8 in patients with middle and high activities were significantly higher than those in control group and patients with low activities (P < 0.05 or P < 0.01), and the protein expression level of caspase-8 in patients with high activities was higher than that in patients with middle activities (P < 0.05). Conclusion: c-FLIP may be involved in the extrinsic apoptotic pathway in rheumatoid arthritis, and can provide reference for the evaluation of disease activities.