ChAT/α7nAChR/核因子κB信号途径在类风湿关节炎发病中的免疫调控

Immunoregulatory mechanism of choline acetyltransferase/alpha 7 nicotinic acetylcholine receptor/nuclear factor-kappa B signaling pathway in the pathogenesis of rheumatoid arthritis

背景:胆碱能抗炎通路广泛参与类风湿关节炎发生发展,然而胆碱乙酰转移酶(choline acetyltransferase,ChAT)/烟碱型乙酰胆碱7受体(α7 nicotinic acetylcholine receptor,α7nAChR)/核因子κB信号途径在类风湿关节炎发病中的免疫调控机制研究未见报道。目的:探究类风湿关节炎发病中ChAT/α7nAChR/核因子κB信号途径的免疫调控机制。方法:取32只雌性Wistar大鼠,采用随机数字表法分为4组,每组8只:除健康对照组外,关节炎模型组、迷走神经切断组和假手术组均采用牛Ⅱ型胶原和弗氏完全/不完全佐剂构建关节炎大鼠模型,造模成功后,迷走神经切断组切断颈部左侧迷走神经,假手术组仅分离迷走神经。术后5周,检测各组大鼠体质量、关节炎评分和关节肿胀度、脾脏及关节病理变化,ELISA检测血清白细胞介素1、白细胞介素6、肿瘤坏死因子α水平,qRT-PCR及Western blot、免疫组化检测ChAT/α7nAChR/核因子κB通路核心基因mRNA及蛋白表达。结果与结论:①与健康对照组相比,关节炎模型组大鼠体质量下降(P<0.05),关节炎评分升高(P<0.01),踝关节肿胀度加重,关节间隙减小伴炎细胞浸润,脾脏白髓及生发中心增大增多,血清白细胞介素、白细胞介素6、肿瘤坏死因子α水平升高(P<0.05,P<0.01),关节中α7nAChR、ChAT、IκBα、核因子κB p50/p65的mRNA及蛋白表达升高(P<0.05,P<0.01),在关节面中显著表达;②与关节炎模型组和假手术组相比,迷走神经切断组大鼠体质量下降(P<0.05),关节炎评分升高(P<0.05,P<0.01),踝关节肿胀度加重伴畸形,关节间隙消失伴炎细胞浸润,脾脏白髓及生发中心增大融合,血清白细胞介素1、白细胞介素6、肿瘤坏死因子α水平升高(P<0.05,P<0.01),关节中α7nAChR、ChAT、IκBα、核因子κB p50/p65 mRNA及蛋白表达升高(P<0.05,P<0.01),在关节面中显著表达;③结果提示,迷走神经通过调控胆碱能抗炎通路刺激ChAT/α7nAChR/核因子κB信号途径,进而参与类风湿关节炎的疾病进程,提示胆碱能抗炎通路可能是治疗类风湿关节炎的潜在靶标。

BACKGROUND:The cholinergic anti-inflammatory pathway is widely involved in the development of rheumatoid arthritis.However,the immunoregulatory mechanism of choline acetyltransferase(ChAT)/α7 nicotinic acetylcholine receptor(α7nAChR)/nuclear factor(NF)-κB signaling pathway in the pathogenesis of rheumatoid arthritis has not been reported.OBJECTIVE:To investigate the immunoregulatory mechanism of ChAT/α7nAChR/NF-κB signaling pathway in the pathogenesis of rheumatoid arthritis.METHODS:Thirty-two female Wistar rats were randomly divided into healthy control group,arthritis model group,vagotomy group and sham operation group,with eight rats in each group.Except for the healthy control group,other groups were treated with bovine type-all collagen and Freund’s complete/incomplete adjuvant to construct the rat arthritis model.After successful molding,the vagus nerve was severed from the left vagus nerve in the neck in the vagotomy group,while only the vagus nerve was isolated in the sham operation group.At 5 weeks after surgery,the body mass,arthritis score,joint swelling degree,pathological changes of the spleen and joint were detected.The levels of interleukin-1,interleukin-6 and tumor necrosis factor-αin serum were detected by ELISA.The mRNA and protein expression levels of ChAT/α7nAChR/NF-κB pathway core genes were detected by qRT-PCR and western blot,respectively,and analyzed by immunohistochemistry.RESULTS AND CONCLUSION:Compared with the healthy control group,the body mass of rats in the arthritis model group was significantly decreased(P<0.05),the arthritis score was significantly increased(P<0.01),the ankle swelling degree was aggravated,the joint space was reduced with inflammatory cell infiltration,and the white pulp and germinal center of the spleen were enlarged and increased.Moreover,the levels of interleukin-1,interleukin-6 and tumor necrosis factor-αwere significantly increased,and the mRNA and protein expressions ofα7nAChR,ChAT,IκBα,and NF-κBp50/p65 on the joint surface were also significantly increased in the arthritis model group compared with the healthy control group(P<0.05,P<0.01).Compared with the arthritis model group and the sham operation group,the body mass of rats in the vagotomy group was significantly decreased(P<0.05),the arthritis score was significantly increased(P<0.05,P<0.01),the swelling degree of the ankle joint was aggravated with deformity,the joint space disappeared with inflammatory cell infiltration,the white pulp and germinal center of the spleen were enlarged and merged.Compared with the arthritis model group and the sham operation group,the levels of interleukin-1,interleukin-6 and tumor necrosis factor-αwere significantly increased in the vagotomy group(P<0.05,P<0.01),as well as the mRNA and protein expressions ofα7nAChR,ChAT,IκBα,and NF-κBp50/p65 were significantly increased on the joint surface(P<0.05,P<0.01).To conclude,the vagus nerve regulates the cholinergic anti-inflammatory pathway and stimulates the ChAT/α7nAChR/NF-κB signaling pathway to participate in the disease progression of rheumatoid arthritis.It indicates that the cholinergic anti-inflammatory pathway may be a potential target for the treatment of rheumatoid arthritis.