摘要
Objective:To observe the effects of moxibustion at Shenshu(BL23)and Zusanli(ST36)on Toll-like receptor 4(TLR4)-myeloid differentiation factor 88(Myd88)signaling pathway-mediated inflammatory factors in the synovial tissue of ankle joints of rats with rheumatoid arthritis(RA),and to explore the molecular and biological mechanisms underlying the anti-inflammatory and analgesic effects.Methods:A total of 24 male Sprague-Dawley(SD)rats were randomly divided into a normal group,a model group,and a moxibustion group,with 8 rats in each group.The RA model was established with exposure to wind,cold,and damp environmental factors,along with Freund’s complete adjuvant.After three days of modeling,mild moxibustion was applied to bilateral Shenshu(BL23)and Zusanli(ST36)in the moxibustion group using moxa sticks of 0.9 cm in diameter for 30 min each time,once a day for 14 d.Structural changes in the synovial tissue and cells were then observed using hematoxylin-eosin staining and transmission electron microscopy,while immunohistochemistry analysis was used to detect tumor necrosis factor(TNF)-α,interleukin(IL)-17,IL-1β,and IL-6 levels.Moreover,the protein expression levels of Myd88,TLR4,and transient potential receptor vanilloid type 1(TRPV1)in the synovial tissue were detected using Western blotting,while their mRNA expression levels were detected using reverse transcription-polymerase chain reaction.Finally,the levels of IL-1β,IL-2,IL-6,IL-17A,and TNF-αin rat serum were detected using enzyme-linked immunosorbent assay.Results:Compared to the normal group,the model group exhibited notable pathological synovial tissue damage,along with significantly higher IL-1β,IL-6,and TNF-αlevels(P<0.01)and a slightly higher IL-17 content(P>0.05).Furthermore,the Myd88,TLR4,and TRPV1 protein and mRNA expression levels and serum IL-1β,IL-2,IL-6,IL-17A,and TNF-αlevels were all significantly higher in the model group than in the normal group(P<0.01).Compared to the model group,the moxibustion group exhibited a lower degree of synovial tissue pathological damage,along with significantly lower IL-1β,IL-6,and TNF-αlevels(P<0.05 or P<0.01)and a lower IL-17 content without statistical significance(P>0.05).Moreover,the Myd88,TLR4,and TRPV1 protein and mRNA expression levels,and serum IL-1β,IL-2,IL-6,IL-17A,and TNF-αlevels were all significantly lower in the moxibustion group than in the model group(P<0.01 or P<0.05).Conclusion:Mild moxibustion at Shenshu(BL23)and Zusanli(ST36)can effectively inhibit TLR4-MyD88 signaling pathway-mediated inflammatory factor expression in the synovial tissue of ankle joints of RA rats.Furthermore,the effect of moxibustion on synovial tissue inflammation in RA rats may be attributed to TRPV1 channel activation.
目的:观察艾灸肾俞和足三里对类风湿性关节炎(RA)模型大鼠踝关节滑膜组织Toll样受体4(TLR4)-髓样分化因子88(Myd88)信号通路介导的炎症因子的影响,探讨艾灸抗炎镇痛效应的分子生物学机制。方法:将24只雄性SD大鼠随机分为正常组、模型组和艾灸组,每组8只。采用风、寒、湿环境因素+弗氏完全佐剂进行RA模型复制。造模3 d后,艾灸组采用直径0.9 cm艾条对大鼠双侧肾俞和足三里施行温和灸,每次30 min,每日1次,连续14 d。苏木素-伊红染色和透射电镜观察大鼠滑膜组织和细胞结构改变,免疫组织化学法检测大鼠滑膜组织中肿瘤坏死因子(TNF)-α、白介素(IL)-17、IL-1β及IL-6含量,免疫印迹法检测滑膜组织中Myd88、TLR4、瞬时电位感受器香草醛受体1(TRPV1)蛋白表达,逆转录聚合酶链反应检测滑膜组织中TLR4、Myd88及TRPV1 mRNA表达,酶联免疫吸附测定检测大鼠血清中IL-1β、IL-2、IL-6、IL-17A及TNF-α含量。结果:相较于正常组,模型组滑膜组织病理损伤明显,滑膜组织中IL-1β、IL-6、TNF-α含量显著升高(P<0.01),IL-17含量轻微升高(P>0.05);Myd88、TLR4、TRPV1蛋白及其mRNA表达水平均显著升高(P<0.01),血清IL-1β、IL-2、IL-6、IL-17A及TNF-α含量均明显升高(P<0.01)。干预后,与模型组比较,艾灸组滑膜组织病理损伤程度降低,滑膜组织中IL-1β、IL-6及TNF-α含量均显著降低(P<0.05或P<0.01),IL-17含量降低,但差异无统计学意义(P>0.05);Myd88、TLR4、TRPV1蛋白及其mRNA表达水平均显著降低(P<0.01或P<0.05),血清IL-1β、IL-2、IL-6、IL-17A和TNF-α含量均明显降低(P<0.01)。结论:艾灸肾俞、足三里可有效抑制RA大鼠踝关节滑膜组织TLR4-MyD88信号通路介导的炎症因子;其对RA模型大鼠滑膜炎症和疼痛的调节作用可能与激活TRPV1通道有关。
基金
国家自然科学基金资助项目,No.82205289
安徽省高等学校自然科学研究重点项目,No.2022AH050526,No.2024AH050915
安徽省自然科学基金青年项目,No.2208085QH265
安徽省高校哲学社会科学研究重点项目,No.2023AH050710
安徽省高校科研计划项目自然科学重点项目,No.2023AH050811。
