文献类型：期刊文献

中文题名：基于自噬-NLRP3炎症小体-IL-1β信号轴探究热补针法对类风湿关节炎寒证模型家兔滑膜炎症的影响

英文题名：Effect of Heat-reinforcing Acupuncture on Synovial Inflammatory in Rheumatoid Arthritis Rabbits with Cold Syndrome Based on Autophagy-NLRP3 Inflammasome-IL-1βSignaling Axis

作者：刘强[1];苏成红[1];秦昕[2];井维尧[1];刘莉梅[1];刘翠[1];杜小正[1];袁博[3];张星华[4];陈平[4];李向军[4];张枫帆[4]

第一作者：刘强

机构：[1]甘肃中医药大学针灸推拿学院,甘肃兰州730000;[2]甘肃省第二人民医院,甘肃兰州730000;[3]甘肃中医药大学附属医院,甘肃兰州730000;[4]甘肃省中医院,甘肃兰州730050

第一机构：甘肃中医药大学针灸推拿学院

年份：2023

卷号：30

期号：12

起止页码：102

中文期刊名：中国中医药信息杂志

外文期刊名：Chinese Journal of Information on Traditional Chinese Medicine

收录：CSTPCD;;CSCD:【CSCD_E2023_2024】；

基金：国家中医药管理局甘肃郑氏针法学术流派传承工作室(2305135901);国家自然科学基金(82060891);甘肃省自然科学基金(21JR7RA568、20JR10RA344)。

语种：中文

中文关键词：类风湿关节炎;热补针法;滑膜炎症;自噬;NLRP3炎症小体;白细胞介素-1β;兔

外文关键词：rheumatoid arthritis;heat-reinforcing acupuncture;synovial inflammation;autophagy;NLRP3 inflammasome;IL-1β;rabbits

摘要：目的观察热补针法对类风湿关节炎(RA)寒证模型家兔滑膜组织自噬-NOD样受体蛋白3(NLRP3)炎症小体-白细胞介素(IL)-1β信号轴的影响,探讨热补针法治疗RA滑膜炎症的作用机制。方法48只新西兰家兔随机分为正常组、模型组、3-甲基腺嘌呤(3-MA)组、雷帕霉素组、热补针法组及平补平泻组,每组8只。以弗氏完全佐剂+卵蛋白诱导联合低温冷冻法复制RA寒证家兔模型。热补针法组和平补平泻组于“足三里”分别施热补针法及平补平泻针法针刺,留针30 min,1次/d,连续14 d;3-MA组和雷帕霉素组分别耳缘静脉注射3-MA和雷帕霉素溶液,1次/2 d,共7次。免疫组化染色检测膝关节滑膜组织IL-1β表达,免疫荧光染色检测滑膜组织NLRP3表达,透射电镜观察滑膜细胞自噬小体形态,Western blot检测滑膜组织自噬蛋白5(Atg5)、UNC-51样激酶1(ULK1)、微管相关蛋白轻链3(LC3)、Beclin-1蛋白表达。结果与正常组比较,模型组家兔膝关节周径显著增加、皮温显著降低(P<0.05),滑膜组织IL-1β、NLRP3表达显著升高(P<0.05),Atg5、ULK1、LC3Ⅱ/LC3Ⅰ、Beclin-1蛋白表达显著降低(P<0.05)。与模型组比较,3-MA组家兔膝关节周径显著增加、皮温显著降低(P<0.05),滑膜组织IL-1β、NLRP3表达显著升高(P<0.05),Atg5、ULK1、LC3Ⅱ/LC3Ⅰ、Beclin-1蛋白表达显著降低(P<0.05);热补针法组、平补平泻组和雷帕霉素组家兔膝关节周径显著缩小、皮温显著升高(P<0.05),滑膜组织IL-1β、NLRP3表达显著降低(P<0.05),Atg5、ULK1、LC3Ⅱ/LC3Ⅰ、Beclin-1蛋白表达显著升高(P<0.05),且热补针法组作用优于平补平泻组。透射电镜观察显示,正常组家兔滑膜细胞胞浆内自噬小体、溶酶体散在分布;模型组和3-MA组家兔滑膜细胞偶见自噬小体分布;雷帕霉素组和热补针法组家兔滑膜细胞可见吞噬泡及双层膜结构的自噬小体,溶酶体数量增加;平补平泻组家兔滑膜细胞偶见吞噬泡、自噬小体及溶酶体等结构。结论热补针法能抑制RA寒证模型家兔膝关节滑膜炎症反应,其机制可能与调节自噬-NLRP3炎症小体-IL-1β信号轴有关。
Objective To observe the effects of heat-reinforcing acupuncture in the treatment of rheumatoid arthritis(RA)rabbits with cold syndrome from the perspective of autophagy-NLRP3 inflammasome-IL-1βsignaling axis;To explore the mechanism of heat-reinforcing acupuncture in the treatment of synovial inflammation in RA.Methods Totally 48 New Zealand rabbits were randomly divided into normal group,model group,3-methyladenine(3-MA)group,rapamycin group,heat-reinforcing acupuncture group,and reinforcing-reducing group,with 8 rats in each group.The rabbit model of RA with cold syndrome was replicated by a combination of Freund’s complete adjuvant,egg albumin induction and hypothermic freezing method.The heat-reinforcing acupuncture group and reinforcing-reducing group were treated with“Zusanli”heat-reinforcing acupuncture and reinforcing-reducing accupuncture respectively,with needle retention for 30 minutes,once a day,and intervention for 14 times;3-MA and rapamycin were injected intravenously into the ear margin once every other day for 7 times in the 3-MA group and rapamycin group respectively.The IL-1βexpression in synovial tissue was detected by immunohistochemistry,and the NLRP3 expression was examined by immunofluorescence,the autophagy of synovial tissue was observed by transmission electron microscopy,Western blot was used to detect Atg5,ULK1,LC3 and Beclin-1 protein expression.Results Compared with the normal group,the circumference of knee joint significantly increased and skin temperature of knee joint significantly decreased in model group(P<0.05),the expression of IL-1βand NLRP3 in synovial tissue significantly increased(P<0.05),the protein expressions of Atg5,ULK1,LC3Ⅱ/LC3Ⅰ,Beclin-1 significantly decreased(P<0.05).Compared with the model group,the circumference of knee joint significantly increased and skin temperature of knee joint significantly decreased in 3-MA group(P<0.05),the expression of IL-1βand NLRP3 in synovial tissue significantly increased(P<0.05),the protein expressions of Atg5,ULK1,LC3Ⅱ/LC3Ⅰ,and Beclin-1 significantly decreased(P<0.05);the circumference of knee joint significantly reduced and skin temperature of knee joint significantly increased in the heat-reinforcing acupuncture group,reinforcing-reducing group and rapamycin group(P<0.05),the expression of IL-1βand NLRP3 in synovial tissue significantly decreased(P<0.05),the protein expressions of Atg5,ULK1,LC3Ⅱ/LC3Ⅰand Beclin-1 significantly increased(P<0.05),and the action of heat-reinforcing acupuncture group was better than that of the reinforcing-reducing group.Transmission electron microscopy observation showed that autophagosomes and lysosomes were scattered in the cytoplasm in synovial cells of normal group was regular;the distribution of autophagosomes was occasionally observed in synovial cells of rabbits in the model group and 3-MA group;phagocytic vesicles and autophagosomes with double-layer membrane structure could be seen in the synovial cells of rabbits in the rapamycin group and the heat-reinforcing acupuncture group,and the number of lysosomes increased;occasional structures such as phagocytic vesicles,autophagosomes,and lysosomes were observed in synovial cells of rabbits in the reinforcing-reducing group.Conclusion Heat-reinforcing acupuncture can effectively reduce knee swelling and inflammatory infiltration in RA with cold syndrome model rabbits.The possible mechanism is related to the regulation of autophagy-NLRP3 inflammasome-IL-1βsignaling axis.