摘要
目的:观察热补针法干预寒湿证类风湿关节炎(RA)家兔“足三里”的疗效及对膝关节滑膜组织中线粒体质量和动力蛋白相关蛋白1(Drp1)/线粒体分裂因子(Mff)通路的影响。方法:将24只新西兰白兔随机分为空白组、模型组、热补针法组、抑制剂组,每组各6只,采用卵蛋白完全弗氏佐剂混合液联合人工气候箱复制寒湿证RA家兔模型。予热补针法组家兔双侧“足三里”热补针法干预,1次/d,共14 d;予抑制剂组家兔腹腔注射线粒体分裂抑制剂Mdivi-1(2.5 mg/kg),1次/2 d,共7次。空白组与模型组采取相同的抓取和固定,不进行其他干预。在造模后及干预后评估家兔膝关节关节炎评分、痛阈及关节周径。采用HE染色观察膝关节滑膜组织病理学变化,透射电镜观察滑膜细胞线粒体超微结构,免疫荧光染色法检测滑膜组织线粒体活性氧(mtROS)表达,荧光素酶法检测滑膜组织ATP含量,免疫荧光染色法观察滑膜组织Drp1与Mff共定位水平,Western blot法检测滑膜组织磷酸化(p)-Drp1 s616/Drp1、Mff蛋白表达。结果:与空白组相比,模型组家兔关节炎评分、膝关节周径、膝关节滑膜组织中mtROS表达、p-Drp1 s616/Drp1比值及Mff蛋白表达、Drp1与Mff共定位水平升高(P<0.01),痛阈及膝关节滑膜组织ATP含量明显降低(P<0.01)。与模型组相比,热补针法组和抑制剂组家兔关节炎评分、膝关节周径、滑膜组织中mtROS表达、p-Drp1 s616/Drp1比值及Mff蛋白表达、Drp1与Mff共定位水平明显降低(P<0.01),痛阈、滑膜组织ATP含量明显升高(P<0.01)。模型组滑膜组织滑膜细胞增殖明显,滑膜内可见炎性细胞浸润及增生的滑膜基质,滑膜细胞中线粒体嵴模糊,线粒体肿胀破裂,线粒体形成分裂为小体;热补针法组及抑制剂组滑膜组织中上述改变均减轻。结论:热补针法可以显著改善寒湿证RA家兔症状,提高滑膜细胞线粒体质量,可能与抑制Drp1/Mff通路活性有关。
Objective To observe the efficacy of heat-reinforcing acupuncture at“Zusanli”(ST36)in rabbits with cold-dampness rheumatoid arthritis(RA)and its impact on mitochondrial quality and the dynamin-related protein 1(Drp1)/mitochondrial fission factor(Mff)pathway in knee synovial tissue.Methods Twenty-four New Zealand white rabbits were randomly divided into a normal group,a model group,a heat-reinforcing acupuncture group,and an inhibitor group,with 6 rabbits in each group.A cold-dampness RA rabbit model was established using complete Freund’s adjuvant-ovalbumin mixture combined with an artificial climate chamber.Rabbits of the heat-reinforcing acupuncture group received heat-reinforcing acupuncture at ST36 once daily for 14 d.Rabbits of the inhibitor group was intraperitoneally injected with the mitochondrial division inhibitor Mdivi-1(2.5 mg/kg)once every 2 d for a total of 7times.Rabbits of the normal and model groups were handled and restrained in the same manner but received no other interventions.Knee arthritis scores,mechanical pain thresholds,and joint circumference were assessed after modeling and intervention.Hematoxylin-eosin(HE)staining was used to observe pathological changes in knee synovial tissue.Transmission electron microscopy was used to observe the ultrastructure of mitochondria in synovial cells.Luciferase method was used to measure ATP content in knee synovial tissue.Immunofluorescence staining was used to detect mitochondrial reactive oxygen species(mtROS)expression and Drp1/Mff co-localization in synovial tissue,and W estern blot was used to measure the relative expression levels of phosphorylated(p)-Drp1 s616/Drp1 and Mff protein.Results Compared with the normal group,the model group showed increased arthritis scores,joint circumference,mtROS expression,p-Drp1 s616/Drp1 and Mff protein expression,and Drp1/Mff co-localization in knee synovial tissue(P<0.01),while mechanical pain thresholds and ATP content in synovial tissue were significantly decreased(P<0.01).Following acupuncture intervention,compared with the model group,the heat-reinforcing acupuncture group and the inhibitor group showed significantly reduced arthritis scores,joint circumference,mtROS expression,p-Drp1 s616/Drp1 and Mff protein expression,and Drp1/Mff co-localization in synovial tissue(P<0.01),while mechanical pain thresholds and ATP content in synovial tissue were significantly increased(P<0.01).In the model group,synovial tissue showed significant synovial cell proliferation,with visible inflammatory cell infiltration and synovial matrix proliferation in the synovium;synovial cells had indistinct mitochondrial cristae,mitochondrial swelling/rupture,and mitochondrial fragmentation into small bodies.These changes in synovial tissue were alleviated in both the heat-reinforcing acupuncture group and the inhibitor group.Conclusion Heat-reinforcing acupuncture can significantly improve symptoms in RA rabbits with cold-dampness and enhance mitochondrial quality in synovial cells,which may be related to the inhibition of Drp1/Mff pathway activity.
作者
井维尧
许从荟
刘翠
刘莉梅
袁博
张星华
张枫帆
陈平
杜小正
JING Wei-yao;XU Cong-hui;LIU Cui;LIU Li-mei;YUAN Bo;ZHANG Xing-hua;ZHANG Feng-fan;CHEN Ping;DU Xiao-zheng(School of Acupuncture-Moxibustion and Tuina,Gansu University of Chinese Medicine,Lanzhou 730000,China;Acupuncture Center,Affiliated Hospital of Gansu University of Chinese Medicine,Lanzhou 730000;Department of Acupuncture and Moxibustion,Gansu Provincial Hospital of Traditional Chinese Medicine,Lanzhou 730050;Department of Rheumatology and Orthopedics,Gansu Provincial Hospital of Traditional Chinese Medicine,Lanzhou 730050)
出处
《针刺研究》
北大核心
2025年第10期1124-1132,共9页
Acupuncture Research
基金
国家自然科学基金项目(No.82060891)
兰州市科技发展指导性项目(No.2023-ZD-207)
甘肃省针灸推拿临床医学中心项目(No.ZJKF2203)。
关键词
类风湿关节炎
热补针法
线粒体质量
Drp1/Mff通路
Rheumatoid arthritis
Heat-reinforcing acupuncture
Mitochondrial quality
Dynamin-related protein 1/mitochondrial fission factor pathway
