基于PI3K/AKT/NFATc1通路探讨针刀疏筋解结术抑制类风湿关节炎大鼠骨破坏的作用及机制  

Effect and mechanism of acupotomy tendon-sparing and knot-dissolving technique in inhibiting bone destruction in rats with rheumatoid arthritis based on the PI3K/AKT/NFATc1 pathway

作　　者：李浩林 陈平[2] 王海东[2] 李伟青[2] 金芳梅[2] 阚丽丽[2] 年芳红 胡乐乐 杨娟娟 景骆羊 LI Hao-lin;CHEN Ping;WANG Hai-dong;LI Wei-qing;JIN Fang-mei;KAN Li-li;NIAN Fang-hong;HU Le-le;YANG Juan-juan;JING Luo-yang(Gansu University of Traditional Chinese Medicine,Lanzhou 730000,China;Gansu Provincial Hospital of Traditional Chinese Medicine,Lanzhou 730050)

机构地区：[1]甘肃中医药大学,兰州730000 [2]甘肃省中医院,兰州730050

出　　处：《针刺研究》2025年第2期131-140,共10页Acupuncture Research

基　　金：国家中医药管理局高水平重点学科建设项目(No.国中医药人教函[2022]226);国家自然科学基金项目(No.82460945);甘肃省科技计划资助项目(No.24JRRA1044);兰州市指导性计划项目(No.2022-ZD-73)。

摘　　要：目的:观察针刀疏筋解结术对类风湿关节炎(RA)大鼠骨破坏及磷脂酰激醇3-激酶(PI3K)/蛋白激酶B(AKT)/活化T细胞核因子c1(NFATc1)通路的影响并探讨相关机制。方法:40只SD大鼠随机分为正常组、模型组、雷公藤组和针刀组,每组10只。采用尾根部注射牛Ⅱ型胶原乳剂建立胶原诱导性RA模型。针刀组予以针刀疏筋解结术治疗,1次/3 d,连续干预9次。雷公藤组予以雷公藤多苷混悬液(8 mg·kg^(-1))灌胃,1次/d,连续灌胃28 d。观察并评价大鼠踝关节肿胀度、关节炎指数评分;采用Micro-CT扫描观察各组大鼠左足踝关节骨破坏程度;HE染色和番红固绿染色分别观察滑膜组织和软骨组织病理形态变化;ELISA法检测滑膜组织肿瘤坏死因子(TNF)-α、白细胞介素(IL)-1β、IL-6、IL-17含量;抗酒石酸酸性磷酸酶(TRAP)染色观察左踝关节破骨细胞数量;Western blot法检测滑膜组织NFATc1、磷酸化(p)-PI3K、p-AKT蛋白的表达水平;免疫组织化学法检测踝关节滑膜组织基质金属蛋白酶-9(MMP9)、组织蛋白酶K(CTSK)、TNF受体关联因子6(TRAF6)的阳性表达。结果:模型组大鼠踝关节及足趾骨质侵蚀严重,表面凹凸不平;滑膜组织中有大量炎性细胞浸润,关节软骨损伤明显,滑膜细胞排列紊乱;软骨基质破坏,软骨层粗糙,软骨下骨结构紊乱。雷公藤组和针刀组以上组织病理变化均减轻。与正常组比较,模型组大鼠的关节肿胀度、关节炎指数,骨表面积和骨体积的比值(BS/BV)、骨小梁数量(Tb.N)、骨小梁分离度(Tb.Sp),滑膜组织中TNF-α、IL-1β、IL-6、IL-17含量,滑膜组织中NFATc1蛋白表达水平及p-PI3K/PI3K、p-AKT/AKT比值,滑膜组织TRAF6、CTSK、MMP9阳性表达均显著升高(P<0.01),骨密度(BMD)、骨体积分数(BV/TV)、骨小梁厚度(Tb.Th)显著降低(P<0.01),关节内破骨细胞数量增加(P<0.01)。与模型组比较,雷公藤组和针刀组大鼠关节肿胀度、关节炎指数,BS/BV、Tb.Sp,滑膜组�Objective To observe the effect of acupotomy tendon-sparing and knot-dissolving technique on bone destruction and phosphatidylinositol 3-kinase(PI3K)/protein kinase B(AKT)/nuclear factor of activated T cells c1(NFATc1)pathway in rats with rheumatoid arthritis(RA),so as to investigate the underlying mechanism.Methods Forty SD rats were randomly divided into normal,model,medication(tripterygium wilfordii),and acupotomy groups,with 10 rats in each group.Except for the normal group,the rest of the rats were injected with bovine type II collagen emulsion at the base of tails to establish a collagen-induced RA model.The acupotomy group was treated with acupotomy tendon-sparing and knot-dissolving technique,once every 3 days,with a continuous intervention for 9 times.The medication group was given tripterygium wilfordii polyglycoside suspension(8 mg·kg^(-1))by gavage,once a day for 28 days continuously.The swelling degree of the ankle joint and the arthritis index score of the rats were observed.Micro-CT scanning was used to observe the degree of bone destruction in the left ankle joint.HE staining and ferruben-solid green staining were used to observe the pathological morphological changes of synovial and cartilage tissue respectively.ELISA was used to detect the contents of tumor necrosis factor(TNF)-α,interleukin(IL)-1β,IL-6 and IL-17 in synovial tissue.Tartrate-resistant acid phosphatase(TRAP)staining was used to observe the number of osteoclasts in the left ankle joint.Western blot was used to detect the expression levels of NFATc1,p-PI3K and p-AKT proteins in synovial tissue.Immunohistochemistry was used to detect the positive expressions of matrix metalloproteinase-9(MMP9),cathepsin K(CTSK)and TNF receptor-associated factor 6(TRAF6)in the synovial tissue of the ankle joint.Results In comparison with the normal group,the bones of the ankle joint and toes of rats were severely eroded,with an uneven surface in the model group;there was a large number of inflammatory cell infiltrations in the synovial tissue,obvio

关 键 词：针刀疏筋解结术 类风湿关节炎 骨破坏 PI3K/AKT/NFATc1通路 破骨细胞 

分 类 号：R245[医药卫生—针灸推拿学]