加味补肾壮筋汤对兔膝骨关节炎Wnt/β-catenin信号通路的影响  

Effect of Modified Bushen Zhuangjin Decoction on Wnt/β-catenin Signaling Pathway in Rabbit Knee Osteoarthritis

作　　者：马金昕 程韶[1,2,3] 李锦锦 苏心哲 张北 胡昆鹏 殷志颖 朱云麒 王上增 MA Jinxin;CHENG Shao;LI Jinjin;SU Xinzhe;ZHANG Bei;HU Kunpeng;YIN Zhiying;ZHU Yunqi;WANG Shangzeng(Henan University of Chinese Medicine,Zhengzhou Henan China 450046;Henan Provincial Hospital of Chinese Medicine/The Second Affiliated Hospital to Henan University of Chinese Medicine,Zhengzhou Henan China 450002;Henan Provincial Engineering Research Center,Zhengzhou Henan China 450002)

机构地区：[1]河南中医药大学,河南郑州450046 [2]河南省中医院/河南中医药大学第二附属医院,河南郑州450002 [3]河南省工程研究中心,河南郑州450002

出　　处：《中医学报》2024年第11期2432-2439,共8页Acta Chinese Medicine

基　　金：国家自然科学基金面上项目(82374490);河南省自然科学基金项目(222300420486,232300420269);河南省中医药科学研究专项课题项目(2021ZY2010,2023ZYZD06,2022ZY2026);河南省高校科技创新团队支持计划资助项目(24IRTSTHN040)。

摘　　要：目的:基于Wnt/β-catenin信号通路探讨加味补肾壮筋汤(modified Bushen Zhuangjin decoction,MBZD)治疗兔膝骨关节炎(knee osteoarthritis,KOA)的作用机制。方法:从36只大耳白兔中随机选取8只作为对照组(Control),其余白兔在右膝关节注射木瓜蛋白酶以建立KOA模型。将造模成功的白兔随机分为模型组(KOA)、加味补肾壮筋汤组(MBZD,3.29 g·kg^(-1))及塞来昔布组(celecoxib,CELE,9.33 mg·kg^(-1)),每组8只。MBZD和CELE组白兔按照相应剂量灌胃,Control和KOA组白兔给予等体积生理盐水,每日1次,连续28 d。取右侧膝关节进行Pelletier评分,HE染色观察软骨与滑膜组织病理形态,免疫组织化学法(immunohistochemistry,IHC)检测软骨与滑膜组织中基质金属蛋白酶-13(matrix metalloproteinase-13,MMP-13)、Ⅱ型胶原α1(collagen typeⅡalpha 1,COL2A1)表达水平,Western blot法检测软骨组织中白细胞介素-1β(interleukin 1β,IL-1β)、MMP-13、COL2A1、Wnt-3a、β-catenin、C-MYC蛋白表达水平,RT-qPCR法检测软骨组织中Wnt-3αmRNA、β-catenin mRNA、C-MYC mRNA表达水平。结果:与Control组比较,KOA组白兔Pelletier评分升高(P<0.05);与KOA组比较,MBZD组和CELE组白兔Pelletier评分降低(P<0.05)。HE染色显示,Control组白兔软骨和滑膜组织形态无明显异常。KOA组白兔软骨严重缺损,滑膜组织异常增生伴炎症细胞浸润。MBZD组和CELE组白兔软骨和滑膜组织损伤明显改善。IHC显示,与Control组比较,KOA组白兔软骨和滑膜组织MMP-13表达水平升高、COL2A1表达水平降低(P<0.05),经MBZD和CELE治疗后得到逆转(P<0.05)。Western blot显示,与Control组比较,KOA组白兔软骨组织IL-1β、MMP-13、Wnt-3α、β-catenin、C-MYC蛋白表达水平升高,COL2A1蛋白表达水平降低(P<0.05),经MBZD和CELE治疗后得到逆转(P<0.05)。RT-qPCR显示,与Control组比较,KOA组白兔软骨组织Wnt-3αmRNA、β-catenin mRNA、C-MYC mRNA水平升高,经MBZD和CELE治疗后得到逆转(P<0.05)。结论:MBZD可改善KObjective:Based on Wnt/β-catenin signaling pathway,to explore the mechanism of modified Bushen Zhuangjin Decoction in treatment of knee osteoarthritis(KOA)in rabbits.Methods:Thirty-six big-eared white rabbits were randomly divided into a control group(Control,8)and a model group(KOA,28),and the latter was injected with papain in the right knee joint in order to establish a KOA model.The successful modeled white rabbits were randomly divided into the model group,the modified Bushen Zhuangjin Decoction group(MBZD,3.29 g·kg^(-1))and celecoxib(CELE,9.33 mg·kg^(-1))group,with 8 rabbits in each group.The rabbits in the MBZD and CELE groups were gavaged according to the corresponding dosage,and the Control and KOA groups were given an equal volume of saline once a day for 28 d.The right knee joints were scored by Pelletier′s score,and the pathological morphology of cartilage and synovial tissue was observed by HE staining,and matrix metalloproteinase-13(matrix metalloproteinase-13)was detected in the cartilage and synovial tissue by immunohistochemistry(IHC).The expression levels of matrix metalloproteinase 13(MMP-13)and collagen type II alpha 1(COL2A1)in cartilage and synovial tissues were detected by immunohistochemistry(IHC),and the expression levels of interleukin 1β(IL-1β),MMP-13,COL2A1,MMP-1β,COL2A1,and MMP-1βin cartilage tissues were detected by Western blot.13,COL2A1,Wnt-3a,β-catenin,and C-MYC protein expression levels in cartilage tissues,and Wnt-3αmRNA,β-catenin mRNA,and C-MYC mRNA expression levels in cartilage tissues by RT-qPCR.Results:The Pelletier score of white rabbits in the KOA group was elevated compared with that of the Control group(P<0.05);and the Pelletier score of white rabbits in the MBZD and CELE groups was decreased compared with that of the KOA group(P<0.05).HE staining showed that there were no obvious abnormalities in the morphology of cartilage and synovial tissues of white rabbits in the Control group,and that white rabbits in the KOA group showed severe defects in cartilage a

关 键 词：膝骨关节炎 加味补肾壮筋汤 WNT/Β-CATENIN信号通路 软骨细胞 滑膜组织 白兔 

分 类 号：R274.9[医药卫生—中西医结合] R285.5[医药卫生—中医骨伤科学]