苗药验方通过Wnt/β-catenin信号通路减轻家兔骨关节炎软骨细胞的损伤  

Effect of Miao Medicine Prescription on Chondrocytes of Osteoarthritis Rabbits Through Wnt/β-catenin Signaling Pathway

作　　者：付钰[1] 古丽玲 罗昌禄[1] 吴晓勇 李宝杰 夏景富 Fu Yu;Gu Liling;Luo Changlu(Department of Rehabilitation Medicine,Guizhou Provincial People’s Hospital,Guiyang 550002,China)

机构地区：[1]贵州省人民医院康复医学科,贵阳550002 [2]贵州中医药大学第一附属医院风湿血液科,贵阳550001 [3]贵州中医药大学第一附属医院苗医学科,贵阳550001

出　　处：《华中科技大学学报（医学版）》2023年第1期48-53,59,共7页Acta Medicinae Universitatis Scientiae et Technologiae Huazhong

基　　金：贵州省科技计划项目(No.黔科合基础[2017]1112)。

摘　　要：目的观察苗药验方对家兔骨关节炎模型软骨细胞的影响,并探讨其防治骨关节炎软骨退变的作用机制。方法36只家兔随机分为正常对照组、模型组、苗药熏蒸组、苗药离子导入组,每组9只。除正常对照组外,其余3组家兔右膝关节腔内注射木瓜蛋白酶制造膝骨性关节炎模型。造模成功后,苗药熏蒸组与苗药离子导入组家兔分别经苗药验方熏蒸治疗及苗药验方离子导入治疗20 d,取各组家兔软骨组织行苏木精-伊红染色,并取各组家兔血清备用。取正常家兔软骨组织培养软骨细胞,将软骨细胞也分为4组:正常对照组、模型组、苗药薰蒸组、苗药离子导入组。除正常对照组外,其余3组软骨细胞用10μg/L白细胞介素-1β(IL-1β)处理24 h制备骨关节炎软骨细胞模型,造模期间分别用相应组家兔血清对软骨细胞进行体外干预,酶联免疫吸附法(ELISA)检测软骨细胞上清液中β-连环蛋白(β-catenin)、肿瘤坏死因子-α(TNF-α)的含量,免疫印迹(Western blot)法检测软骨细胞中骨形态发生蛋白-2(BMP-2)、基质金属蛋白酶-3(MMP-3)、基质金属蛋白酶-13(MMP-13)、细胞外因子-16(Wnt-16)、β-catenin蛋白表达水平,实时荧光定量聚合酶链反应(qPCR)检测软骨细胞中BMP-2、MMP-3、MMP-13、Wnt-16、β-catenin mRNA表达水平。结果与模型组比较,苗药熏蒸组、苗药离子导入组家兔关节软骨表面纤维化及其软骨细胞结构破坏均得到不同程度改善。与模型组比较,苗药薰蒸组软骨细胞上清液TNF-α、β-catenin的含量显著减少(均P<0.05),软骨细胞BMP-2、MMP-3、β-catenin蛋白表达以及BMP-2、Wnt-16 mRNA表达水平显著下调(均P<0.05);与模型组比较,苗药离子导入组软骨细胞上清液中TNF-α、β-catenin的含量显著减少(均P<0.05),软骨细胞BMP-2、MMP-3、MMP-13、β-catenin蛋白表达水平以及BMP-2、MMP-3、Wnt-16 mRNA表达水平显著下调(均P<0.05)。结论苗药验方薰蒸疗法及�Objective To observe the effect of Miao medicine prescription on chondrocytes in rabbit osteoarthritis model through Wnt/β-catenin signaling pathway,and to explore the mechanism of Miao medicine prescription in preventing and treating cartilage degeneration of osteoarthritis.Methods Thirty-six rabbits were randomly divided into normal control group,model group,Miao medicine fumigation group and Miao medicine iontophoresis group,with nine rabbits in each group.Except for normal control group,the other three groups were injected with papain into the right knee joint to make knee osteoarthritis(KOA)model.After successful modeling,the rabbits in the Miao medicine fumigation group and the Miao medicine iontophoresis group were treated with Miao medicine prescription fumigation and Miao medicine prescription iontophoresis for 20 d respectively.The cartilage tissues of rabbits in each group were processed for HE staining,and the serum samples of rabbits in each group were prepared.Chondrocytes were collected and cultured from cartilage of three rabbits,and the chondrocytes were divided into four groups:normal control group,model group,Miao medicine fumigation group,Miao medicine iontophoresis group.Except for the normal control group,chondrocytes in the other three groups were treated with 10μg/L IL-1βfor 24 h to construct osteoarthritis chondrocyte model,and the chondrocytes were intervened in vitro with serum from rabbits of corresponding group during the modeling period.ELISA was used to detect the amount ofβ-catenin、TNF-αin the supernatant of chondrocytes.The protein expression levels of BMP-2、MMP-3、MMP-13、Wnt-16、β-catenin in chondrocytes were determined by Western blotting,and the mRNA levels of BMP-2、MMP-3、MMP-13、Wnt-16、β-catenin in chondrocytes were detected by qPCR.Results Compared with model group,the surface fibrosis of articular cartilage and the destruction of chondrocyte structure in Miao medicine fumigation group and Miao medicine iontophoresis group were improved in varying degree

关 键 词：苗药 熏蒸疗法 离子导入 骨关节炎 软骨细胞 WNT/Β-CATENIN信号通路 

分 类 号：R684.3[医药卫生—骨科学]