补肾活血汤通过Runx2/Osterix促进骨质疏松模型大鼠的骨折愈合  

Bushen Huoxue Decoction promotes osteoporotic fracture healing in rats through Runx2/Osterix

作　　者：胡流超 罗毅文[2] 程英雄[2] 吴志方[2] 罗辉 沈玮 Hu Liuchao;Luo Yiwen;Cheng Yingxiong;Wu Zhifang;Luo Hui;Shen Wei(Graduate School of Guangzhou University of Chinese Medicine,Guangzhou 510405,Guangdong Province,China;Hospital,Guangzhou University of Chinese Medicine,Guangzhou 510240,Guangdong Province,China)

机构地区：[1]广州中医药大学研究生院,广东省广州市510405 [2]广州中医药大学附属骨伤科医院,广东省广州市510240

出　　处：《中国组织工程研究》2019年第11期1717-1722,共6页Chinese Journal of Tissue Engineering Research

基　　金：国家自然基金项目(81473699);项目负责人:罗毅文;广东省科技厅项目(2014A020221022);项目负责人:程英雄~~

摘　　要：背景:Runx2/Osterix是促进成骨细胞分化的重要的转录因子,研究发现中药能通过上调Runx2/Osterix促进骨折愈合。补肾活血汤是临床上治疗骨折的经验方药,能促进骨形成,通过探究其作用机制更好服务临床。目的:通过慢病毒载体介导Runx2基因沉默,研究补肾活血汤对骨质疏松模型大鼠骨折愈合影响及分子机制。方法:将广州中医院药大学动物实验中心提供的40只4月龄成年雌性SD大鼠随机等为5组,模型组、补肾活血汤组、Runx2沉默组、Runx2沉默+补肾活血汤组及对照组,对照组只建立股骨骨折模型,其他各组切除双侧卵巢并造成股骨骨折制作骨质疏松骨折大鼠模型。建模后补肾活血汤组、Runx2沉默+补肾活血汤组予以补肾活血汤灌胃,模型组、对照组及Runx2沉默组予以等体积蒸馏水灌胃;Runx2沉默组、Runx2沉默+补肾活血汤组于骨折处注射Runx2重组慢病毒,模型组、对照组及补肾活血汤组于相同部位处注射等体积PBS。建模4周后,Micro CT测定股骨标本骨密度、组织矿物质密度及骨体积分数(BV/TV),q PCR、Western blot测定骨痂成骨标志基因Runx2及Osterix mRNA及蛋白的表达水平。结果与结论:(1)与对照组相比,模型组骨密度、组织矿物质密度、Runx2和Osterixm RNA及蛋白表达量显著降低(P <0.01);(2)与模型组相比,补肾活血汤组骨体积分数(BV/TV)、Runx2、Osterix mRNA表达量升高(P<0.05),而骨密度、组织矿物质密度、Runx2及Osterix蛋白表达量显著升高(P<0.01);Runx2沉默组骨体积分数和Osterix mRNA表达量降低(P <0.05),骨密度、组织矿物质密度、Runx2 mRNA及蛋白表达量、Osterix蛋白表达量显著降低(P <0.01);(3)与Runx2沉默组比较,Runx2沉默+补肾活血汤组上述全部指标的m RNA及蛋白表达均显著升高(P <0.01);(4)结果表明,慢病毒介导Runx2基因沉默使大鼠骨质疏松骨折愈合延缓,补肾活血汤能上调Runx2/Osterix表达,逆转Runx2基因沉默�BACKGROUND:Runx2/Osterix is an important transcription factor promoting osteoblast differentiation.Chinese herbs have been shown to promote fracture healing through up-regulation of Runx2/Osterix.Bushen Huoxue Decoction is a prescription used to treat fractures and it can promote bone formation.Exploring underlying mechanism is helpful for its clinical application.OBJECTIVE:To study the effect of Bushen Huoxue Decoction on osteoporotic fracture healing in rats and the underlying molecular mechanism.METHODS:Forty 4-month-old adult female Sprague-Dawley rats provided by Laboratory Animal Centre of Guangzhou University of Chinese Medicine were randomly divided into five groups:model,Bushen Huoxue Decoction,Runx2 gene silencing,Runx2 gene silencing+Bushen Huoxue Decoction,and control groups.The femoral fracture model was established in the control group,while the osteoporotic femoral fracture model was established by ovariectomy in the other groups.Rats in the Bushen Huoxue Decoction and Runx2 gene silencing+Bushen Huoxue Decoction groups were treated with Bushen Huoxue Decoction,while rats in the model,control and Runx2 gene silencing groups were treated with equal volume of distilled water.Rats in the Runx2 gene silencing and Runx2 gene silencing+Bushen Huoxue Decoction groups were injected with Runx2 recombinant lentivirus,while rats in the model,control,and Bushen Huoxue Decoction groups were injected with equal volume of PBS at the same site.After 4 weeks,bone mineral density,tissue mineral density and bone volume fraction were measured by Micro CT.The mRNA and protein expression levels of the callus osteogenesis markers Runx2 and Osterix were detected by qPCR and western blot assay,respectively.RESULTS AND CONCLUSION:Compared with the control group,bone mineral density,tissue mineral density,and expression levels of Runx2 and Osterix mRNA and protein in the model group were significantly decreased(P<0.01).Compared with the model group,bone volume fraction and Runx2 and Osterix mRNA expression levels were signif

关 键 词：补肾活血汤 骨质疏松 骨质疏松性骨折 骨质疏松性骨折愈合 慢病毒 核心结合因子 骨形成 组织工程 基因沉默 骨折愈合 

分 类 号：R453[医药卫生—治疗学] R364[医药卫生—临床医学]