独活寄生汤对膝骨关节炎大鼠SDF-1/CXCR4信号通路的影响

Effects of Duhuo Jisheng Decoction on SDF-1/CXCR4 Signaling Pathway in Rats with Knee Osteoarthritis

目的:观察独活寄生汤对膝骨关节炎(KOA)大鼠SDF-1/CXCR4信号通路相关调节因子的影响,探讨独活寄生汤治疗KOA可能的作用机制。方法:选取2月龄SPF级SD雄性大鼠45只,适应性喂养1周后,随机分为空白组(15只)和造模组(30只)。造模组采用双膝关节腔注射4%木瓜蛋白酶复制KOA模型,术后2周随机分为模型组(15只)和治疗组(15只)。治疗组给予独活寄生汤,按照9.3 g·kg-1·d-1的剂量进行灌胃。空白组和模型组给予等量的0.9%生理盐水进行灌胃;治疗8周后,10%水合氯醛腹腔注射麻醉后,收集关节液,-80℃保存备用,采用ELISA法检测SDF-1含量;收集膝关节滑膜组织,迅速置入液氮中保存,采用Real-time PCR法检测SDF-1 mRNA表达,Western blot法检测SDF-1蛋白表达;收集胫骨平台和股骨下端软骨组织,迅速置入液氮中保存,采用Real-time PCR法检测CXCR4、MMP-3、MMP-9和MMP-13 mRNA表达,Western blot法检测CXCR4、MMP-3、MMP-9、MMP-13蛋白表达。结果:ELISA结果显示:与模型组相比,独活寄生汤可有效降低大鼠关节液中SDF-1含量(P<0.01)。Real-time PCR和Western blot结果表明:与模型组相比,独活寄生汤能抑制大鼠滑膜组织中SDF-1和软骨组织中CXCR4、MMP-3、MMP-9、MMP-13 mRNA和蛋白的表达(P<0.01)。结论:独活寄生汤可通过调控SDF-1/CXCR4信号通路,抑制KOA炎症反应,起到治疗KOA的作用。

Objective:To observe the effect of Duhuo Jisheng Decoction(DHJSD)on the related regulatory factors in SDF-1/CXCR4 signaling pathway in rats with knee osteoarthritis,so as to explore its mechanism on knee osteoarthritis treatment.Methods:After one week of acclimation,two-month-old male SPF grade Sprague Dawley rats(n=45)were randomly divided into blank group(n=15)and knee osteoarthritis model group(n=30).Knee osteoarthritis model was induced in double knees of rats by injecting 0.2 mL of 4%papain solution on days 1,4 and 7.Two week later,the animals were randomly divided into model group(n=15)and treatment group(n=15).The treatment group received a clinical oral dose of DHJSD(9.3 g/kg/day)intragastrically.The blank group and model group received an equivalent amount of 0.9%saline.After eight weeks treatment,the animals were anaesthetized by intraperitoneal injection 10%chloralhydrate,joint fluid was obtained and storaged in-80℃;synovial tissue and cartilage tissue of tibial plateau and lower femur were obtained and storaged in liquid nitrogen immediately.The content of SDF-1 was measured by ELISA.The mRNA expression of SDF-1 in synovial tissue and CXCR4、MMP-3、MMP-9、MMP-13 in cartilage tissue were measured by Real-time PCR.The protein levels of SDF-1 in synovial tissue and CXCR4、MMP-3、MMP-9、MMP-13 in cartilage tissue were measured by Western blot.Results:ELISA results show that DHJSD could effectively inhibit the expression of SDF-1 compared with the model group(P<0.01).Real-time PCR results suggested that DHJSD could inhibit the mRNA expression of SDF-1 in synovial tissue and CXCR4、MMP-3、MMP-9、MMP-13 in cartilage tissue compared with the model group(P<0.01).Western blot results suggested that DHJSD could inhibit the protein expression of SDF-1 in synovial tissue and CXCR4、MMP-3、MMP-9、MMP-13 in cartilage tissue compared with the model group(P<0.01).Conclusion:The above results suggested that DHJSD could inhibit the inflammatory reaction in knee osteoarthritis by regulating SDF-1/CXCR4 signaling pathway,which may explain its clinical efficacy in knee osteoarthritis treatment.