右归丸对膝骨关节炎模型鼠软骨组织钙离子调控蛋白表达的影响

Effects of Yougui Pill on Calcium Regulatory Protein of Chondrocytes in Rats with Knee Osteoarthritis

关键词:  右归丸  膝骨关节炎  钙离子代谢  热休克蛋白90α

Key words:  Yougui pill  knee osteoarthritis  calcium metabolism  Hsp90α

摘要:

目的 观察右归丸对膝骨关节炎（knee osteoarthritis，KOA）模型鼠显微结构和相关蛋白表达的影响，进一步揭示右归丸防治KOA的机制。方法 将大鼠随机分为假手术组、模型组、硫酸氨基葡萄糖组，右归丸（高、中、低剂量）组，每组10只。采用改良Hulth法制备KOA大鼠模型6周后，分别予相应药物灌胃8周。HE染色法观察软骨组织的形态学变化，并进行Mankin评分；应用免疫组化法检测各组大鼠软骨组织热休克蛋白90α（heat shock protein 90α，Hsp90α）、钙调蛋白（calmodulin，CaM）、钙调蛋白激酶Ⅱ（calmodulin kinase-Ⅱ，CaMK-Ⅱ）和肌球蛋白磷酸酶Rho相互作用蛋白（myosin phosphatase Rho-interacting protein，Mprip）的表达；应用Western blotting检测各组大鼠软骨组织Hsp90α的表达。结果 与假手术组比较，模型组大鼠软骨组织Mankin评分、Hsp90α的蛋白表达显著升高，CaM和Mprip的蛋白表达明显降低（P<0.05）；模型组大鼠关节软骨边缘严重破坏，软骨细胞排列紊乱。与模型组相比，右归丸高剂量组大鼠软骨组织Mankin评分、Hsp90α的蛋白表达均明显降低，CaM和Mprip的蛋白表达明显升高（P<0.05），软骨结构趋于正常，软骨细胞分布仅偶见不均，关节软骨表面欠光滑。结论 右归丸通过促进KOA模型鼠软骨组织的Mprip和CaM蛋白表达、抑制其Hsp90α的蛋白表达来调控钙离子代谢，从而有效保护KOA模型鼠关节软骨，延缓关节软骨退变。

Abstract:

OBJECTIVE To observe the microstructure and the expression of related proteins of articular cartilage in the knee osteoarthritis rats, which were treated with Yougui pill, and to further reveal its mechanism. METHODS Sixty SD rats were randomly divided into six groups:sham operation group, model group, glucosamine sulfate group, Yougui pill(high-dose, middle-dose and low-dose) group, 10 rats in each group. The modified Hulth method was used to make KOA models for 6 weeks. The rats were gavaged with corresponding drugs for 8 weeks. The morphological change of cartilage was observed by HE staining, it was evaluated by Mankin score. The expression of heat shock protein 90a(Hsp90a), calmodulin(CaM), calmodulin kinase-Ⅱ(CaMK-Ⅱ), myosin phosphatase Rho-interacting protein(Mprip) was analyzed by immunohistochemistry. The expression of Hsp90α was checked by Western blotting. RESULTS Compared with the sham operation group, the Mankin score was obviously raised in the model group, the protein expression of Hsp90α was evidently increased in the model group, the protein expression of CaM and Mprip was evidently decreased in the model group(P<0.05), articular cartilage was seriously damaged, and chondrocytes were arranged in disorder. Compared with the model group, the Mankin score and the expression of Hsp90α were declined obviously in the high-dose Yougui pill group, but the protein expression of CaM and Mprip was evidently increased in the high-dose group(P<0.05), cartilage structure was tended to be normal, the chondrocytes distribution was uneven, and articular cartilage surface was not smooth. CONCLUSION Yougui pill is effective for delaying articular cartilage degeneration. Yougui pill can regulate calcium metabolism by up-regulating the expression of Mprip and CaM and down-regulating the expression of Hsp90α.