丹参酮ⅡA抑制软骨细胞炎症、软骨基质降解的机制及对大鼠骨关节炎的影响

TanshinoneⅡA Ameliorates Osteoarthritis in Rats to Inhibit Chondrocyte Inflammation and Cartilage Matrix Degradation

目的探究丹参酮ⅡA(TanⅡA)通过调节β-arrestin2表达对软骨细胞炎症及软骨基质降解的机制,以及对大鼠骨关节炎(osteoarthritis,OA)的影响。方法白细胞介素-1β(IL-1β)诱导大鼠关节软骨细胞制备关节炎体外模型。24只SD大鼠随机分为对照组、模型组和TanⅡA组,每组8只。模型组和TanⅡA组采用改良Hulth法建立大鼠膝骨关节炎模型,TanⅡA组造模成功后予TanⅡA 0.5 mg/kg灌胃,模型组造模成功后和对照组(未造模)予等量0.9%氯化钠注射液灌胃。细胞增殖检测软骨细胞活力;Western blot检测一氧化氮合酶(iNOS)、环氧化酶-2(COX-2)、基质金属蛋白酶(MMPs)、Ⅱ型胶原、Aggrecan、β-arrestin2、p-p65和p65蛋白表达,酶联免疫吸附法检测前列腺素E 2(PGE 2)含量;一氧化氮(NO)测定试剂盒检测NO含量;HE染色及甲苯胺蓝染色检测大鼠软骨组织病理组织学变化。结果与对照组相比,IL-1β100.00μmol/L可显著抑制软骨细胞活力(P<0.05),IL-1β可显著升高大鼠软骨细胞中NO、PGE 2、iNOS、COX-2、MMPs和p-p65/p65的表达(P<0.05),IL-1β可显著降低Ⅱ型胶原、Aggrecan和β-arrestin2的表达(P<0.05);与IL-1β组相比,TanⅡA(25.00μmol/L和50.00μmol/L)呈剂量依赖性升高软骨细胞活力、Ⅱ型胶原、Aggrecan和β-arrestin2的表达(P<0.05),剂量依赖性降低大鼠软骨细胞中NO、PGE 2、iNOS、COX-2、MMPs和p-p65/p65的表达(P<0.05)。对照组大鼠软骨表面光滑,软骨细胞排列整齐;模型组大鼠软骨表面粗糙,软骨层变薄,软骨细胞排列紊乱。与对照组相比,模型组大鼠Mankin评分显著升高(P<0.05);与模型组相比,TanⅡA组大鼠Mankin评分显著降低(P<0.05)。结论TanⅡA通过调节β-arrestin2表达抑制软骨细胞炎症及软骨基质降解,改善OA大鼠病情。

Objective To explore the mechanism of tanshinoneⅡA(TanⅡA)on chondrocyte inflammation and cartilage matrix degradation by regulating the expression ofβ-arrestin2,and its effect on osteoarthritis(OA)in rats.Methods Rat articular chondrocytes were induced by interleukin-1β(IL-1β)to establish arthritis model in vitro.Twenty-four SD rats were randomly divided into control group,model group and TanⅡA group,with 8 rats in each group.Rat knee osteoarthritis model in the model group and TanⅡA group were established by modified Hulth method.TanⅡA 0.5 mg/kg was given by gavage in TanⅡA group,and the model group and the control group were given the same amount of 0.9%sodium chloride injection by gavage.The viability of chondrocytes and cell proliferation were detected.The expression of inducible nitric oxide synthase(iNOS),cyclooxygenase-2(COX-2),matrix metalloproteinases(MMPs),typeⅡcollagen,Aggrecan,β-arrestin2,p-p65 and p65 was detected by Western blot.The content of PGE 2 was detected by enzyme-linked immunosorbent assay.The content of nitric oxide(NO)was detected by NO assay kit.Histopathological changes in rat cartilage tissue was detected by HE staining and toluidine blue staining.Results Compared with the control group,IL-1β(100.00μmol/L)significantly inhibited the viability of chondrocytes(P<0.05).In addition,IL-1βsignificantly increased the expression of NO,PGE 2,iNOS,COX-2,MMPs and p-p65/p65(P<0.05),and significantly decreased the expression of typeⅡcollagen,Aggrecan andβ-arrestin2(P<0.05).Compared with IL-1βgroup,TanⅡA(25.00μmol/L and 50.00μmol/L)increased expression of viability,typeⅡcollagen,Aggrecan,andβ-arrestin2 in chondrocytes in a dose-dependent manner,while decreased expression of NO,PGE 2,iNOS,COX-2,MMPs and p-p65/p65 in chondrocytes in a dose-dependent manner(P<0.05).In control group,the cartilage surface was smooth and the chondrocytes were arranged in order;in model group,the cartilage surface was rough,the cartilage layer became thin and the chondrocytes were arranged in disorder.Compared with control group,the Mankin score of the model group was significantly increased(P<0.05);compared with model group,the Mankin score of the TanⅡA group was significantly decreased(P<0.05).Conclusion TanⅡA can inhibit chondrocyte inflammation and cartilage matrix degradation by regulating the expression ofβ-arrestin2,thereby improving the condition of OA rats.