白藜芦醇调控SDF-1/CXCR4通路对颞下颌关节骨关节炎模型大鼠软骨细胞凋亡的影响

Impact of resveratrol on chondrocyte apoptosis in temporomandibular joint osteoarthritis model rats by regulating SDF-1/CXCR4 pathway

[目的]探究白藜芦醇(RES)对颞下颌关节骨关节炎(TMJOA)大鼠软骨细胞凋亡的影响及在该过程中对基质细胞衍生因子-1(SDF-1)/CXC趋化因子受体4(CXCR4)轴的调节机制。[方法]将36只Wistar雄性大鼠随机分为对照组、模型组、CXCR4抑制剂组(AMD3100组)、RES低剂量组、RES高剂量组、RES高剂量+重组SDF-1组(RES高+SDF-1组),每组6只。苏木精-伊红(HE)和番红O-固绿染色观察软骨组织病理学变化;分离培养软骨细胞;流式细胞术检测细胞凋亡;酶联免疫吸附法(ELISA)检测细胞中一氧化氮合酶(iNOS)、一氧化氮(NO)、细胞色素C(Cyt C)水平;蛋白免疫印迹分析(Western Blot)检测软骨细胞中caspase-9、Bcl-2、Bax、SDF-1、CXCR4蛋白表达。[结果]与对照组比较,模型组软骨组织结构层次紊乱,细胞数量减少,软骨着色明显减退,损伤评分、细胞凋亡率、iNOS、NO、Cyt C的水平及caspase-9、Bax、SDF-1、CXCR4蛋白表达升高,Bcl-2蛋白表达降低(P<0.05);与模型组比较,AMD3100组和RES低、高剂量组软骨组织结构层次逐渐明显,细胞数量增多,软骨着色增加,损伤评分、细胞凋亡率、iNOS、NO、Cyt C的水平及caspase-9、Bax、SDF-1、CXCR4蛋白表达降低,Bcl-2蛋白表达升高(P<0.05);进一步加入重组蛋白SDF-1发现,SDF-1表达的升高逆转了高剂量RES对信号通路以及软骨细胞凋亡的抑制作用(P<0.05)。[结论]RES能够通过阻断SDF-1/CXCR4信号通路来抑制TMJOA大鼠的软骨细胞凋亡。

[Objective]To explore the impacts of resveratrol(RES)on chondrocyte apoptosis in rats with temporomandibular joint osteoarthritis(TMJOA)and its regulatory mechanism on SDF-1/CXCR4 axis in this process.[Methods]Thirty-six Wistar male rats were randomly grouped into control group,model group,CXCR4 inhibitor group(AMD3100 group),RES low dose group,RES high dose group,and RES high dose+recombinant SDF-1 group(RES high+SDF-1 group),with 6 rats in each group.Histopathological changes of cartilage were observed by hematoxylin-eosin(HE)and safranine O-solid green staining;chondrocytes were isolated and cultured;cell apoptosis was detected by flow cytometry;enzyme-linked immunosorbent assay(ELISA)was applied to detect the levels of nitric oxide synthase(iNOS),nitric oxide(NO)and cytochrome C(Cyt C)in cells;Western blot was applied to detect the expression of caspase-9,Bcl-2,Bax,SDF-1,CXCR4 proteins in chondrocytes.[Results]Compared with the control group,the cartilage tissue structure of the model group was disordered and the number of cells was reduced.The staining of cartilage was obviously reduced.The injury score,cell apoptosis rate,iNOS,NO,Cyt C levels and the expression of caspase-9,Bax,SDF-1,CXCR4 proteins increased.The expression of Bcl-2 protein decreased(P<0.05).Compared with the model group,the structure level of cartilage tissue in AMD3100 group and RES low and high dose groups were gradually obvious.The number of cells increased,and the staining of cartilage increased.The injury score,apoptosis rate,iNOS,NO,Cyt C levels and the expression of caspase-9,Bax,SDF-1,CXCR4 proteins decreased,and the expression of Bcl-2 protein increased(P<0.05).Further addition of recombinant protein SDF-1 showed that the increased expression of SDF-1 reversed the inhibitory effects of high-dose RES on signal pathway and chondrocyte apoptosis(P<0.05).[Conclusion]RES can inhibit chondrocyte apoptosis in TMJOA rats by blocking SDF-1/CXCR4 signal pathway.