重楼皂苷VII对IL-1β诱导的关节软骨细胞凋亡和分泌炎症介质的影响

Effects of polyphyllin VII on IL-1β-induced apoptosis and secretion of inflammatory mediators

目的探讨重楼皂苷VII对IL-1β诱导的关节软骨细胞凋亡和分泌炎症介质的影响。方法分离培养人膝关节软骨细胞,分成对照组、模型组、低(0.1μg/mL)、中(0.2μg/mL)、高剂量组(0.4μg/mL)、中剂量+Vector组、中剂量+TLR4组,CCK-8法检测细胞增殖,流式细胞术检测细胞凋亡,ELISA法分析TNF-α、PGE2水平,比色法检测NO水平,Western blot检测TLR4、C-Caspase-3、MMP-13、MMP-1、MMP-3蛋白表达。结果与对照组比较,模型组细胞存活率下降,细胞凋亡率上升,TLR4、C-Caspase-3、MMP-13、MMP-1、MMP-3蛋白水平和TNF-α、NO、PGE_(2)水平升高。与模型组比较,低、中、高剂量组细胞存活率上升,细胞凋亡率降低,TLR4、C-Caspase-3、MMP-13、MMP-1、MMP-3蛋白水平和TNF-α、NO、PGE_(2)水平降低。与中剂量+Vector组比较,中剂量+TLR4组细胞存活率降低,细胞凋亡率升高,TLR4、C-Caspase-3、MMP-13、MMP-1、MMP-3蛋白水平和TNF-α、NO、PGE_(2)水平升高。结论重楼皂苷VII抑制IL-1β诱导的关节软骨细胞凋亡、分泌炎症介质和MMP-13、MMP-1、MMP-3蛋白表达,机制与降低TLR4有关。

Objective To investigate the effects of polyphyllin VII on apoptosis and secretion of inflammatory mediators of articular chondrocytes induced by IL-1β.Methods Human knee articular cartilage chondrocytes were isolated and cultured.Cells were divided into Control,Model group,low-(0.1μg/mL),middle-(0.2μg/mL),and high-dose(0.4μg/mL)group,middle dose+Vector group,and middle dose+TLR4 group.Cell proliferation was detected with CCK-8 method.Cell apoptosis was detected with flow cytometry,The levels of TNF-αand PGE2 were analyzed with ELISA.The level of NO was detected with colorimetry.The protein expressions of TLR4,C-Caspase-3,MMP-13,MMP-1,and MMP-3 were detected with Western blotting.Results Compared with those in the Control group,cell survival rate in the Model group decreased,the cell apoptosis rate increased,and the levels of TLR4,C-Caspase-3,MMP-13,MMP-1,and MMP-3 protein and TNF-α,NO,PGE2 increased.Compared with those in IL-1βgroup,the cell survival rate in low-,middle-,and high-dose groups gradually increased,the apoptosis rate gradually decreased,and the levels of TLR4,C-Caspase-3,MMP-13,MMP-1,MMP-3 protein and TNF-α,NO and PGE2 decreased.Compared with the middle dose+Vector group,the middle dose+TLR4 group had a lower cell survival rate and an increased cell apoptosis rate.The levels of TLR4,C-Caspase-3,MMP-13,MMP-1,MMP-3 protein and TNF-α,NO,PGE2 increased.Conclusion Polyphyllin VII inhibits IL-1β-induced apoptosis,secretion of inflammatory mediators,and the expression of MMP-13,MMP-1,and MMP-3 proteins of articular chondrocytes.The mechanism is related to the reduction of TLR4.