灵芝多糖对地塞米松诱导的MC3T3-E1成骨细胞凋亡的影响及可能机制

Effect and its possible mechanism of Ganoderma lucidum polysaccharide on dexamethasone-induced apoptosis of MC3T3-E1 osteoblasts

目的探讨灵芝多糖(GLP)对地塞米松(DEX)诱导的MC3T3-E1成骨细胞凋亡的影响及可能的作用机制。方法采用CCK-8法检测不同浓度GLP对DEX诱导的MC3T3-E1成骨细胞凋亡的保护作用,选出GLP最佳浓度;实验分为空白对照组(Control)、地塞米松组(DEX)与灵芝多糖+地塞米松组(GLP+DEX)。试剂盒检测各组细胞半胱氨酸天冬氨酸蛋白酶-3(Caspase-3)和半胱氨酸天冬氨酸蛋白酶-9(Caspase-9)的活性;Hoechst染色及流式细胞仪检测各组细胞的凋亡情况;Western blot检测Bcl-2、BAX、caspase-3、JAK2、p-JAK2、STAT3、p-STAT3的表达。结果DEX可上调MC3T3-E1成骨细胞caspase-3和caspase-9活性,诱导MC3T3-E1成骨细胞凋亡,上调凋亡相关蛋白Bax、caspase-3的表达,下调抗凋亡蛋白Bcl-2的表达,上调JAK2-STAT3信号通路相关蛋白p-JAK2、p-STAT3的表达,灵芝多糖干预可逆转以上变化的发生。结论灵芝多糖可能通过抑制JAK2/STAT3通路抑制地塞米松诱导的MC3T3-E1成骨细胞凋亡。

Objective To explore the effect and its possible mechanism of Ganoderma lucidum polysaccharides(GLP)on apoptosis of MC3T3-E1 osteoblasts induced by dexamethasone(DEX).Methods CCK-8 method was used to detect the protective effect of different concentrations of GLP on DEX-induced apoptosis of MC3T3-E1 osteoblasts,and the best concentration of GLP was selected.The experiment was divided into blank control group(Control),dexamethasone group(DEX)and Ganoderma lucidum polysaccharide+dexamethasone group(GLP+DEX).The activity of caspase-3 and caspase-9 in each group were detected.Hoechst staining and flow cytometry were used to detect the apoptosis of cells in each group.The expressions of BAX,Bcl-2,Caspase-3,JAK2-STAT3,JAK2,p-JAK2,p-STAT3 and STAT3 were detected by Western blot.Results DEX up-regulated the activities of caspase-3 and caspase-9 in MC3T3-E1 osteoblasts,induced the apoptosis of MC3T3-E1 osteoblasts,promoted the expressions of Bax and caspase-3,down-regulate the expression of Bcl-2 and up-regulated the expressions of p-JAK2 and p-STAT3.The intervention of Ganoderma lucidum polysaccharide can reverse the above changes.Conclusion Ganoderma lucidum polysaccharides may inhibit the dexamethasone-induced apoptosis of MC3T3-E1 osteoblasts by inhibitting JAK2/STAT3 pathway.