红芪多糖对脂多糖诱导的牙周膜细胞凋亡及对Wnt信号通路的影响

Effect of Hedysari Polysaccharide on Apoptosis of Periodontal Ligament Cells Induced by LPS and the Expression of Wnt Signaling Pathway

目的:探讨红芪多糖对脂多糖(LPS)诱导的牙周膜细胞凋亡及对Wnt信号通路的影响。方法:体外分离培养人牙周膜细胞。实验分为3组,依次为对照组、脂多糖组和红芪多糖组。对照组细胞用正常的细胞培养液培养细胞;脂多糖组和红芪多糖组细胞用含有脂多糖浓度为10mg/L的培养液培养细胞;红芪多糖组细胞培养液中加入红芪多糖终浓度为10mg/L的培养液。流式细胞仪检测细胞凋亡情况,探针法检测细胞内的活性氧(ROS)水平。试剂盒检测细胞中碱性磷酸酶活性和分泌的肿瘤坏死因子-α(TNF-α)水平。Western blot检测细胞中cmyc、β-连环蛋白(β-catenin)、Bcl-2相关X蛋白(Bax)单克隆抗体、B细胞淋巴瘤/白血病-2(Bcl-2)蛋白水平。结果:脂多糖组细胞存活率明显低于对照组(P<0.01)。脂多糖组细胞凋亡率和Bax水平、ROS水平明显高于对照组(P<0.01)。红芪多糖组细胞凋亡率和Bax水平、ROS水平明显低于脂多糖组(P<0.01)。脂多糖组细胞Bcl-2、c-myc、β-catenin水平和碱性磷酸酶活性明显低于对照组(P<0.01)。红芪多糖组细胞Bcl-2、c-myc、β-catenin水平和碱性磷酸酶活性明显高于脂多糖组(P<0.01)。脂多糖组细胞分泌的TNF-α水平明显高于对照组(P<0.01)。红芪多糖组细胞分泌的TNF-α水平明显低于脂多糖组(P<0.01)。结论:红芪多糖能够抑制脂多糖诱导的人牙周膜细胞凋亡,抑制细胞分泌TNF-α,作用机制可与Wnt信号通路、ROS水平有关。

Objective：To study the effect of Hedysari polysaccharide on apoptosis of periodontal ligament cells induced by LPS and the expression of Wnt signaling pathway.Methods：Human periodontal ligament cells were cultured and divided into three groups,control group,LPS group with normal cell culture medium containing 10μg/mL LPS and Hedysari polysaccharide group with 10μg/mL LPS and 10μg/mL Hedysari polysaccharide.Cell apoptosis was detected by flow cytometry.Intracellular ROS levels were detected by the probe method.The activity of alkaline phosphatase and the levels of TNF-αin the cells were detected by ELISA.The levels of c-myc,β-catenin,Bax and Bcl-2were detected by Western Blot.Results：The apoptosis rate,expression of Bax and ROS level in the LPS group were significantly higher than those in the control group and lower in Hedysari polysaccharide group（P〈0.01）.The levels of Bcl-2,c-myc,β-catenin and alkaline phosphatase in the LPS group were significantly lower than those in the control group and significantly higher in Hedysari polysaccharide group（P〈0.01）.The level of TNF-αin LPS group was significantly higher than those in the control group and Hedysari polysaccharide group（P〈0.01）.Conclusion：Hedysari polysaccharide can inhibit LPS induced apoptosis and the secretion of TNF-αof human periodontal ligament cells.The mechanism may relate to Wnt signaling pathway and ROS level.