摘要
目的探讨P2X4受体在高糖介导PC12细胞损伤中的作用及其可能的作用机制。方法通过高糖处理PC12细胞建立拟糖尿病细胞损伤模型,应用MTS比色法检测不同糖浓度对细胞的损伤作用并确定最佳建模浓度。对各组细胞进行不同干预后,通过ELISA方法检测各组细胞上清液中TNF-α和IL-1β的含量;通过q PCR和Western Blot方法检测细胞内P2X4mRNA、受体蛋白和P65蛋白变化。结果不同糖浓度处理细胞24h后,细胞存活率随糖浓度增大逐渐降低;通过高糖建模并经不同处理发现,与对照组相比,高糖组细胞存活率明显下降,高糖+5-BDBD组与之相比显著上升;通过q PCR和Western Blot检测发现,与对照组相比,高糖组细胞P2X4mRNA、受体蛋白和NF-κB蛋白的表达均明显上升,高糖+5-BDBD组与之相比有所下降,且与对照组无明显差异。结论 P2X4受体参与高糖介导PC12细胞损伤,高糖能诱导PC12细胞NF-κB转录复合体形成,进而介导P2X4受体表达上调,使细胞过度激活并大量释放TNFα、IL-1β等炎性因子对细胞产生毒副作用。
Objective To investigate the effect of P2X4 receptor in high glucose-induced PC12 cell injury and its possible mechanism.Methods Establish the cell injury model of PC12 cells via high glucose treatment,using MTS colorimetric assay to detect the injury effect of different glucose concentrations on cells and to determine the optimal concentration of the model.After treatment of with P2X4 receptor antagonist 5-BDBD,ELISA method was used to detect the content of TNFα and IL-1β in supernatant of each group of cells;q PCR and western blot methods were used to detect the expression of P2X4 mRNA,receptor protein and NF-κB-P65 protein in each group of cells.Results After treatment with different concentrations of glucose for 24 h,the survival rate of the cell decreased as the glucose concentration increased.The survival rate of the high glucose group was significantly decreased when compared with the control group.The survival rate of high glucose ＋ 5-BDBD group was higher than that of the high glucose group.q PCR and western blot results showed that the expression of P2X4 mRNA,receptor protein and NF-κB-P65 protein in high glucose group were significantly increased when compared with the control group.The expression of P2X4 mRNA,receptor protein and NF-κB-P65 protein in high glucose ＋ 5-BDBD group was decreased,the difference was not statistically significant when compared with the control group.Conclusion Our study showed that P2X4 receptors might be involved in high glucose-induced PC12 cells injury,and the up-regulated NF-κB transcription complex may play an important role in activation of PC12 cells.Activated PC12 cells release inflammatory cytokines（i.e.TNFα and IL-1β） that are toxic to cells.
出处
《标记免疫分析与临床》
CAS
2016年第8期932-937,共6页
Labeled Immunoassays and Clinical Medicine
基金
四川省卫计委资助项目(编号:30305020654)
