摘要
目的研究分泌型磷脂酶A2-IB（sPLA2-IB）及M型磷脂酶A2受体（PLA2R）在人肾小球足细胞损伤中的作用及可能的信号传导通路。方法体外培养条件永生性人足细胞，按刺激的sPLA2-IB浓度分组，以PBS或不同浓度sPLA2-IB（10^-9、10^-7、10^-5mol／L）分别刺激2h。划痕实验检测足细胞迁移率，Hoechst．33342及流式细胞术检测足细胞凋亡率，Western印迹检测足细胞p-cPLA2（x、P—p38及p53蛋白表达。将细胞分为对照组、siRNA阴性对照组、PLA2RsiRNA组，采用PLA2RsiRNA或对照siRNA转染足细胞，再给予10～mol／LsPLA2-IB刺激足细胞2h，用Western印迹检测p-cPLA2c~、p-p38及p53蛋白表达水平。结果与PBS对照组比较，10^-7、10^-5mol／L组sPLA2-IB降低了足细胞迁移能力，增加了足细胞的凋亡率，且呈剂量依赖性（均P〈0．05）。此外，与PBS对照组比较，10^-7、10^-5mol／L组p-cPLA2（x、p-p38及p53蛋白表达增加（均P〈0．05），且随着sPLA2-IB浓度的增高，其蛋白表达含量亦逐渐增高。采用PLA2RsiRNA转染沉默足细胞PLA2R表达后，再以sPLA2．IB刺激，p-cPLA2（x、p-p38及p53蛋白表达均较对照组下降（均P〈0．05）。结论sPLA2-IB增加人足细胞凋亡率并降低细胞迁移能力，可能是通过PLA2R活化p38-cPLA2α-p53信号通路导致的。
Objective To investigate the role of the group IB secretory phospholipase A2 （sPLA2-IB） and M-type phospholipase A2 receptor （PLA2R） in human podocyte injury and its possible signal transduction pathway. Methods Differentiated human podocytes were exposed to PBS or different sPLA2- IB concentration conditions （10^-9, 10^-7, 10^-5 mol/L） for 2 hours. The wound healing assay was used to measure cell migration rate; Apoptosis in cultured human podocytes was assessed by Hoechst 33342 staining and flow cytometry; Western blot was used to analyze the protein expression of p- cPLA2α, p- p38, and p53. Then control siRNA or PLA2R siRNA were transfected to podocytes. Podocytes were divided into normal control group, negative control siRNA group and PLA2R siRNA group. Twenty four hous later, the cells were stimulated by 10^-5 mol/L sPLA2- IB for 2 hours. The protein expression of p-cPLA2α, p-p38, and p53 were detected by Western blot. Results Compared to PBS control group, the migration ability of podocytes decreased when stimulated with sPLA2-IB （10^-7 mol/L-10^-5 mol/L）, and the apoptosis of podocytes increased in a concentration-dependent manner, the protein level of p-cPLA2α, p-p38 and p53 protein increased too. After the knockdown of PLA2R by
PLA2R siRNA transfection, stimulated the podocytes with the same dosage of sPLA2-IB, the protein expression of p- cPLA2α, p-p38 and p53 all decreased. Conclusion sPLA2- IB stimulation can increase human podocyte apoptosis and decrease its migration ability. The possible mechanism might be through p38-cPLA2α-p53 pathway.
出处
《中华肾脏病杂志》
CSCD
北大核心
2017年第9期691-697,共7页
Chinese Journal of Nephrology
基金
基金项目：国家自然科学基金面上项目（81670635）
福建省自然科学基金项目（2016J01532）
福建省卫生计生中青年骨干人才培养项目（2016.ZQN.36）
关键词
足细胞
磷脂酶A2
受体
磷脂酶A2
凋亡
Podocytes
Phospholipase A2
Receptor, phospholipase A2
Apoptosis
