缺氧缺血性脑病新生大鼠肾脏 Bax 和 Caspase3-蛋白的表达与肾细胞凋亡的作用机制

摘要

目的：探讨缺氧缺血性脑病（ HIE）新生大鼠肾脏Bax和Caspase-3蛋白的表达与肾细胞凋亡的作用机制。方法新生7 d Wistar大鼠制备HIE模型。分为假手术对照组和HIE组，于HIE后6、12、24和48 h处死大鼠，取肾脏组织，应用流式细胞术检测肾细胞凋亡情况；应用免疫组织化学法检测肾细胞Bax和Caspase3-蛋白的表达情况；ELISA法检测TNF-α、IL -1β的分泌；应用Western blot技术检测肾细胞凋亡与p38MAPK信号通路的关系。结果 HIE组6 h肾脏阳性凋亡细胞开始增多，24 h达到高峰，各时间点阳性凋亡细胞均高于对照组（ P ＜0侣．05）； HIE组12 h Bax和Caspase-3阳性细胞表达开始增多，24 h达到高峰，各时间点阳性细胞均高于对照组（ P ＜0．05）；HIE组6 h TNF-α、IL-1β分泌开始增多，TNF-α12 h达到高峰，IL-1β24 h达到高峰各时间点TNF-α、IL-1β均高于对照组（ P ＜0．05）；HIE组6 hp-p38MAPK蛋白表达开始增多并达到高峰，各时间点p-p38MAPK蛋白均高于对照组（ P ＜0．05）。结论 HIE新生大鼠肾脏细胞Bax和Caspase-3蛋白表达增加，提示Bax和Caspase-3蛋白表达对肾脏细胞凋亡有促进作用，其可能通过p-p38MAPK信号通路实现的。%Objective To investigate the expression of Bax and Caspase-3 in kidney of neonatal rats with hypoxic-ischemic encephalopathy ( HIE) as well as the action mechanism of apoptosis in renal cells .Methods HIE models were established in seven-day-old Wistar rats ,then these rats were randomly divided into sham operation control group and HIE group.The rats were executed at 6h,12h ,24h,48h after HIE and kidney tissues were obtained .The apoptosis condition of renal cells was detected by flow cytometry ,and expression of Bax and Caspase-3 protein was determined by immunohistochemistry , the levels of TNF-α and IL-1βwere detected by ELISA , and renal cells apoptosis and P 38MAPK signaling pathway were examined by Western Blot.Results In HIE group,positive apoptosis cells were increased at 6h and reached peak at 24h, which were significantly higher than those in control group in different time points ( P <0.05).More Bax and Caspase-3 positive cells appeared at 12h in HIE group and peaked at 24h,and positive cells in different time points were significantly higher than those in control group( P <0.05).At 6h,the levels of TNF-αand IL-1βwere increased in HIE group,and TNF-αreached peak at 12h,however,IL-1βreached peak at 24h,and the levels of TNF-αand IL-1βin different time points were significantly higher than those in control group ( P <0.05).The expression levels of p-P38MAPK protein were increased at 6h in HIE group and reached the peak ,and the expression levels of p-P38MAPK protein were significantly higher than those in control group in different time points ( P <0.05).Conclusion The expression levels of Bax and Caspase-3 in renal cells of neonate rats with HIE are increased ,which suggests that the expression of Bax and Caspase-3 contributes to the apoptosis of renal cells via p-P38 MAPK signaling pathway .