PI3K/Akt信号通路在脓毒症肾脏损伤诱导的自噬中的调节作用  

PI3K/Akt signaling pathway regulates autophagy induced by acute kidney injury in septic rats

作　　者：向镜芬[1,2] 杨祥[1,2] 龚剑锋[1,2] 雷伟健 邓艳琼[1,2] 牟丹[1,2] 钟国权 孟启勇[1,2] 

机构地区：[1]暨南大学第五附属医院 [2]清远市人民医院重症医学科,广东清远511500

出　　处：《中国病理生理杂志》2014年第6期1052-1058,共7页Chinese Journal of Pathophysiology

基　　金：清远市医疗卫生领域自筹经费科技计划项目(No.1301)

摘　　要：目的:研究脓毒症造成肾脏损伤时的自噬情况以及磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(Akt)信号通路的调节作用。方法:对大鼠盲肠进行结扎与穿刺(CLP),对肾脏组织切片进行HE染色,并测定血清尿素氮和肌酐。通过Western blotting定量分析CLP大鼠肾脏损伤发生后不同时点自噬相关分子微管相关蛋白轻链3(LC3)Ⅰ/Ⅱ、beclin-1和Akt蛋白磷酸化的表达情况;体外用LPS诱导人近端肾小管上皮细胞株HK-2发生自噬,检测不同浓度LPS和不同刺激时间自噬相关分子LC3Ⅰ/Ⅱ和Akt蛋白磷酸化的表达情况;进一步使用PI3K抑制剂、Akt抑制剂和LPS刺激HK-2细胞观察自噬相关蛋白的表达情况及细胞的凋亡水平。结果:同对照组相比,CLP大鼠显微镜下可见肾损伤的典型病理改变,血清尿素氮和肌酐均有上升。CLP肾脏损伤发生后,自噬相关蛋白LC3Ⅰ/Ⅱ、beclin-1含量及Akt磷酸化水平均有上升。LPS刺激HK-2细胞后,随着刺激浓度的增加,p-Akt(308)表达量逐渐提高,而LC3Ⅰ/Ⅱ及p-Akt(472)的表达量在10 mg/L LPS刺激组最高。随着刺激时间的延长,p-Akt(308)表达量逐渐提高;LC3Ⅰ/Ⅱ表达量同p-Akt(472)在刺激8 h时最高;使用PI3K抑制剂及Akt抑制剂后,LPS诱导的LC3表达显著下调,HK-2细胞凋亡明显增加。结论:CLP肾脏损伤发生时可以诱导自噬发生,PI3K/Akt信号通路在其中发挥重要调节作用。AIM：To investigate the autophagy induced by sepsis and acute kidney injury , and the regulation of phosphoinositide 3-kinase （PI3K）/protein kinase B （Akt） signaling pathway in this process.METHODS： The rats were subjected to cecal ligation and puncture （ CLP） or sham operation .Histopathologic changes of the renal tissues were examined by HE staining .Blood urea nitrogen （ BUN） and serum creatinine （ SCr） were measured by chemical colorimetry.The protein expression of microtubule-associated protein light chain 3 Ⅰ/Ⅱ （LC3Ⅰ/Ⅱ）, beclin-1 and p-Akt at different time points after CLP was detected by Western blotting .In vitro, human proximal tubular epithelial cell line HK-2 were treated with LPS to induce autophagy .The protein expression of LC 3 Ⅰ/Ⅱ and p-Akt in the HK-2 cells after LPS treatment at different time points and different concentrations was detected by Western blotting .These molecules in HK-2 cells and apoptosis of HK-2 cells treated with LPS plus PI3K inhibitor or Akt inhibitor were also detected .RESULTS： Compared with sham group , the severe changes of renal histopathological injuries in CLP groups were observed , the levels of BUN and SCr in CLP groups were significantly increased .LC3 I/II, beclin-1 and phosphorylation of Akt gradually increased after CLP.After treatment with LPS, the expression of p-Akt （308） in the HK-2 cells gradually increased in a dose-and time-dependent fashion.The expression of beclin-1 and p-Akt （472） reached a peak at 8 h or 10 mg/L LPS treatment.Treatment with PI3K or Akt inhibitor down-regulated the expression of LC3 and promoted the apoptosis of HK-2 cells.CONCLUSION：Autophagy in the kidney is induced by sepsis and acute kidney injury .PI3/Akt signaling pathway may be involved in this process .

关 键 词：自噬 PI3K AKT通路 脓毒症 肾损伤 

分 类 号：R692.6[医药卫生—泌尿科学]