CAJ | 학술논문

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。
목적：연구농독증조성신장손상시적자서정황이급린지선기순3-격매（PI3K）／단백격매B（Akt）신호통로적조절작용。방법：대대서맹장진행결찰여천자（ CLP），대신장조직절편진행HE염색，병측정혈청뇨소담화기항。통과Western blotting정량분석CLP대서신장손상발생후불동시점자서상관분자미관상관단백경련3（LC3）Ⅰ／Ⅱ、beclin-1화Akt단백린산화적표체정황；체외용LPS유도인근단신소관상피세포주HK-2발생자서，검측불동농도LPS화불동자격시간자서상관분자LC3Ⅰ／Ⅱ화Akt단백린산화적표체정황；진일보사용PI3K억제제、Akt억제제화LPS자격HK-2세포관찰자서상관단백적표체정황급세포적조망수평。결과：동대조조상비，CLP대서현미경하가견신손상적전형병리개변，혈청뇨소담화기항균유상승。 CLP신장손상발생후，자서상관단백LC3Ⅰ／Ⅱ、beclin-1함량급Akt린산화수평균유상승。 LPS자격HK-2세포후，수착자격농도적증가，p-Akt（308）표체량축점제고，이LC3Ⅰ／Ⅱ급p-Akt（472）적표체량재10 mg／L LPS자격조최고。수착자격시간적연장，p-Akt（308）표체량축점제고；LC3Ⅰ／Ⅱ표체량동p-Akt（472）재자격8 h시최고；사용PI3K억제제급Akt억제제후，LPS유도적LC3표체현저하조，HK-2세포조망명현증가。결론：CLP신장손상발생시가이유도자서발생， PI3K／Akt신호통로재기중발휘중요조절작용。
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 colorime-try.The protein expression of microtubule-associated protein light chain 3 I/II (LC3 I/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.Treat-ment with PI3K or Akt inhibitor down-regulated the expression of LC3 and promoted the apoptosis of HK-2 cells.CON-CLUSION:Autophagy in the kidney is induced by sepsis and acute kidney injury .PI3/Akt signaling pathway may be in-volved in this process .