摘要
目的探讨低氧诱导因子-1(hypoxia inducible factor-1α,HIF-1α)在喹啉酸诱导大鼠肾上腺嗜铬细胞瘤PC12细胞损伤中的作用。方法用不同浓度的喹啉酸(2.5,5和10mmol·L-1)处理PC12细胞24 h诱导细胞损伤,采用噻唑蓝还原法和乳酸脱氢酶漏出率检测法测定细胞损伤程度,采用丙二醛和超氧化物歧化酶试剂盒检测细胞内氧自由基水平,Hoechst 33258单荧光染色法观察细胞凋亡,免疫荧光染色法检测HIF-1α在细胞内的表达,免疫印迹法检测细胞HIF-1α、蛋白激酶B(Akt)、磷酸化Akt(phosphorylated-Akt,pAkt)、淋巴瘤/白血病-2(B cell lymphoma/lewkmia-2,Bcl-2)和Bcl-2相关X蛋白(Bcl-2 associated X protein,Bax)的表达。结果喹啉酸可剂量依赖性地诱导PC12细胞损伤,导致细胞内氧自由基增多和细胞凋亡。同时,喹啉酸可使PC12细胞HIF-1α表达上调并聚集于核内;p-Akt表达增加,Bax/Bcl-2表达比例增加。结论 HIF-1α和Akt介导了喹啉酸诱导PC12的细胞凋亡;此外,氧化应激过程也可能参与了细胞损伤。
Aim To investigate the role of HIF-1α in PC12 cell injury induced by quinolinic acid. Methods PC12 cells were treated with quinolinic acid at the doses of 2. 5,5 and 10 mmol · L^- 1,the cell viability was determined by MTT reduction assay and LDH assay, the intracellular levels of oxygen species was measured by assessing SOD and MDA levels,cell apoptosis was determined by Hoechst 33258 staining,the intracellular distribution of HIF-1α was examined by HIF-1α immunostaining,and the expressions of HIF-1α,Akt,p-Akt,Bcl-2 and Bax were determined by immunoblotting analysis. Results Quinolinic acid induced cell injury in PC12 cells in a dose-dependent manner,and potentiated oxygen radical production and cell apoptosis. In addition,quinolinic acid enhanced HIF-1α expression and accumulation in nuclei. The pAkt expression and Bax / Bcl-2 ratio was increased by quinolinc acid in PC12 cells. Conclusions HIF-1αand Akt mediate qunolinc acid-induced cell apoptosis in PC12 cells. And cellular oxidative stress may contribute to the injury as well.
出处
《中国药理学通报》
CAS
CSCD
北大核心
2015年第4期493-499,共7页
Chinese Pharmacological Bulletin
基金
国家自然科学基金资助项目(No 81300059)
江苏大学高级人才基金项目(No 08JDG005
11JDG092)
