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探讨低氧诱导因子-1α(hypoxia inducible factor-1α,HIF-1α)在喹啉酸诱导人神经母细胞瘤SH-SY5Y细胞损伤中的作用。将体外培养的人神经母细胞瘤SH-SY5Y细胞分为对照组、喹啉酸低、中、高剂量组及HIF-1α抑制剂二甲氧基雌二醇(2-methoxyestradiol,2ME2)预处理喹啉酸高剂量组,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,Hoechst 33342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(protein kinase B,Akt)、磷酸化Akt(p-Akt)、Bcl-2和Bax的表达。结果显示,喹啉酸可剂量、时间依赖性地诱导SH-SY5Y细胞损伤,导致细胞凋亡。同时,喹啉酸可使SH-SY5Y细胞HIF-1α表达上调并发生核转位、p-Akt表达增加及Bax/Bcl-2表达比例增加,而2ME2可抑制喹啉酸诱导的SH-SY5Y细胞损伤及降低HIF-1α、p-Akt和Bax/Bcl-2的表达。由此说明,HIF-1α/Akt通路介导了喹啉酸诱导SHSY5Y的细胞凋亡。HIF-1α抑制剂(2ME2)能够减轻喹啉酸致SH-SY5Y细胞损伤程度,减少细胞凋亡。
To investigate the role of hypoxia inducible factor-1α (HIF-1α) in quinolinic acid-induced injury of human neuroblastoma SH-SY5Y cells. Human neuroblastoma SH-SY5Y cells cultured in vitro were divided into control group, low, medium and high-dose quinolinic acid groups and 2-methoxyestradiol (2ME2) pretreatment Quinolinic acid high dose group, the degree of cell damage was determined by the method of thiazolyl blue reduction and lactate dehydrogenase leakage rate detection. Hoechst 33342 single fluorescent staining was used to observe the apoptosis. The expression of HIF-1α in the cells was detected by immunofluorescence staining The expression of HIF-1α, protein kinase B (Akt), phosphorylated Akt (p-Akt), Bcl-2 and Bax were detected by Western blotting. The results showed that quinolinic acid could induce SH-SY5Y cell injury in a time-and dose-dependent manner, leading to apoptosis. Meanwhile, quinolinic acid up-regulated the expression of HIF-1α in SH-SY5Y cells and led to nuclear translocation. The expression of p-Akt increased and the ratio of Bax / Bcl-2 expression increased while 2ME2 inhibited quinolinic acid-induced SH-SY5Y cells Damage and reduce the expression of HIF-1α, p-Akt and Bax / Bcl-2. Thus, HIF-1α / Akt pathway mediates quinolinic acid-induced apoptosis in SHSY5Y cells. HIF-1α inhibitor (2ME2) can reduce the degree of injury of SH-SY5Y cells induced by quinolinic acid and reduce apoptosis.