二十二碳六烯酸对体外大鼠C6胶质瘤细胞的促凋亡作用

Pro-apoptosis effects of docosahexaenoic acid on rat C6 glioma cells in vitro

目的探讨二十二碳六烯酸(DHA)对大鼠C6胶质瘤细胞的促凋亡作用。方法 10、25、50、75和100μmol/L DHA处理C6细胞24、48和72 h后检测细胞活力;100μmol/L DHA处理C6细胞24 h后,应用TUNEL方法检测凋亡细胞,流式细胞术检测凋亡细胞的比例;应用免疫荧光和Western blotting方法检测剪切后含半胱氨酸的天冬氨酸蛋白水解酶3(cleaved Caspase-3)在凋亡细胞中的表达情况。结果 25、50、75和100μmol/L DHA处理C6细胞24、48和72 h后,抑制细胞活力;100μmol/L DHA处理细胞24 h后,能够检测到TUNEL阳性的凋亡细胞;流式细胞术检测后发现,DHA能够明显上调凋亡细胞的比例;此外,细胞经100μmol/L DHA处理后,免疫荧光能够检测到cleaved Caspase-3阳性细胞,Western blotting能够检测到cleaved Caspase-3阳性蛋白条带。结论 DHA具有促进大鼠C6胶质瘤细胞凋亡的作用。

Objective To clarify the pro-apoptotic effects of docosahexaenoicacid（ DHA） on rat C6 glioma cells in vitro. Methods After treatment of 10,25,50,75 and 100 μmol/L DHA for 24,48 and 72 hours,C6 cell viability was detected. After treatment of 100 μmol/L DHA for 24 hours, apoptotic cells were labeled by the terminaldeoxynucleotidyl transferase mediated nick end labeling（ TUNEL） method,and the proportion of apoptotic cells was counted by flow cytometry. The expression of cleaved Caspase-3 in apoptotic cells was detected by immunocytochemistry and Western blotting. Results 25,50,75 and 100 μmol/L DHA inhibited the C6 cell viability at 24,48 and 72 hours. The TUNEL positive cells were found after treatment of 100 μmol/L DHA for 24 hours. Flow cytometry analysis showed that the proportion of apoptotic cells was increased. Moreover,after treatment of 100 μmol/L DHA,cleaved Caspase-3 positive cells were found by immunocytochemistry. Cleaved Caspase-3 positive protein band was investigated by Western blotting.Conclusion DHA may induce the apoptosis of C6 cells in vitro.