CAJ | 학술논문

目的探讨组蛋白去乙酰化酶抑制剂Apicidin对胶质瘤细胞系U87 Nanog和p53表达及细胞增殖、周期、凋亡的影响. 方法体外常规培养U87细胞,MTT检测0.2、0.5、1.0、2.0μmol/Apicidin处理24、48、72 h后细胞抑制率的变化,同时设对照组,RT-PCR和Western blotting法检测1.0μmol/LApicidin处理48 h后细胞Nanog与p53 mRNA和蛋白水平的改变.DAPI染色检测1.0 μmol/LApicidin作用后细胞凋亡的改变;流式细胞仪检测0.2、0.5、1.0 μmol/LApicidin作用48 h后U87细胞凋亡率和细胞周期的变化. 结果 MTT显示不同浓度Apicidin处理后细胞生长出现抑制,并呈剂量、时间依赖性,48 h细胞增殖的半数抑制浓度(IC50)为(1.74±0.13)μmol/L;RT-PCR检测发现1.0μmol/LApicidin组细胞Nanog mRNA表达较对照组降低,p53 mRNA表达增高.Western blotting检测显示与对照组比较,Nanog蛋白水平降低,而p53蛋白水平增高,差异有统计学意义(P＜0.05).DAPI染色发现1.0μmol/LApicidin组细胞核出现凋亡表现;流式细胞仪检测发现,0.5、1.0μmol/LApicidin作用48 h后U87细胞凋亡率为11.57％、19.67％,高于对照组(2.2％).S期细胞比例分别为(42.92±0.56)％、(56.95±0.53)％,高于对照组的(32.68±0.49)％,差异有统计学意义(P＜0.05). 结论 Apicidin可以显著抑制U87胶质瘤细胞系生长,诱导细胞周期阻滞和凋亡产生,其机制可能与抑制干细胞标志性基因Nanog的表达和抑癌基因p53的转录及蛋白表达增加有关.
목적 탐토조단백거을선화매억제제Apicidin대효질류세포계U87 Nanog화p53표체급세포증식、주기、조망적영향. 방법 체외상규배양U87세포,MTT검측0.2、0.5、1.0、2.0μmol/Apicidin처리24、48、72 h후세포억제솔적변화,동시설대조조,RT-PCR화Western blotting법검측1.0μmol/LApicidin처리48 h후세포Nanog여p53 mRNA화단백수평적개변.DAPI염색검측1.0 μmol/LApicidin작용후세포조망적개변;류식세포의검측0.2、0.5、1.0 μmol/LApicidin작용48 h후U87세포조망솔화세포주기적변화. 결과 MTT현시불동농도Apicidin처리후세포생장출현억제,병정제량、시간의뢰성,48 h세포증식적반수억제농도(IC50)위(1.74±0.13)μmol/L;RT-PCR검측발현1.0μmol/LApicidin조세포Nanog mRNA표체교대조조강저,p53 mRNA표체증고.Western blotting검측현시여대조조비교,Nanog단백수평강저,이p53단백수평증고,차이유통계학의의(P＜0.05).DAPI염색발현1.0μmol/LApicidin조세포핵출현조망표현;류식세포의검측발현,0.5、1.0μmol/LApicidin작용48 h후U87세포조망솔위11.57％、19.67％,고우대조조(2.2％).S기세포비례분별위(42.92±0.56)％、(56.95±0.53)％,고우대조조적(32.68±0.49)％,차이유통계학의의(P＜0.05). 결론 Apicidin가이현저억제U87효질류세포계생장,유도세포주기조체화조망산생,기궤제가능여억제간세포표지성기인Nanog적표체화억암기인p53적전록급단백표체증가유관.
Objective To investigate the effect of histone deacetylase inhibitor Apicidin on apoptosis,cycle,proliferation and Nanog and p53 expressions of U87 glioblastoma cell line.Methods U87 cells were cultured routinely in vitro; MTT assay was employed to detect the changes of cell inhibition after 0.2,0.5,1.0 and 2.0 μmol/L Apicidin treatment; RT-PCR and Western blotting were employed to measure the mRNA and protein expressions of Nanog and p53 in cells of the Apicidin treatment group (treated with 1.0 μmol/L Apicidin for 48 h) and control group; DAPI staining was employed to observe the changes of cell apoptosis after being treated with 1.0 μmol/L Apicidin; cell cycle arrest and apoptosis rate were examined by flow cytometry after being treated with 0.2,0.5 and 1.0 μmol/L Apicidin for 48 h.Results MTT assay showed that the cell growth was inhibited after being treated with different doses of Apicidin,with dose-and time-dependent manners; the median inhibitory concentration (IC50) of proliferation 48 hafter treatment was approximately 1.74±0.13 μmol/L.RT-PCR showed that 1.0 μmol/L Apicidin treatment group had significantly lower Nanog mRNA expression and higher p53 mRNA expression than control group (P＜0.05); Western blotting showed that 1.0 μmol/L Apicidin treatment group had significantly lower Nanog protein expression and higher p53 protein expression than control group (P＜0.05).DAPI staining showed apoptotic nuclei in cells of 1.0 μ mol/L Apicidin treatment group.Flow cytometry indicated that the apoptosis rate in the 0.5 and 1.0 μmol/L Apicidin treatment groups (11.57 ％ and 19.67 ％) was significantly higher than that in the control group (2.2 ％).The percentage of cells being arrested at S phase was 42.92±0.56％ and 56.95±0.53％ in the 0.5 and 1.0 μmol/L Apicidin treatment groups,which was significantly higher than that in the control group (32.68±0.49％,P＜0.05).Conclusion Histone deacetylase inhibitor Apicidin could sequentially repress the proliferation of glioma cell line U87 and induce cell cycle arrest and apoptosis in U87 cells,whose molecular mechanisms might relate to activate the tumor suppressor p53 expression and inhibit Nanog expression.