CAJ | 학술논문

目的研究亚砷酸诱导鼻咽癌CNE-2Z细胞株中抑癌基因Ras相关结构域家族1A基因(RAS association domain family gene 1A,RASSFIA)的表达.方法体外培养的CNE-2Z细胞分别加入不同浓度的亚砷酸,并作用不同时间.应用台盼蓝染色法筛选出亚砷酸抑制CNE-2Z细胞生长的最佳浓度,检测IC_(50)值;流式细胞术检测细胞周期的改变;用终浓度为2 μmol/L、1 μmol/L、0.5 μmol/L的亚砷酸加入鼻咽癌CNE-2Z细胞系和不加药物的空白对照组,作用48 h后,采用甲基化特异性PCR法检测亚砷酸处理前后RASSF1A基因去甲基化的情况,采用反转录PCR检测鼻咽癌CNE-2Z细胞系中RASSFIA基因亚砷酸处理前后mRNA水平的表达情况,采用Western blot方法检测亚砷酸处理前后RASSF1A基因蛋白质水平的表达情况.结果亚砷酸对细胞增殖的抑制作用具有明显的时间和剂量效应.不同浓度的亚砷酸作用细胞24 h、48 h、72 h后,其IC_(50)值分别为(1.50±0.05)、(1.09±0.13)、(0.65±0.04)μmol/L,亚砷酸使细胞周期阻滞于S期和G2/M期;RASSF1A经亚砷酸作用后甲基化随着作用浓度的增高逐渐减弱,而去甲基化随着作用浓度的增高逐渐增强,且亚砷酸作用后RASSF1A在mRNA水平和蛋白质水平表达明显增强.空白对照组和不同浓度亚砷酸作用CNE-2Z细胞的处理组之间差异均有统计学意义(P值均<0.05),不同浓度亚砷酸作用CNE-2Z细胞的处理组之间差异均有统计学意义(P值均<0.05).结论亚砷酸可诱导鼻咽癌CNE-2Z细胞株中RASSF1A表达增强,从而阻滞鼻咽癌细胞的增殖分化.
목적 연구아신산유도비인암CNE-2Z세포주중억암기인Ras상관결구역가족1A기인(RAS association domain family gene 1A,RASSFIA)적표체.방법 체외배양적CNE-2Z세포분별가입불동농도적아신산,병작용불동시간.응용태반람염색법사선출아신산억제CNE-2Z세포생장적최가농도,검측IC_(50)치;류식세포술검측세포주기적개변;용종농도위2 μmol/L、1 μmol/L、0.5 μmol/L적아신산가입비인암CNE-2Z세포계화불가약물적공백대조조,작용48 h후,채용갑기화특이성PCR법검측아신산처리전후RASSF1A기인거갑기화적정황,채용반전록PCR검측비인암CNE-2Z세포계중RASSFIA기인아신산처리전후mRNA수평적표체정황,채용Western blot방법 검측아신산처리전후RASSF1A기인단백질수평적표체정황.결과 아신산대세포증식적억제작용구유명현적시간화제량효응.불동농도적아신산작용세포24 h、48 h、72 h후,기IC_(50)치분별위(1.50±0.05)、(1.09±0.13)、(0.65±0.04)μmol/L,아신산사세포주기조체우S기화G2/M기;RASSF1A경아신산작용후갑기화수착작용농도적증고축점감약,이거갑기화수착작용농도적증고축점증강,차아신산작용후RASSF1A재mRNA수평화단백질수평표체명현증강.공백대조조화불동농도아신산작용CNE-2Z세포적처리조지간차이균유통계학의의(P치균<0.05),불동농도아신산작용CNE-2Z세포적처리조지간차이균유통계학의의(P치균<0.05).결론 아신산가유도비인암CNE-2Z세포주중RASSF1A표체증강,종이조체비인암세포적증식분화.
Objective To investigate the effect of arsenic trioxide (As_2O_3) on expression of anti-oncogene RAS association domain family gene 1A(RASSF1A) in nasopharyngeal carcinoma cell line CNE-2Z. Methods CNE-2Z cells were treated with various concentrations of As_2O_3 for different times. The IC_(50) values were detected by trypan blue stain assay. Cell cycle redistribution was analyzed by flow cytometry. The final concentration 2 μmol/L,1 μmol/L,0. 5 μmol/L of As_2O_3 was added to CNE-2Z cell for succedent experiments. The controls and no drugs of CNE-2Z cells were cultivated for 48 h. Methylation specific PCR was used to detect the change of methylation status of RASSF1A gene. The expression of RASSF1A gene was detected by reverse transcription PCR and Western blot at mRNA and protein level. Results The suppression of cell proliferation by As_2O_3 was time and dose-dependent. After being treated with As_2O_3,the IC_(50) values of As_2O_3 were (1.50±0.05), (1.09±0.13), (0.65±0.04) μmol/L at 24, 48, and 72 h, respectively. As_2O_3 also arrested CNE-2Z cells in G2/M phase of cell cycle. After the effect of As_2O_3, the methylation of RASSF1A gene became weaker by increasing the concentration of As_2O_3; and the expression of RASSF1A gene became stronger at mRNA and protein level. Between different concentration of As_2O_3 group and no drugs group, the differences had statistical significance (P<0.05). Along with increasing the concentration of As_2O_3, the expression of RASSF1A gene became stronger at mRNA and protein level, the methylation of RASSF1A gene became weaker and weaker. Conclusions As_2O_3 can activate the expression of RASSF1A gene to inhibit the cell cyc]e progress of uasopharyngeal carcinoma cell line.