中华皮肤科杂志
中華皮膚科雜誌
중화피부과잡지
Chinese Journal of Dermatology
2014年
12期
868-872
,共5页
nutlin-3%黑色素瘤,实验性%细胞周期%细胞凋亡%细胞运动
nutlin-3%黑色素瘤,實驗性%細胞週期%細胞凋亡%細胞運動
nutlin-3%흑색소류,실험성%세포주기%세포조망%세포운동
Nutlin-3%Melanoma,experimental%Cell cycle%Apoptosis%Cell movement
目的 观察顺式咪唑啉衍生物nutlin-3对人A375黑素瘤细胞生物学行为的影响,研究其机制.方法 将A375细胞分为实验组和对照组,实验组细胞接受2.5、5、10 μmol/L nutlin-3处理,对照组细胞采用二甲基亚砜处理.分别于作用24、48、72 h后,噻唑蓝(MTT)法检测细胞增殖情况;Western印迹检测p53蛋白表达的改变;流式细胞仪检测细胞周期分布以及细胞凋亡;Transwell法检测迁移性变化.采用重复测量的方差分析进行统计学检验.结果 2.5、5、10μmol/L nutlin-3处理A375细胞24、48、72 h后,MTT法显示不同时间点间的增殖抑制率差异有统计学意义(F=67.43,P＜ 0.01),不同浓度间的抑制率差异有统计学意义(F=135.58,P＜ 0.01),浓度越高抑制率越高;时间和浓度之间有交互作用(F=26.95,P＜0.01).Western印迹、流式细胞仪、Transwell法检测显示,不同时间点之间A375细胞的p53表达、G2期细胞百分率、凋亡率、迁移抑制率差异均有统计学意义(F值分别为1255.00、831.38、809.45、1100.00,均P＜0.01),除p53外,时间越长各项指标值越高;不同浓度间各项指标值差异均有统计学意义(F值分别为9196.00、267.99、723.83、1667.00,均P＜0.01),浓度越高各项指标值越高;时间与浓度之间交互作用有统计学意义(F值分别为826.79、21.602、44.48、313.09,均P＜ 0.01).结论 Nutlin-3可能通过p53蛋白积累途径抑制人A375细胞的增殖和迁移,促进细胞周期停滞和细胞凋亡.
目的 觀察順式咪唑啉衍生物nutlin-3對人A375黑素瘤細胞生物學行為的影響,研究其機製.方法 將A375細胞分為實驗組和對照組,實驗組細胞接受2.5、5、10 μmol/L nutlin-3處理,對照組細胞採用二甲基亞砜處理.分彆于作用24、48、72 h後,噻唑藍(MTT)法檢測細胞增殖情況;Western印跡檢測p53蛋白錶達的改變;流式細胞儀檢測細胞週期分佈以及細胞凋亡;Transwell法檢測遷移性變化.採用重複測量的方差分析進行統計學檢驗.結果 2.5、5、10μmol/L nutlin-3處理A375細胞24、48、72 h後,MTT法顯示不同時間點間的增殖抑製率差異有統計學意義(F=67.43,P＜ 0.01),不同濃度間的抑製率差異有統計學意義(F=135.58,P＜ 0.01),濃度越高抑製率越高;時間和濃度之間有交互作用(F=26.95,P＜0.01).Western印跡、流式細胞儀、Transwell法檢測顯示,不同時間點之間A375細胞的p53錶達、G2期細胞百分率、凋亡率、遷移抑製率差異均有統計學意義(F值分彆為1255.00、831.38、809.45、1100.00,均P＜0.01),除p53外,時間越長各項指標值越高;不同濃度間各項指標值差異均有統計學意義(F值分彆為9196.00、267.99、723.83、1667.00,均P＜0.01),濃度越高各項指標值越高;時間與濃度之間交互作用有統計學意義(F值分彆為826.79、21.602、44.48、313.09,均P＜ 0.01).結論 Nutlin-3可能通過p53蛋白積纍途徑抑製人A375細胞的增殖和遷移,促進細胞週期停滯和細胞凋亡.
목적 관찰순식미서람연생물nutlin-3대인A375흑소류세포생물학행위적영향,연구기궤제.방법 장A375세포분위실험조화대조조,실험조세포접수2.5、5、10 μmol/L nutlin-3처리,대조조세포채용이갑기아풍처리.분별우작용24、48、72 h후,새서람(MTT)법검측세포증식정황;Western인적검측p53단백표체적개변;류식세포의검측세포주기분포이급세포조망;Transwell법검측천이성변화.채용중복측량적방차분석진행통계학검험.결과 2.5、5、10μmol/L nutlin-3처리A375세포24、48、72 h후,MTT법현시불동시간점간적증식억제솔차이유통계학의의(F=67.43,P＜ 0.01),불동농도간적억제솔차이유통계학의의(F=135.58,P＜ 0.01),농도월고억제솔월고;시간화농도지간유교호작용(F=26.95,P＜0.01).Western인적、류식세포의、Transwell법검측현시,불동시간점지간A375세포적p53표체、G2기세포백분솔、조망솔、천이억제솔차이균유통계학의의(F치분별위1255.00、831.38、809.45、1100.00,균P＜0.01),제p53외,시간월장각항지표치월고;불동농도간각항지표치차이균유통계학의의(F치분별위9196.00、267.99、723.83、1667.00,균P＜0.01),농도월고각항지표치월고;시간여농도지간교호작용유통계학의의(F치분별위826.79、21.602、44.48、313.09,균P＜ 0.01).결론 Nutlin-3가능통과p53단백적루도경억제인A375세포적증식화천이,촉진세포주기정체화세포조망.
Objective To estimate the effect of a cis-imidazoline derivative,nutlin-3,on the biological behavior of A375 human melanoma cells,and to investigate its mechanism.Methods Cultured A375 cells were divided into several test groups treated with nutlin-3 at different concentrations (2.5,5,10 μmol/L) for 24,48 and 72 hours,and a control group treated with dimethyl sulfoxide (DMSO) only.Then,methyl thiazolyl tetrazolium (MTT) assay was performed to evaluate cellular proliferative activity,Western blot to measure the expression of p53 protein,flow cytometry to estimate cell cycle phase distribution and apoptosis rate,and Transwell assay to evaluate migratory activity,of A375 cells.Statistical analysis was carried out by repeated-measures analysis of variance (ANOVA).Results After treatment with nutlin-3 of 2.5,5 and 10 μmol/L for 24,48 and 72 hours,significant differences were observed among different time points at each concentration and among different concentrations at the same time point in proliferation inhibition rate (F =67.43,135.58,respectively,both P ＜ 0.01),p53 protein expression level (F =1255.00,9196.00,respectively,both P ＜ 0.01),percentage of cells at G2 phase (F =831.38,267.99,respectively,both P ＜ 0.01),apoptosis rate (F =809.45,723.83,respectively,both P ＜ 0.01),migration inhibition rate (F =1100.00,1667.00,respectively,both P ＜ 0.01).The influence of nutlin-3 on cellular proliferative activity increased with the increase in its concentration,and that on percentage of cells at G2 phase,apoptosis rate and migratory activity increased with the increase in its concentration and treatment duration.There was a significant interaction between the treatment duration and concentration of nutlin-3 for p53 protein expression level in (F =826.79,P ＜ 0.01),percentage of cells at G2 phase in (F =21.602,P ＜ 0.01),apoptosis rate in (F =44.48,P ＜ 0.01),migratory activity of (F =313.09,P ＜ 0.01),and cellular proliferative activity of (F =26.95,P ＜ 0.01),A375 cells.Conclusion Nutlin-3 may inhibit the proliferation and migration of,but promote cell cycle arrest and apoptosis in,A375 cells,through accumulation of p53 protein.
