rhTRAIL对黑色素瘤A-375细胞凋亡的影响

Effects of rhTRAIL on apoptosis of melanoma cell line A-375

目的:研究重组人肿瘤坏死因子相关的凋亡诱导配体(rhTRAIL)对黑色素瘤A-375细胞生长及诱导细胞凋亡的影响,并探讨其可能机制。方法:将培养的黑色素瘤A-375细胞分为对照组和rhTRAIL低、中、高剂量组(0.25、0.50、1.00mg·L-1),经不同剂量rhTRAIL作用后,观察细胞形态;MTT法检测细胞存活分数;TUNEL法检测细胞凋亡率;流式细胞仪检测细胞凋亡及细胞周期变化。结果:对照组细胞呈梭形贴壁或半贴壁生长,rhTRAIL组一部分细胞变圆,贴壁不佳,细胞数低于对照组。与对照组比较,rhTRAIL明显抑制黑色素瘤A-375的生长,1.00mg·L-1组的生长抑制率最高,0.50mg·L-1组次之,0.25mg·L-1组最低,3组间比较差异有显著性(P<0.05)。流式细胞检测结果显示,rhTRAIL蛋白可诱导A-375细胞凋亡,rhTRAIL低、中、高剂量组细胞凋亡率分别为6.68%、24.59%和28.91%,且凋亡率随剂量增大而升高,3组间比较差异有显著性(P<0.05)。rhTRAIL低、中、高剂量组细胞处于G1期的百分率(45.26%、60.67%和69.10%)明显高于对照组(37.41%)(P<0.05)。结论:rhTRAIL对A-375细胞生长有显著的抑制作用,呈剂量依赖性,其机制可能与细胞周期阻滞、诱导细胞凋亡有关。

Objective To investigate the effect of recombinant human TNF-related apoptosis-inducing ligand （rhTRAIL） protein on the growth and apoptosis of melanoma cell line A-375. Methods The A-375 cells were divided into four groups including control group and rhTRAIL 0.25, 0.50, and 1.00 mg · L^-1 groups and the morphological changes of A-375 cells were observed by contrast phase microscope after treated with different doses of rhTRAIL. The survival fraction of A-375 cells was measured by MTT assay; Apoptotic rate was determined by TUNEL method and the cell cycle and apoptosis index were analyzed by technique of flow cytometry （FCM）. Results The cells in control group were fusiform adherence or half adherence and abundant, the state of some cells in rhTRAIL group was worse than that in control group. The inhibitory rates of growth in rhTRAIL groups with different doses （0.25, 0.50, 1.00 mg · L^-1 ） were （15.36±3.21） %, （27.34±2.53） %, and （43.68±3.51） %, respectively, there were significant differences between three groups （P〈0.05） . The FCM results showed that the rhTRAIL could induce the apoptosis in dose-dependent manner, and the apoptotic rates were 6.68 %, 24.59 %, and 28.91 %, respectively, when the rhTRAIL doses were 0.25, 0. 50, and 1.00 mg · L^-1, there were significant differences between three groups （P〈0.05）. The GI phase cell propotions in rhTRAIL 0.25, 0.50,and 1.00 mg · L^-1 groups （45.26%, 60.67%, 69.10 %） were higher than that in control group （37.41%） . Conclusion The rhTRAIL has the significant effect on inhibiting the growth of A-375 cells, and the inhibitory rate is dose-dependent. The mechanism may be concerned with cell cycle arrest and inducing apoptosis upon A-375 cells.