CAJ | 학술논문

目的探讨沙利度胺联合干扰素(IFN)对急性髓系白血病细胞系Kasumi-1细胞的增殖抑制作用及其机制.方法沙利度胺、IFN以及两药联合处理Kasumi-1细胞,采用CCK-8法检测细胞增殖抑制率,流式细胞术检测细胞凋亡率,ELISA法检测细胞培养上清血管内皮生长因子(VEGF)水平,Western blot法检测凋亡相关蛋白的表达.结果在50～ 500 μg/ml范围内沙利度胺可抑制Kasumi-1细胞增殖,并呈浓度依赖性[24 h的IC50值为(451.13±6.92)μg/ml、48 h的IC50值为(362.50±14.52) μg/ml];在500～5 000 U/ml范围内,IFN浓度依赖性地抑制Kasumi-1细胞增殖[24 h的IC50值为(2 209±127)U/ml、48 h的IC50值为(1 393±63)U/ml].350 μg/ml沙利度胺和1 400 U/ml IFN作用48h,Kasumi-1细胞的凋亡率分别为(14.68±2.61)％和(21.71±0.71)％,与对照组[(0.89±0.38)％]相比,差异有统计学意义(P值均＜0.01);两药联合作用后Kasumi-1细胞的增殖抑制率为(88.50±2.40)％,凋亡率为(41.95±3.41)％,与对照组和各单药组比较,差异均有统计学意义(P值均＜0.01).沙利度胺、IFN单药及两药联合组VEGF水平分别为(141.11±3.70)、(119.90±2.00)和(94.61±5.46)ng/L,两药联合组明显低于单药组(P值均＜ 0.05).沙利度胺、IFN单药作用Kasumi-1细胞48 h能不同程度下调Bcl-2的表达,上调p-P38(同时P38表达量下降)、Bax、细胞色素C、活化型(cleaved)Caspase-3、8、9的表达,与对照组比较差异均有统计学意义(P值均＜0.05);沙利度胺联合IFN该作用进一步加强(P值均＜0.05).结论沙利度胺和IFN可协同抑制Kasumi-1细胞增殖、诱导其凋亡,其可能是通过线粒体和死亡受体途径,并活化P38信号通路诱导细胞凋亡和抑制Kasumi-1细胞VEGF的自分泌来实现.
목적 탐토사리도알연합간우소(IFN)대급성수계백혈병세포계Kasumi-1세포적증식억제작용급기궤제.방법 사리도알、IFN이급량약연합처리Kasumi-1세포,채용CCK-8법검측세포증식억제솔,류식세포술검측세포조망솔,ELISA법검측세포배양상청혈관내피생장인자(VEGF)수평,Western blot법검측조망상관단백적표체.결과 재50～ 500 μg/ml범위내사리도알가억제Kasumi-1세포증식,병정농도의뢰성[24 h적IC50치위(451.13±6.92)μg/ml、48 h적IC50치위(362.50±14.52) μg/ml];재500～5 000 U/ml범위내,IFN농도의뢰성지억제Kasumi-1세포증식[24 h적IC50치위(2 209±127)U/ml、48 h적IC50치위(1 393±63)U/ml].350 μg/ml사리도알화1 400 U/ml IFN작용48h,Kasumi-1세포적조망솔분별위(14.68±2.61)％화(21.71±0.71)％,여대조조[(0.89±0.38)％]상비,차이유통계학의의(P치균＜0.01);량약연합작용후Kasumi-1세포적증식억제솔위(88.50±2.40)％,조망솔위(41.95±3.41)％,여대조조화각단약조비교,차이균유통계학의의(P치균＜0.01).사리도알、IFN단약급량약연합조VEGF수평분별위(141.11±3.70)、(119.90±2.00)화(94.61±5.46)ng/L,량약연합조명현저우단약조(P치균＜ 0.05).사리도알、IFN단약작용Kasumi-1세포48 h능불동정도하조Bcl-2적표체,상조p-P38(동시P38표체량하강)、Bax、세포색소C、활화형(cleaved)Caspase-3、8、9적표체,여대조조비교차이균유통계학의의(P치균＜0.05);사리도알연합IFN해작용진일보가강(P치균＜0.05).결론 사리도알화IFN가협동억제Kasumi-1세포증식、유도기조망,기가능시통과선립체화사망수체도경,병활화P38신호통로유도세포조망화억제Kasumi-1세포VEGF적자분비래실현.
Objective To explore the inhibitory effect of thalidomide combined with interferon (IFN) on the human acute myeloid leukemia cell line Kasumi-1 and its mechanism.Methods The inhibitiory effect of Kasumi-1 cells by thalidomide,interferon or combination was detected by CCK-8 method,the apoptosis by flow cytometry,the expression of apoptosis related proteins by Western blot,vascular endothelial growth factor (VEGF) concentration in culture supernatant by ELISA.Results Thalidomide inhibited the proliferation of Kasumi-1 in a dose-dependent manner from 50 μg/ml to 500 μg/ml with an IC50 of (451.13±6.92)μg/ml at 24 h and (362.50± 14.52)μg/ml at 48 h.IFN also demonstrated the inhibitory capacity in a dose-dependent manner from 500 U/ml to 5 000 U/ml,with an IC50 of(2 209±127)U/ml at 24 h and(1 393±63)U/ml at 48 h.The apoptosis rates of Kasumi-1 cells treated with thalidomide 350 μg/ml or IFN 1 400 U/ml for 48 h were (14.68±2.61)％ and (21.71 ±0.71)％,respectively,significantly higher than control group (P＜0.01).In combination group the inhibition and the apoptosis rate were (88.50±2.40)％ and (41.95±3.41)％,significantly higher than control and each single agent group (P＜0.01).The VEGF concentrations of combination group [(94.61 ± 5.46) ng/L] decreased significantly,as compared to thalidomide group [(141.11±3.70) ng/L] and IFN group [(119.90±2.00) ng/L] (P ＜ 0.05).Western blot analysis showed Bcl-2 expression of Kasumi-1 cells decreased,while p-P38,Bax,cytochrome C,cleaved-Caspase-3,8,9 increased after treated with thalidomide 350 μg/ml or IFN 1 400 U/ml for 48 h.When treated with the combination agents,the expression of Bcl-2 further decreased and p-P38,Bax,cytochrome C,cleaved-Caspase-3,8,9 further increased as compared with each single agent (P ＜ 0.05).Conclusions Thalidomide and IFN could synergistically inhibit the proliferation of Kasumi-1 cells probably through inducing apoptosis via the mitochondrial pathway,death receptor pathway and P38 MAPK pathway,as well as inhibiting VEGF secretion.