雷公藤内酯酮对SHI-1细胞增殖及凋亡的影响

Effect of Triptonide on Proliferation and Apoptosis of SHI-1 Cells

目的:探讨雷公藤内酯酮(TN)对人急性单核细胞白血病(AML)细胞株SHI-1细胞增殖、周期、凋亡以及凋亡相关蛋白表达水平的影响,并探究其可能的作用机制。方法:采用噻唑蓝(MTT)比色法检测20,40,80,160,320 nmol·L^(-1)的雷公藤内酯酮作用SHI-1细胞48,72 h对细胞增殖的抑制作用,流式细胞术碘化丙啶(PI)单染法检测雷公藤内酯酮作用前后SHI-1细胞周期的变化,流式细胞术AnnexinV/PI双染法检测雷公藤内酯酮作用前后SHI-1细胞凋亡的变化,蛋白免疫印迹法(Western blot)检测80,160 nmol·L^(-1)雷公藤内酯酮作用于SHI-1细胞前后半胱氨酸蛋白酶-3(Caspase-3),半胱氨酸蛋白酶-8(Caspase-8)和核转录因子-κB(NF-κB)的蛋白表达。结果:分别作用48,72 h,与空白组比较,40,80,160,320 nmol·L^(-1)雷公藤内酯酮对SHI-1细胞增殖均具有显著抑制作用(P<0.01),且具有剂量依赖性;雷公藤内酯酮160,320 nmol·L^(-1)可诱导SHI-1细胞凋亡(P<0.01),并且雷公藤内酯酮160 nmol·L^(-1)可使SHI-1细胞S期比例降低(P<0.01)。此外,与空白组比较,雷公藤内酯酮80,160 nmol·L^(-1)组NF-κB蛋白表达水平显著下调(P<0.05,P<0.01),雷公藤内酯酮160 nmol·L^(-1)组Caspase-3,Caspase-8蛋白表达水平显著下调(P<0.01)。结论:雷公藤内酯酮可以体外抑制SHI-1细胞增殖并诱导其凋亡,其作用机制可能与雷公藤内酯酮降低SHI-1细胞S期比例,下调Caspase-3,Caspase-8以及NF-κB的蛋白表达水平有关。

Objective:This study aims to investigate the effect of triptonide(TN)on proliferation,cell cycle,apoptosis and expressions of apoptosis-related proteins of human acute monocytic leukemia(AML)cell line SHI-1,and to explore its possible mechanism of action. Method: The thiazolyl blue(MTT)colorimetric assay was applied to detect the inhibitory effect of 20,40,80,160,320 nmol·L^(-1) triptonide on the proliferation of SHI-1 cells for 48,72 h. Changes in SHI-1 cell cycle before and after triptonide treatment were detected by flow cytometry propidium iodide(PI)simple staining,and changes in SHI-1 cell apoptosis before and after triptonide treatment were detected by flow cytometry with AnnexinV/PI double staining. Western blot was applied to detect the protein expression of cysteine protease(Caspase)-3,Caspase-8 and nuclear transcription factor kappaB(NF-κB)in SHI-1 cells before and after treatment with 80,160 nmol·L^(-1) triptonide. Result: Compared with the blank group,40,80,160,320 nmol·L^(-1) triptonide significantly inhibited the proliferation of SHI-1 cells(P<0.01)in a dose-dependent manner for 48,72 h,while 160,320 nmol·L^(-1) triptonide induced apoptosis of SHI-1 cells(P<0.01)for 48,72 h,and 160 nmol·L^(-1) triptonide could decrease the S phase ratio of SHI-1 cells(P<0.01). In addition,compared with the blank group,80,160 nmol·L^(-1) triptonide induced the downregulation of NF-κB significantly(P<0.01), 160 nmol·L^(-1) triptonide induced the downregulation of Caspase-3, Caspase-8 significantly(P<0.01). Conclusion:Triptonide can inhibit the proliferation and induce apoptosis in vitro of SHI-1 cells,which may be related to the reduction of the cells in S phase proportion by triptonide,and the downregulation of the expression levels of Caspase-3,Caspase-8 and NF-κB proteins.