氨基葡萄糖硫酸盐诱导K562细胞凋亡的实验研究  

Study on the Apoptosis of K562 Cells Induced by Glucosamine Sulfate

作　　者：梁蓉[1] 王哲[1] 黄高[1] 乔岩[2] 王爱勤[2] 董宝侠[1] 郭英[1] 王娟红[1] 

机构地区：[1]第四军医大学西京医院,710033 [2]中国科学院兰州化学物理研究所,730000

出　　处：《实用癌症杂志》2005年第1期20-23,27,共5页The Practical Journal of Cancer

基　　金：国家自然科学基金资助项目 (No .30 30 0 1 4 1 )

摘　　要：目的　探讨氨基葡萄糖硫酸盐(GS)对慢性髓系白血病K 5 62细胞凋亡诱导作用和分子机理。方法　采用台盼蓝拒染法测定细胞生长曲线来观察GS对K5 62细胞的生长抑制作用;取对数生长期K 5 62细胞,在0 .5 ,1.0 ,5 .0mmol/LGS不同浓度作用48h后,利用细胞计数、电镜、吖啶橙/溴化乙啶(AO/EB)染色、DNA梯状电泳、流式细胞仪等方法来观察GS对K 5 62细胞的抑制效应和促凋亡作用。并用Westernblot检测活化的caspase 3和bcl- 2基因的表达。结果　0 .5 ,1.0 ,5 .0mmol/LGS对K 5 62细胞的生长有抑制作用,并呈时间和剂量依赖性(P <0 .0 1)。采用光镜、电镜和AO /EB染色法发现5mmol/LGS作用K 5 62细胞后,出现典型的凋亡形态学改变。AnnexinV染色后流式细胞仪能检测到早期凋亡细胞,细胞周期显示:G1期细胞比例增高,S期比例减低,并有凋亡峰。有明显的梯状DNA。同时GS诱导K 5 62细胞凋亡后出现活化的Caspase 3 ,而Bcl- 2蛋白表达减弱,甚至消失。结论　GS能诱导白血病K- 5 62细胞凋亡,该过程伴有Caspase- 3的活化和Bcl- 2蛋白表达降低。Objective To investigate the effect and mechanisms of glucosamine sulfate(GS) on chronic myloid leukemia K562 cells.Methods Exposured K562 cells in an exponential growth to 0.5,1.0,5.0 mmol/L GS and then test the inhibitive effect by cell counting.Morphologic evidence for apoptosis of GS inducing on K562 cells was determined by transmission electron microscope.The detection of DNA Ladder was tested by agarose gel electrophoresis.Sub-G1,cell cycle and Annexin V was detected by flow cytometry.Western blot was performed to detect the expression of activated caspase-3 and bcl-2.Results Treatment with 0.5,1.0, 5.0 mmol/L GS resulted in a concentration-dependent reduction in K562 cells growth(P<0.01).The characteristic morphological changes of apoptosis were observed in K562 cells treated by GS by Wright-Gimsa and AO/EB staining.The proportion of G 1 cells was increased and the sub-G1 phase was 5.9%.K562 cells treated by GS were observed to have characteristic morphological changes of apoptosis.Treatment of K562 cells with 5 mM GS for 2～3 days resulted in the induction of apoptosis as assayed qualitatively by agarose gel electrophoresis and quantitatively by Annexin V.K562 cells treated by GS for 2-3 days expressed activated Caspase-3.However,the expression of Bcl-2 was downregulated.Conclusion GS could induce apoptosis of chronic myelogenous leukemia K562 cells with activation of Caspase-3 and downregulation of Bcl-2.

关 键 词：K562细胞凋亡 氨基葡萄糖硫酸盐 实验研究 CASPASE-3 CASPASE-3 BCL-2蛋白表达 AO/EB染色法 慢性髓系白血病 Western bcl-2基因 Annexin 流式细胞仪 凋亡诱导作用 生长抑制作用 细胞生长曲线 台盼蓝拒染法 BLOT检测 对数生长期 

分 类 号：R971.1[医药卫生—药品] R977.4[医药卫生—药学]