中华肿瘤杂志
中華腫瘤雜誌
중화종류잡지
CHINESE JOURNAL OF ONCOLOGY
2009年
8期
582-586
,共5页
孙萍%张良明%孙等军%董亮亮
孫萍%張良明%孫等軍%董亮亮
손평%장량명%손등군%동량량
沙利度胺%SMMC-7721细胞%凋亡%血管内皮生长因子
沙利度胺%SMMC-7721細胞%凋亡%血管內皮生長因子
사리도알%SMMC-7721세포%조망%혈관내피생장인자
Thalidomide%SMMC-7721 cells%Apoptosis%Vascular endothelial growth factor
目的 研究沙利度胺对人肝癌细胞株SMMC-7721体外生长的抑制作用及其可能的机制.方法 将不同浓度的沙利度胺作用于人肝癌细胞株SMMC-7721,采用四甲摹偶氮唑蓝(MTT)法检测沙利度胺对SMMC-7721细胞的增殖抑制作用.将SMMC-7721细胞培养至对数生长期,采用DNA琼脂糖凝胶电泳、荧光显微镜观察、流式细胞仪检测等方法 观察沙利度胺处理后SMMC-7721细胞的凋亡梯度、形态学变化和凋亡率,并对凋亡调控蛋白caspase-3的表达进行测定.采用酶联免疫吸附(ELISA)法测定不同浓度的沙利度胺处理后SMMC-7721细胞表达血管内皮生长因子(VEGF)的变化.结果 沙利度胺的浓度从3.125μg/ml增至200μg/ml时,其对SMMC-7721细胞的增殖抑制率从11.7%增至34.2%;当沙利度胺的浓度>25 μg/ml时,其对SMMC-7721细胞的增殖抑制作用明显强于空白对照组(P<0.05).200 μg/ml的沙利度胺处理SMMC-7721细胞24 h后,行琼脂糖凝胶电泳,可见到DNA梯形条带;48 h后梯形条带更明显,并且在荧光显微镜下可见SMMC-7721细胞出现核固缩和核裂解现象.200μg/ml的沙利度胺处理SMMC-7721细胞12、24、48和72 h时,碘化丙啶(PI)法检测SMMC-7721细胞的凋亡率分别为3.1%±0.5%、8.4%±1.3%、19.4%±3.5%和25.8%±2.1%,24 h起的凋亡率均明显高于空白对照组SMMC-7721细胞48 h的自然凋亡率(1.6%±0.6%,均P<0.05).50、100和200μg/ml的沙利度胺处理SMMC-7721细胞48 h时,Annexin V-FITC/PI双标法检测SMMC-7721细胞的凋亡率分别为8.7%±1.2%、16.8%±2.5%和25.4%±4.5%,均明显高于空白对照组SMMC-7721细胞48 h的自然凋亡率(2.1%±0.5%,均P<0.05).随着沙利度胺浓度的增加,表达caspase-3蛋白的SMMC-7721细胞数量不断增加,而SMMC-7721细胞中VEGF的含量却逐渐下降.结论 沙利度胺可能通过诱导肝癌细胞的凋亡、抑制肿瘤血管的生成而发挥双重抗肿瘤生长的作用.
目的 研究沙利度胺對人肝癌細胞株SMMC-7721體外生長的抑製作用及其可能的機製.方法 將不同濃度的沙利度胺作用于人肝癌細胞株SMMC-7721,採用四甲摹偶氮唑藍(MTT)法檢測沙利度胺對SMMC-7721細胞的增殖抑製作用.將SMMC-7721細胞培養至對數生長期,採用DNA瓊脂糖凝膠電泳、熒光顯微鏡觀察、流式細胞儀檢測等方法 觀察沙利度胺處理後SMMC-7721細胞的凋亡梯度、形態學變化和凋亡率,併對凋亡調控蛋白caspase-3的錶達進行測定.採用酶聯免疫吸附(ELISA)法測定不同濃度的沙利度胺處理後SMMC-7721細胞錶達血管內皮生長因子(VEGF)的變化.結果 沙利度胺的濃度從3.125μg/ml增至200μg/ml時,其對SMMC-7721細胞的增殖抑製率從11.7%增至34.2%;噹沙利度胺的濃度>25 μg/ml時,其對SMMC-7721細胞的增殖抑製作用明顯彊于空白對照組(P<0.05).200 μg/ml的沙利度胺處理SMMC-7721細胞24 h後,行瓊脂糖凝膠電泳,可見到DNA梯形條帶;48 h後梯形條帶更明顯,併且在熒光顯微鏡下可見SMMC-7721細胞齣現覈固縮和覈裂解現象.200μg/ml的沙利度胺處理SMMC-7721細胞12、24、48和72 h時,碘化丙啶(PI)法檢測SMMC-7721細胞的凋亡率分彆為3.1%±0.5%、8.4%±1.3%、19.4%±3.5%和25.8%±2.1%,24 h起的凋亡率均明顯高于空白對照組SMMC-7721細胞48 h的自然凋亡率(1.6%±0.6%,均P<0.05).50、100和200μg/ml的沙利度胺處理SMMC-7721細胞48 h時,Annexin V-FITC/PI雙標法檢測SMMC-7721細胞的凋亡率分彆為8.7%±1.2%、16.8%±2.5%和25.4%±4.5%,均明顯高于空白對照組SMMC-7721細胞48 h的自然凋亡率(2.1%±0.5%,均P<0.05).隨著沙利度胺濃度的增加,錶達caspase-3蛋白的SMMC-7721細胞數量不斷增加,而SMMC-7721細胞中VEGF的含量卻逐漸下降.結論 沙利度胺可能通過誘導肝癌細胞的凋亡、抑製腫瘤血管的生成而髮揮雙重抗腫瘤生長的作用.
목적 연구사리도알대인간암세포주SMMC-7721체외생장적억제작용급기가능적궤제.방법 장불동농도적사리도알작용우인간암세포주SMMC-7721,채용사갑모우담서람(MTT)법검측사리도알대SMMC-7721세포적증식억제작용.장SMMC-7721세포배양지대수생장기,채용DNA경지당응효전영、형광현미경관찰、류식세포의검측등방법 관찰사리도알처리후SMMC-7721세포적조망제도、형태학변화화조망솔,병대조망조공단백caspase-3적표체진행측정.채용매련면역흡부(ELISA)법측정불동농도적사리도알처리후SMMC-7721세포표체혈관내피생장인자(VEGF)적변화.결과 사리도알적농도종3.125μg/ml증지200μg/ml시,기대SMMC-7721세포적증식억제솔종11.7%증지34.2%;당사리도알적농도>25 μg/ml시,기대SMMC-7721세포적증식억제작용명현강우공백대조조(P<0.05).200 μg/ml적사리도알처리SMMC-7721세포24 h후,행경지당응효전영,가견도DNA제형조대;48 h후제형조대경명현,병차재형광현미경하가견SMMC-7721세포출현핵고축화핵렬해현상.200μg/ml적사리도알처리SMMC-7721세포12、24、48화72 h시,전화병정(PI)법검측SMMC-7721세포적조망솔분별위3.1%±0.5%、8.4%±1.3%、19.4%±3.5%화25.8%±2.1%,24 h기적조망솔균명현고우공백대조조SMMC-7721세포48 h적자연조망솔(1.6%±0.6%,균P<0.05).50、100화200μg/ml적사리도알처리SMMC-7721세포48 h시,Annexin V-FITC/PI쌍표법검측SMMC-7721세포적조망솔분별위8.7%±1.2%、16.8%±2.5%화25.4%±4.5%,균명현고우공백대조조SMMC-7721세포48 h적자연조망솔(2.1%±0.5%,균P<0.05).수착사리도알농도적증가,표체caspase-3단백적SMMC-7721세포수량불단증가,이SMMC-7721세포중VEGF적함량각축점하강.결론 사리도알가능통과유도간암세포적조망、억제종류혈관적생성이발휘쌍중항종류생장적작용.
Objective The aim of this study was to investigate the effect of thalidomide on the growth of human hepatoma cell line SMMC-7721 cells in vitro, and to explore the curative possibility of hepatocellular carcinoma with thalidomide. Methods SMMC-7721 cells were treated with Thalidomide at different concentrations. The cell growth and proliferation was assessed by MTT assay. DNA ladder, apoptosis rate and changes of cell nuclei were studied by agarose electrophresis, fluorescence microscopy and flow cytometry, respectively. The expression of caspase-3 was analyzed with flow cytometry. The VEGF content of SMMC-7721 cells in culture medium was tested by ELSIA. Results When the concentration of Thalidomide .solution was increased from 3. 125 μg/ml to 200 μg/ml, the cell growth was inhibited by from 11.7% to 34.2%. Compared with the control group, the thalidomide solution at a concentration of 25,50, I00 and 200 μg/ml solution significantly inhibited the proliferation of SMMC-7721 cells ( P < 0. 05 ). A ladder pattern of DNA fragments appeared after SMMC-7721 cells exposed to 200 μg/ml thalidomide for 24 h, especially for 48 h. Fluorescence microscopy revealed that the cell nuclei were condensed and fragmented after the cells were exposed to 200 μg/mi thalidomide for 48 h. In cells treated with 200 μg/ml thalidomide for 12,24,48 and 72 h, the apoptotie rate was 3.1% ± 0. 5%,8.4% ± 1.3%, 19.4% ± 3.5% and 25.8% ± 2. 1%, respectively, significantly higher than that in the negative control group 1.6% ± 0.6%. The cells treated with thalidomide at a concentration of 50, 100, 200 μg/ml for 48 h, the apoptotie rate was 8.7% ± 1.2%, 16. 8% ± 2. 5% and 25.4% ± 4.5%, respectively, increasing in a dose-dependent manner, also significantly than that in the cells of control group 2.1% ± 0.5%, (all were P < 0.05 ). The easpase-3 positivity of SMMC-7721 cells treated with thalidomide was increasing along with the increase of treatment time or drug concentration, but not in the control cells. The VEGF content in SMMC-7721 cells was lowering when thalidomide was used in an increasing concentration. Conclusion Under the conditions used in this study, thalidomide can inhibit the proliferation of SMMC-7721 cells in vitro. Induction of apoptosis and inhibition of angiogenesis may be possibly two mechanisms for its anticancer action.
