百里醌抑制乳腺癌血管生长的实验研究  

Anti-angiogenic effect of thymoquinone on breast cancer

作　　者：吴志豪[1] 李香利[1] 郑敏[1] 

机构地区：[1]温州医学院附属第二医院乳腺外科,浙江温州325000

出　　处：《中国病理生理杂志》2016年第1期106-111,共6页Chinese Journal of Pathophysiology

基　　金：温州市科技计划(No.Y20130278)

摘　　要：目的：探讨百里醌对乳腺癌血管生长的作用于及其机制。方法：不同浓度百里醌作用于人脐静脉内皮细胞（HUVECs）及人乳腺癌细胞株MCF-7后,应用CCK-8法检测细胞活力,流式细胞术检测细胞凋亡,小管形成实验检测内皮细胞体外小管形成能力,Western blot法检测内皮细胞中cleaved caspase-3、p-ERK及p-AKT的蛋白水平;建立裸鼠乳腺癌移植瘤模型,完全随机法分成对照组和百里醌组,免疫组织化学法检测乳腺肿瘤组织中CD31的表达,TUNEL法检测移植瘤细胞凋亡。结果：百里醌（20~80 nmol/L）对MCF-7细胞生长无明显抑制作用,20 nmol/L、40 nmol/L和80 nmol/L百里醌作用于内皮细胞24 h后,细胞存活率分别为（74.3±5.9）%、（68.7±5.6）%和（47.9±4.3）%;20 nmol/L、40 nmol/L和80 nmol/L百里醌作用于MCF-7细胞24 h后,细胞凋亡率分别为（2.6±0.3）%、（2.4±0.3）%和（4.6±0.4）%,而相应的内皮细胞凋亡率分别为（21.5±3.7）%、（23.8±2.9）%和（27.8±3.1）%,内皮细胞较乳腺癌细胞对百里醌反应更敏感（P〈0.05）;20 nmol/L、40 nmol/L和80nmol/L百里醌作用于内皮细胞1 h后,内皮细胞小管形成长度分别为（0.88±0.12）mm、（0.76±0.10）mm和（0.54±0.08）mm,与对照组相比,差异有统计学意义（P〈0.05）;80 nmol/L百里醌作用于内皮细胞24 h后,内皮细胞中cleaved caspase-3的水平上调,AKT和ERK磷酸化被抑制,与对照组比较差异有统计学意义（P〈0.05）;免疫组化检测移植瘤组织中CD31结果显示,百里醌组IA值明显少于对照组（P〈0.05）;TUNEL检测结果显示,百里醌组IA值明显增大（P〈0.05）。结论：百里醌可抑制体内外乳腺癌血管生长,降低p-ERK及p-AKT的水平可能是其作用机制之一。AIM： To investigate the anti-angiogenic effect of thymoquinone on human breast cancer and the possible mechanism. METHODS： The activities of human umbilical vein endothelial cells（ HUVECs） and human breast cancer MCF-7 cells were assessed by CCK-8 assay. The apoptotic rate was measured by flow cytometry. Furthermore,the assay of capillary tube formation was used to observe the effect of thymoquinone on the tube formation of HUVECs. The protein levels of cleaved caspase-3,p-ERK and p-AKT were detected by Western blot. MCF-7 cells were subcutaneously injected into nude mice to establish breast xenograft tumors. After 3 weeks of implantation,the mice were randomized into control group and thymoquinone group. After the mice were sacrificed,immunohistochemistry was used to detect the expression of CD31 in the tumors,and the TUNEL test kit was used to explore cell apoptosis in the tumor tissues. RESULTS： Thymoquinone at concentrations of 20 ~ 80 nmol / L exerted no growth inhibitiory effect on MCF-7 cells. However,the cell viability of HUVECs were（ 66. 1 ± 8. 3） %,（ 53. 7 ± 3. 4） % and（ 41. 6 ± 4. 9） % when the concentrations of thymoquinone were 20,40 and 80 nmol / L,respectively. The apoptotic ratio of MCF-7 cells were（ 2. 6 ± 0. 3） %,（ 2. 4 ±0. 3） % and（ 4. 6 ± 0. 4） % and the apoptotic ratio of HUVECs were（ 21. 5 ± 3. 7） %,（ 23. 8 ± 2. 9） % and（ 27. 8 ±3. 1） % when the concentrations of thymoquinone were 20,40 and 80 nmol / L,respectively. HUVECs were more sensitive to thymoquinone-induced apoptosis and inhibition in the cell activity than MCF-7 cells. Incubation of HUVECs with different concentrations of thymoquinone（ 20,40 and 80 nmol / L） for 1 h decreased their tube formation capacity（ P 0. 05）.The protein level of cleaved caspase-3 was up-regulated,but the phosphorylation of AKT and ERK were inhibited in a concentration-dependent manner. The immunohistochemical analysis of CD31 showed significant difference of the integral absorbance between contro

关 键 词：乳腺肿瘤 百里醌 内皮细胞 肿瘤血管生成 

分 类 号：R730.23[医药卫生—肿瘤]