5-氮杂-2’-脱氧胞苷诱导乳腺癌细胞自噬与DNA损伤相关联  

Autophagy of breast cancer cells induced by 5-aza-2’-deoxycytidine is associated with DNA damage

作　　者：梁文颖[1] 熊静波[1] 赵嘉佳[1] 

机构地区：[1]南方医科大学基础医学院细胞生物教研室,广州510515

出　　处：《肿瘤》2012年第7期495-500,共6页Tumor

基　　金：国家自然科学基金资助项目(编号:81071810;81172058)

摘　　要：目的:探讨5-氮杂-2’-脱氧胞苷对乳腺癌细胞自噬的影响及其可能的机制。方法:用依托泊苷、顺铂和5-氮杂-2’-脱氧胞苷处理乳腺癌细胞后,采用彗星实验测定细胞DNA损伤,蛋白质印迹法检测p53和p21蛋白表达;细胞自噬测定用3种方法:(1)蛋白质印迹法检测微管相关蛋白1轻链3-Ⅱ(microtubule-associated protein 1 light chain3-Ⅱ,LC3-Ⅱ)蛋白的表达;(2)用单丹磺酰戊二胺对乳腺癌细胞染色后,在荧光显微镜下计数自噬泡阳性细胞,并计算自噬泡阳性细胞百分比;(3)将pEGFP-LC3质粒转染至乳腺癌细胞后,在荧光显微镜下计数含绿色荧光蛋白的阳性细胞,并计算绿色荧光蛋白阳性细胞的百分比。结果:依托泊苷和顺铂均可诱导乳腺癌细胞DNA损伤和自噬,与对照组比较,药物处理组细胞的彗星长度增长(P<0.01),p53、p21和LC3-Ⅱ蛋白表达上调。5-氮杂-2’-脱氧胞苷也可诱导乳腺癌细胞DNA损伤和自噬,与对照组比较,处理组细胞的彗星长度增长(P<0.01),p53、p21和LC3-Ⅱ蛋白表达上调,自噬泡阳性细胞百分比和绿色荧光蛋白阳性细胞百分比上升,差异均有统计学意义(P<0.05和P<0.01)。结论:5-氮杂-2’-脱氧胞苷可诱导乳腺癌细胞自噬,其机制可能与其所诱导的DNA损伤有关。Objective： To investigate the effect of 5-aza-2＇-deoxycytidine on the autophagy of human breast cancer cells, and to explore the possible mechanism. Methods： Breast cancer cells were treated with etoposide, cisplatin and 5-aza-2＇-deoxycytidine. DNA damage was detected by comet assay, and the expressions of p53 and p21 proteins were examined by Western blotting. The autophagy of breast cancer cells was monitored by three methods： （1） The expression of microtubule-associated protein 1 light chain 3-H （LC3-Ⅱ）was detected by Western blotting; （2）The breast cancer cells presenting autophagosome vacuoles were counted under a fluorescence microscope after staining with monodansylcadaverine （MDC）, and the percentage of cells presenting autophagosome vacuoles was calculated; （3） The green fluorescent protein （GFP）-positive breast cancer cells after transfection with pEGFP-LC3 were counted under a fluorescence microscope, and the percentage of GFP-positive breast cancer cells was calculated. Results： Etoposide and cisplatin induced the DNA damage and autophagy in breast cancer cells. Compared with the untreated breast cancer cells, the comet tail length was increased （P〈O.01） and the expression levels of p53, p21 and LC3-H proteins were all up-regulated in the breast cancer cells treated with etoposide and cisplatin. The 5-aza-2＇-deoxycytidine could also induce the DNA damage and autophagy in breast cancer cells, and the comet tail length was increased （P〈O.01）, the expression levels of p53, p21 and LC3-Ⅱ proteins were up-regulated, as well as the percentages of MDC-positive breast cancer cells and GFP-positive breast cancer cells were both increased （P〈0.05, P〈0.01）. Conclusion： 5-Aza-2＇- deoxycytidine can induce the autophagy of breast cancer cells, and the mechanism may be associated with DNA damage.

关 键 词：乳腺肿瘤 自噬 DNA损伤 5-氮杂-2’-脱氧胞苷 

分 类 号：R737.9[医药卫生—肿瘤]