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目的:研究腺病毒载体AdING4对人MCF-7乳腺癌细胞的生长抑制及化疗增敏作用。方法:将搭载有ING-4基因的重组腺病毒载体AdING4感染人MCF-7乳腺癌细胞,用荧光显微镜观察感染后的MCF-7细胞形态学变化;RT-PCR和Western-Blot法检测ING-4基因在MCF-7细胞中的转录和表达;RT-PCR法检测凋亡相关基因在MCF-7细胞中的表达;CCK法测定Ad-ING4感染MCF-7乳腺癌细胞后所发挥的细胞增殖抑制作用。流式细胞技术检测ING-4对MCF-7乳腺癌细胞的促凋亡作用。CCK-8法分别测定病毒感染前后的MCF-7乳腺癌细胞的药物半数抑制浓度IC50,并观察Ad-ING4与化疗药物合用后对MCF-7细胞增殖抑制和化疗增敏现象。结果:MCF-7细胞在转染ING-4基因后,明显出现变圆、脱落、皱缩、聚集等现象;外源性ING-4基因在MCF-7细胞中获得成功表达;外源性ING-4基因作用下MCF-7细胞的增殖受到了明显抑制,凋亡率有所升高,凋亡相关基因Bax的表达水平明显上调,Bcl-2、Survivin的表达水平明显下调。ING-4基因感染MCF-7细胞后,使MCF-7细胞对相关化疗药物的敏感度更高;ING-4基因与化疗药物合用后对MCF-7细胞的增殖抑制作用,较之单用化疗药物更为明显。结论:MCF-7细胞在转染ING4基因后其增殖受到了明显抑制并更易凋亡,该现象可能是通过改变Bax,Bcl-2及Survivin表达水平来实现的,且对化疗药物的敏感性更高。
Objective: To study the growth inhibition and chemosensitivity of adenovirus vector AdING4 on human MCF-7 breast cancer cells. METHODS: The recombinant adenovirus AdING4 harboring ING-4 gene was transfected into MCF-7 breast cancer cells. The morphology of infected MCF-7 cells was observed by fluorescence microscopy. The expression of ING-4 was detected by RT-PCR and Western- 4 gene in MCF-7 cells; RT-PCR was used to detect the expression of apoptosis-related genes in MCF-7 cells; CCK assay was used to determine the cell proliferation induced by Ad-ING4 in MCF-7 breast cancer cells Inhibition. Flow cytometry was used to detect the pro-apoptotic effect of ING-4 on MCF-7 breast cancer cells. CCK-8 method was used to determine the IC50 of MC50 of MCF-7 breast cancer cells before and after virus infection. The inhibitory effect of Ad-ING4 combined with chemotherapeutic agents on MCF-7 cell proliferation and chemosensitization was observed. RESULTS: After transfection with ING-4 gene, MCF-7 cells showed obvious rounding, shedding, shrinkage and aggregation. The exogenous ING-4 gene was successfully expressed in MCF-7 cells. -4 gene significantly inhibited the proliferation of MCF-7 cells, the apoptosis rate was increased, the expression of apoptosis-related gene Bax was significantly increased, Bcl-2, Survivin expression was significantly down-regulated. In MCF-7 cells infected by ING-4 gene, the sensitivity of MCF-7 cells to related chemotherapeutic drugs was higher. The inhibitory effect of ING-4 gene and chemotherapeutic agents on MCF-7 cell proliferation was higher than that of chemotherapy alone Drugs are more obvious. CONCLUSION: The proliferation of MCF-7 cells after ING4 transfection was significantly inhibited and more likely to apoptosis, which may be achieved by changing the expression levels of Bax, Bcl-2 and Survivin, and more sensitive to chemotherapeutics high.