中国肿瘤生物治疗杂志2017，Vol.24Issue(12)：1375-1380,6.DOI:10.3872/j.issn.1007-385x.2017.12.006
RNA干扰Livin基因表达增强骨肉瘤MG-63耐药细胞对多柔比星的化疗敏感性
Livin gene silence by RNA interference enhances the chemotherapeutic sensitivity of drug resistant MG-63 osteosarcoma cells to doxorubicin
摘要
Abstract
Objective:To investigate the effect of Livin gene (an inhibitor of apoptosis protein) on reversing the drug-resistance of osteosarcoma.Methods:Drug-resistant MG-63 cell strain was established in vitro by continuous exposure to doxorubicin at gradually increased concentrations.The resistance index of drug-resistant MG-63 cells was examined by MTT method;Livin protein expressions in MG-63 cells and durg-resistant MG-63 cells were determined by Western blotting.Livin shRNA eukaryotic expression vector (pSilencer3.l-H1 neo-Livin si) was constructed and then transfected into drug-resistant MG-63 cells by using Lipofectmine 2000.Expression change of Livin mRNA and protein in drug-resistant MG-63 cells before and after the transfection was respectively measured by Real-time PCR and Western blotting.The distribution of cell cycle and apoptosis were determined by flow cytometry.The analysis of chemotherapeutic sensitivity of drug-resistant MG-63 cell to doxorubicin was performed by MTT.Results:The recombinant eukaryotic expression vector pSilencer3.1-H1 neo-Livin si was successfully constructed.Resistance index to doxorubicin of drug-resistant MG-63 cells (MG-63/R) was 81.32±5.33.Livin shRNA could significantly inhibit the mRNA and protein expression of Livin in MG-63/R cells compared with untransfected group and non-specific transfected group (down-regulated by 72％ and 69％ at mRNA and protein level respectively,all P＜0.05).The flow cytometry analysis showed there was significantly higher apoptosis rate in Livin shRNA transfected group than that of untransfected group and non-specific transfected group ([22.4±3.2]％ vs [4.2±1.1]％,[4.7±0.6]％,P＜0.05).After adding doxorubicin,the proliferation of three groups of cells was all inhibited at different levels in time-dependent manner;However,the cell survival rate in Livin shRNA transfected group was significantly lower than that in other two groups (P＜0.05).Conclusion:Livin shRNA could efficiently promote the apoptosis of drug-resistant MG-63 cells,and thus increase its sensitivity to chemotherapy drugs.关键词
骨肉瘤/Livin基因/RNA干扰/多柔比星/耐药Key words
osteosarcoma/Livin gene/RNA interference/doxorubicin/drug resistance分类
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孙廓,袁初平,李怀强,廖琦..RNA干扰Livin基因表达增强骨肉瘤MG-63耐药细胞对多柔比星的化疗敏感性[J].中国肿瘤生物治疗杂志,2017,24(12):1375-1380,6.基金项目
国家自然科学基金资助项目(No.81660443) （No.81660443）
江西省卫计委科技项目资助(No.20155278).Project supported by the National Natural Science Foundation of China(No.81660443),and the Health and Family Planning Commision of Jiangxi Province(No.20155278) （No.20155278）
