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目的观察人脐血间充质干细胞携带人肿瘤坏死因子α(TNF-α)基因对绒癌耐药细胞株耐药指数的影响。方法应用AdMax系统构建携带TNF-α基因的腺病毒载体。鉴定、分离、培养、扩增、纯化人脐血间充质干细胞(hUCB-MSCs)。携带TNF-α基因的重组腺病毒感染hUCB-MSCs后,与绒癌耐药细胞株JEG3/VP16共培养,空载腺病毒(Ad-EGFP)感染的hUBC-MSCs做为对照,MTT法测定共培养48h后JEG-3/VP16对依托泊苷(VP16)、氟尿嘧啶(5-FU)、甲氨蝶呤(MTX)耐药情况。结果(1)酶切、聚合酶链式反应(PCR)鉴定构建的重组腺病毒携带TNF-α基因。(2)重组腺病毒可高效感染hUCB-MSCs。(3)JEG-3/VP16和转染TNFa基因的hUCB-MSCs共培养,JEG-3/VP16对VP16、5-FU、MTX的IC50分别下降至未共培养前IC50的1/12.8,1/6.75和1/10.1。结论携带hTNFα基因的hUCB-MSCs与绒癌耐药细胞株JEG-3/VP16共培养后,逆转了JEG-3/VP16的耐药性。
Objective To observe the effect of human tumor necrosis factor-α (TNF-α) gene in human umbilical cord blood-derived mesenchymal stem cells on the drug resistance index of choriocarcinoma cell line. Methods AdMax system was used to construct adenoviral vector carrying TNF-α gene. Identification, isolation, culture, amplification and purification of human umbilical cord blood mesenchymal stem cells (hUCB-MSCs). After hUCB-MSCs were infected with the recombinant adenovirus carrying TNF-α gene, hUBC-MSCs infected with Ad-EGFP were co-cultured with the choriocarcinoma cell line JEG3 / VP16. MTT assay After 48h, the drug resistance of JEG-3 / VP16 to VP16, 5-fluorouracil and methotrexate (MTX) were studied. Results (1) The recombinant adenovirus constructed by restriction enzyme digestion and polymerase chain reaction (PCR) identified the TNF-α gene. (2) Recombinant adenovirus can efficiently infect hUCB-MSCs. (3) IC50 of JEG-3 / VP16 on VP16, 5-FU and MTX decreased to 1/12.8, 1 / 6.75 and 1 / 10.1. Conclusion The co-culture of hUCB-MSCs with hTNFα gene and the choriocarcinoma cell line JEG-3 / VP16 reversed the drug resistance of JEG-3 / VP16.