hMSH2重组质粒对人卵巢癌细胞顺铂敏感性的影响

Effects of hMSH2 gene on cisplatin sensitivity of drug-resistant ovarian cancer cells

目的:研究h MSH2重组质粒对卵巢癌顺铂敏感性方面的影响。方法:构建重组真核表达质粒pCAN-h MSH2,通过酶切及测序法对重组质粒进行鉴定。采用脂质体转染技术对卵巢癌耐药细胞SKOV3/DDP进行瞬时转染,对照组为未转染细胞和SKOV3敏感细胞;h MSH2在细胞内的表达变化由Western blotting和RTPCR进行检测;四甲基偶氮唑蓝(M TT)法对细胞的顺铂敏感性进行检测;耐药细胞的凋亡情况通过H oechst染色法检测。结果:重组质粒p CAN-h MSH2转染进SKOV3/DDP后,经RT-PCR和Western blotting检测证实转染成功,h MSH2的表达得到增强;MTT结果提示转染后的卵巢癌耐药细胞对顺铂的敏感性明显增强;Hoechst染色发现,转染后耐药细胞的凋亡明显增强。结论 :h MSH2基因转染后能提高其在SKOV3/DDP细胞中的表达,增强SKOV3/DDP细胞对顺铂的敏感性,促进在顺铂作用下的凋亡。

Objective： To investigate the effect of h MSH2 on cisplatin sensitivity of drug- resistant ovarian cancer cells. Methods： The eukaryotic expresion plasmid p CAN- h MSH2 carrying the coding sequence was constructed and evaluated by enzyme analysis and nucleotide sequencing. The SKOV3 / DDP cells were transfected with the recombinant plasmid p CAN- h MSH2 and p CAN vector by using Lipofectamine 2000. The expressions of h MSH2 gene were detected by RT- PCR and Western blotting. Chemosensitivity of SKOV3 / DDP cells to cisplatin was evaluated by methyl thiazolyl tetrazolium（ MTT）. The apoptosis of cells was detected by Hoechst dyeing after treatment with cisplatin. Results： The expression level of hMSH2 gene of SKOV3 / DDP cells was increased significantly after transient transfection with p CAN- hMSH2. MTT assay showed the chemosensitivity to cisplatin in SKOV3 / DDP cells transfected with hMSH2 was enhanced. Hoechst dyeing showed that the apopototic rate of h MSH2 transfected cells was increased. Conclusion： hMSH2 transfection can increase the expression level of hMSH2 gene of SKOV3 / DDP cells and enhance the chemosensitivity to cisplatin and improve apoptosis.