Up-regulation of  hMLH1 Gene Expression on Cisplatin Sensitivity of Drug-resistant Ovarian Cancer Cells

Abstract: We constructed the eukaryotic expression plasmid pCAN-hMLH1 carrying the coding sequence and investigated its reversing effect on cisplatin-resistant ovarian cancer cells. With the recombinant DNA techniques，the hMLH1 gene in pET28-hMLH1 plasmid was subcloned into pCAN vector. The correctness of recombinant plasmid was evaluated by enzyme analysis and nucleotide sequencing. The SKOV3/DDP cells were transfected with the recombinant plasmid pCAN-hMLH1 and pCAN vectors by lipofectamine 2000. The expressions of hMLH1 were detected by RT-PCR and Western blot. Chemosensitivity of SKOV3/DDP cells to cisplatin was measured by methyl thiazolyl tetrazolium (MTT). Cell apoptosis was detected by Hoechst dyeing. The mRNA and protein levels of hMLH1 of SKOV3/DDP cells transient transfected with pCAN-hMLH1 were increased significantly. The chemosensitivity to cisplatin was enhanced after the hMLH1 gene being transfected into SKOV3/DDP cells. Hoechst dyeing showed that the apopototic rate of hMLH1 transfected cells was increased. The plasmid pCAN-hM- LH1 has been successfully constructed and expressed in SKOV3/DDP cells effectively. hMLH1 gene can increase the chemosensitivity to cisplatin and improve apoptosis.

CSTR: 32200.14.cjcb.2014.02.0011