The chronic-hypoxia-resistant gastric cancer cell line was established,and its biological characteristics were explored and compared with the parental cell line.Gastric cancer cell lines were cultured under the degressive oxygen concentration.Cell doubling time was calculated by cell counting method.Chemo-resistance ability of cells was tested by MTT assay.Irradiation tolerance of cells was evaluated by colony forming method.Cell cycle distribution was tested with flow cytometry.Invasive ability was tested by Transwell method.The expression levels of GLUT-1 and HIF-1α were detected by using Western blot.MNK45/HYP cells successfully survived under the 1% concentration of oxygen and its cell doubling time was 35.01±1.02 h,while that of MNK45 was 27.35±0.83 h（P〈0.01）.The percentage of MNK45/HYP cells in G0/G1 stage was（58.3±6.1）%,and that of MNK45 cells was（42.2±6.0）%（P〈0.05）.Comparing with the parental cells MNK45,drug resistance indexes of 5-Fu,PTX,OXA,Sn38,GEM and VP16 in MNK45/HYP cells were respectively 5.3,1.3,3.6,2.2,4.8 and 4.4.Colony forming ability of MNK45/HYP cells after irradiation was also significantly higher than MNK45 cells.The invasive number of MNK45/HYP cells was 107.7±17.5,while that of MNK45 cells was 59.0±9.9.The expression levels of GLUT-1 and HIF-1α in MNK45/HYP cells were significantly higher than those in MNK45 cells.MNK45/HYP cells hold biological characteristics of hypoxia tumor with good tolerance to chronic hypoxia,and can be used for the research of solid tumor under chronic hypoxia condition.
Objective The aim of the current study was to establish an oxaliplatin-resistant hepatoma cell line(HepG2/OXA) and investigate the potential mechanisms of its drug resistance.Methods The hepatoma cell subline, HepG2/OXA, resistant to oxaliplatin(OXA), was established from a parent cell line HepG2, by stepwise exposure to gradually increasing concentrations of OXA over a half-year period. Chemosenstivity of the cytotoxic drugs, OXA, cisplatin(CDDP), adriamycin(ADM), and 5-fuorouracil(5-FU), was determined in HepG2 and HepG2/OXA cells, by the Cell counting kit-8(CCK8) assay. Cell cycle distribution of HepG2 and HepG2/OXA cells was analyzed by Flow cytometry(FCM). The expression levels of several drug resistance-related proteins, such as P-glycoprotein(P-gp), multidrug resistant protein 1(MRP1), and excision repair-cross complementing 1(ERCC1) protein in the two cell lines were tested by the western blot assay.Results The IC50 of OXA in HepG2/OXA and HepG2 were 136.84 μmol/L and 23.86 μmol/L, respectively. The resistance index(RI) was 5.34. HepG2 was also demonstrated to be cross-resistant to other antitumor agents, such as 5-FU, ADM, and CDDP. The percentage of HepG2/OXA cells in the S phase was significantly decreased compared to HepG2 cells(25.58% ± 2.36% vs 14.37% ± 2.54%, P < 0.05), while the percentage of cells in the G0/G1 and G2/M phases showed no statistical difference(respectively 55.29% ± 4.98% vs 56.73% ± 4.56%, P > 0.05, and 24.63% ± 4.81% vs 28.26% ± 3.82%, P > 0.05). The ERCC1 was found to be over expressed in HepG2/OXA cells, while there was no difference in the expressions of P-gp and MRP1 between the multiple drug resistance(MDR) phenotype cell line and its parental cell line.Conclusion HepG2/OXA showed an MDR ability; the over expression of ERCC1 might be associated with the platinum resistance of the cells, but P-gp and MRP1 are not.
