Development and characterization of multidrug resistant human hepatocarcinoma cell line in nude mice

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AIM: To establish a multidrug resistant (MDR) cell subline from the human hepatocarcinoma cell line (HepG2)in nude mice.METHODS: HepG2 cell cultures were incubated with increasing concentrations of adriamycin (ADM) to develop an ADM-resistant cell subline (HepG2/ADM) with crossresistance to other chemotherapeutic agents. Twenty male athymic BALB/c-nu/nu mice were randomized into HepG2/nude and HepG2/ADM/nude groups (10 in each group). A cell suspension (either HepG2 or HepG2/ADM)was injected subcutaneously into mice in each group.Tumor growth was recorded, and animals were sacrificed 4-5 wk after cell implantation. Tumors were prepared for histology, and viable tumor was dispersed into a single-cell suspension. The IC50 values for a number of chemotherapeutic agents were determined by 2, 3-bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide inner salt (MTT) assay. Rhodamine-123retention/efflux and the level of resistance-associated proteins were determined by flow cytometry. The mRNA expression of mdr1, mrp and Irp genes was detected using reverse transcriptase polymerase chain reaction (RT-PCR) in HepG2/nude and HepG2/ADM/nude groups.RESULTS: The appearances of HepG2/nude cells were slightly different from those of HepG2/ADM/nude cells.Similar tumor growth curves were determined in both groups. A cross-resistance to ADM, vincristine, cisplatin and 5-fiuorouracil was seen in HepG2/ADM/nude group.The levels of P-glycoprotein and multidrug resistanceassociated proteins were significantly increased. The mRNA expression levels of mdr1, mrp and Irp were higher in HepG2/ADM/nude cells.CONCLUSION: ADM-resistant HepG2 subline in nude mice has a cross resistance to chemotherapeutic drugs.Tt may be used as an in vivo model to investigate the mechanisms of MDR, and explore the targeted approaches to overcoming MDR.
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