Establishment of a cisplatin-induced multidrug resistance cell line SK-Hep1/DDP

Chin J Cancer. 2010 Feb;29(2):167-71. doi: 10.5732/cjc.009.10630.

Abstract

Background and objective: Multidrug resistance (MDR) is a major obstacle in the chemotherapy of cancer patients. The aim of this study was to establish a mutidrug-resistant cell line SK-Hep1/DDP and explore its molecular mechanism of the MDR.

Methods: SK-Hep1/DDP cell line was induced by pulse treatment using a high concentration of cisplatin (DDP) in vitro. The chemoresistance indexes of cells were evaluated by CCK-8 assays. The protein of MDR1 (ABCB1), MRP1 (ABCC1), MRP2 (ABCC2) and Bax were detected by Western blotting, and the effect of MDR1 inhibitor cyclosporine A (CsA) on expression of MDR1 proteins in SK-Hep1 and SK-Hep1/DDP cell lines. Flow cytometry was performed to determine the distribution of the cell cycle and cell apoptosis ratio.

Results: The SK-Hep1/DDP cells were 13.76 times more resistant to DDP in comparison with SK-Hep1 cells, and SK-Hep1/DDP cells also exhibited cross-resistance to many other chemotherapeutic agents (adramycin and 5-fuorouracil). MDR1, MRP1, and MRP2 protein expressions were significantly higher in the SK-Hep1/DDP than in the SK-Hep1 (P < 0. 01), but Bax was lower in the SK-Hep1/DDP than in the SK-Hep1(P < 0. 01). There was no obvious influence between SK-Hep1 and SK-Hep1/DDP cells in the expression of MDR1 by MDR1 inhibtor CsA (P > 0. 05). The percentages of cells in G(2)/M and S phase were significantly increased in SK-Hep1/DDP in comparison with those in SK-Hep1 [(20.67 +/- 5.69)% vs. (12.14 +/- 3.36)%; (42.20 +/- 2.65)% vs. (27.91 +/- 2.16)%; P < 0. 01]. After the cells were exposed to 10 μg/mL DDP for 24 h, the cell apoptosis rate of SK-Hep1/DDP was decreased in comparison with SK-Hep1, but it was increased in those with pretreatment of MDR1 inhibitor CsA as compared with those without pretreatment.

Conclusions: A reliable multi-drug resistant human hepatoma cell line SK-Hep1/DDP is successfully established. The MDR mechanisms of this cell lines are closely related to the over-expression of MDR1 MRP1 and MRP2, lower expression of Bax and the attenuated cell apoptosis induced by chemotherapeutic agents.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • ATP Binding Cassette Transporter, Subfamily B, Member 1 / antagonists & inhibitors
  • ATP Binding Cassette Transporter, Subfamily B, Member 1 / metabolism
  • Antineoplastic Agents / pharmacology
  • Apoptosis / drug effects
  • Cell Cycle / drug effects
  • Cell Line, Tumor*
  • Cell Proliferation / drug effects
  • Cisplatin / pharmacology*
  • Cyclosporine / pharmacology
  • Doxorubicin / pharmacology
  • Drug Resistance, Multiple*
  • Drug Resistance, Neoplasm*
  • Fluorouracil / pharmacology
  • Humans
  • Liver Neoplasms / metabolism
  • Liver Neoplasms / pathology*
  • Multidrug Resistance-Associated Proteins / metabolism
  • Vincristine / pharmacology
  • bcl-2-Associated X Protein / metabolism

Substances

  • ATP Binding Cassette Transporter, Subfamily B, Member 1
  • Antineoplastic Agents
  • Multidrug Resistance-Associated Proteins
  • bcl-2-Associated X Protein
  • multidrug resistance-associated protein 2
  • Vincristine
  • Doxorubicin
  • Cyclosporine
  • Cisplatin
  • Fluorouracil
  • multidrug resistance-associated protein 1