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[Establishment of human multidrug-resistant hepatocellular carcinoma cell line (HepG2/Adm) and biological
Bao-Jin Zhai1, Feng Wu, Ze-Yong Shao
1The Center for Tumor Therapy, The Second Affiliated Hospital, Chongqing University of Medical Science, Chongqing, 400016, PR China. zhaibaojin@yahoo.com.cn
Background & Objective:
Multidrug resistance (MDR) is considered to be the major obstacle for chemotherapy. In order to reverse tumor MDR in vitro, we designed this study to establish human multidrug-resistant hepatocellular carcinoma cell line (HepG2/Adm) and to investigate its biological characteristics.
Methods:
An adriamycin-resistant human hepatocellular carcinoma cell subline (HepG2/Adm) was established in vitro using gradually increased concentration of adriamycin (ADM) in culture. Cell growth was measured and multidrug resistance to multianticancer agents was evaluated by MTT assay. Flow cytometry (FCM) was performed to determine cell cycle,and to assess the expression of P-glycoprotein (P-gp), multidrug resistance-associated protein (MRP),lung-related protein (LRP),and glutathione S-transferase (GST), respectively. MDR mRNA expression related to these proteins were determined with reverse transcription polymerase chain reaction (RT-PCR).
Results:
Compared to HepG2, HepG2/Adm had a prolonged doubling time of 30.1 hours. The number of cells in S-phase was significantly decreased (5.6+/-0.03)% in HepG2/Adm while those in G1 and G2-phase increased [(4.2+/-0.09)% and (1.5+/-0.08)%, respectively]. Furthermore, HepG2/Adm was resistant to many anti-tumor agents, and its IC(50) of ADM was 26 times higher than that of parent cell line HepG2. Significant overexpression of P-gp, MRP, and GST were detected. RT-PCR showed that mRNA expression of P-gp, MRP, LRP, and GST were significantly increased in HepG2/Adm.
Conclusion:
HepG2/Adm is human multidrug-resistant, and its resistance may be closely related to the overexpression of P-gp, MRP, and GST.
Insights
This study established a human multidrug-resistant liver cancer cell line (HepG2/Adm) to combat chemotherapy resistance. The new cell line overexpresses key drug resistance proteins, offering a model for developing new cancer therapies.
Area of Science:
- Oncology
- Molecular Biology
- Cell Biology
Background:
- Multidrug resistance (MDR) is a significant challenge in chemotherapy effectiveness.
- Developing models to study and overcome tumor MDR is crucial for improving cancer treatment outcomes.
Purpose of the Study:
- To establish a human multidrug-resistant hepatocellular carcinoma cell line (HepG2/Adm) in vitro.
- To investigate the biological characteristics and drug resistance mechanisms of the established cell line.
Main Methods:
- Adriamycin (ADM) was used to gradually induce resistance in HepG2 cells to create the HepG2/Adm subline.
- Cell proliferation was assessed using MTT assays, and drug resistance was evaluated against various anticancer agents.
- Flow cytometry (FCM) analyzed cell cycle distribution and expression of P-glycoprotein (P-gp), multidrug resistance-associated protein (MRP), lung-related protein (LRP), and glutathione S-transferase (GST).
- Reverse transcription polymerase chain reaction (RT-PCR) quantified mRNA expression of resistance-related proteins.
Main Results:
- The HepG2/Adm cell line exhibited a 26-fold higher IC(50) for ADM compared to the parent HepG2 cells, indicating significant resistance.
- HepG2/Adm cells showed altered cell cycle distribution with decreased S-phase and increased G1 and G2 phases.
- Overexpression of P-gp, MRP, and GST proteins and their corresponding mRNAs was detected in HepG2/Adm cells.
Conclusions:
- The established HepG2/Adm cell line is a valuable in vitro model of human multidrug-resistant hepatocellular carcinoma.
- The observed multidrug resistance in HepG2/Adm cells is strongly associated with the overexpression of P-gp, MRP, and GST.
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