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Differential miRNA expression profiles in hepatocellular carcinoma cells and drug-resistant sublines
Lijuan Zhuo1, Jingfeng Liu, Bin Wang
1Department of Pathology, School of Basic Medical Sciences, Fujian Medical University, Fuzhou 350004, PR China.
Abstract:
Altered expression of miRNAs may contribute to multidrug resistance (MDR) in human cancers. This study investigated the association between miRNAs and MDR in five different drug-resistant hepatocellular carcinoma (HCC) cell sublines. The HCC Huh-7 cell line was treated with adramycin (ADM), cisplatin (DDP), carboplatin (CBP), mitomycin C (MMC) or vincristine (VCR) at increasing concentrations to develop drug-resistant sublines. The cell viability MTT assay was used to detect drug resistance. Five different drug-resistant HCC sublines, Huh-7/ADM, Huh-7/CBP, Huh-7/DDP, Huh-7/MMC and Huh-7/VCR, were established. Cells that were resistant to one drug were also found to be resistant to the other drugs. miRNA microarrays were analyzed to identify differential miRNA expression profiles in these cell lines, and real-time PCR was used to validate miRNA microarray data. miRNA microarray analysis showed that there were 53 upregulated miRNAs in Huh-7/ADM, 56 in Huh-7/CBP, 58 in Huh-7/DDP, 58 in Huh-7/MMC and 49 in Huh-7/VCR, whereas there were 52 downregulated miRNAs in Huh-7/ADM, 50 in Huh-7/CBP, 41 in Huh-7/DDP, 55 in Huh-7/MMC and 56 in Huh-7/VCR. Moreover, 26 simultaneously upregulated and 25 simultaneously downregulated miRNAs were noted in the Huh-7/ADM, Huh-7/CBP, Huh-7/DDP and Huh-7/MMC sublines compared to the parental Huh-7 cell line. In contrast, among these 51 upregulated and downregulated miRNAs, 12 miRNAs were upregulated and 13 miRNAs were downregulated in Huh-7/VCR. Upregulation of miR-27b, miR-181a, miR-146b-5p, miR-181d and miR-146a expression was verified using real-time RT-PCR in the parental and the five drug-resistant cell lines. In conclusion, the present study demonstrates that the differentially expressed miRNA profiles in these five drug-resistant HCC sublines could be useful to further investigate the association of miRNA expression with drug resistance in HCC.
Insights
Altered microRNA (miRNA) expression is linked to multidrug resistance (MDR) in hepatocellular carcinoma (HCC). This study identified distinct miRNA profiles in drug-resistant HCC cell lines, suggesting their role in cancer drug resistance.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- MicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression.
- Altered miRNA expression is implicated in the development of multidrug resistance (MDR) in various human cancers.
- Hepatocellular carcinoma (HCC) remains a significant health challenge, with MDR contributing to treatment failure.
Purpose of the Study:
- To investigate the association between miRNA expression profiles and MDR in hepatocellular carcinoma (HCC).
- To establish and characterize drug-resistant HCC cell sublines for studying miRNA involvement in MDR.
Main Methods:
- Development of five distinct drug-resistant HCC cell sublines (Huh-7/ADM, Huh-7/CBP, Huh-7/DDP, Huh-7/MMC, Huh-7/VCR) through stepwise drug exposure.
- Assessment of drug resistance using the MTT cell viability assay.
- Identification of differentially expressed miRNAs using miRNA microarrays.
- Validation of miRNA expression data through real-time quantitative reverse transcription PCR (RT-PCR).
Main Results:
- Established five drug-resistant HCC sublines exhibiting cross-resistance to multiple chemotherapeutic agents.
- miRNA microarray analysis revealed significant alterations in miRNA expression profiles across the resistant sublines compared to the parental cell line.
- A substantial number of miRNAs were found to be simultaneously upregulated or downregulated across multiple resistant sublines.
- Real-time RT-PCR confirmed the differential expression of specific miRNAs, including miR-27b, miR-181a, miR-146b-5p, miR-181d, and miR-146a.
Conclusions:
- The study successfully generated drug-resistant HCC cell models.
- Differentially expressed miRNA profiles were identified in these resistant HCC sublines.
- These findings highlight the potential role of specific miRNAs in the development and progression of multidrug resistance in HCC, providing targets for future research.
