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Transcriptome Analysis of Liver Cancer Cell Huh-7 Treated With Metformin
Chun-Qing Li1, Zhi-Qin Liu2, Sha-Sha Liu1,3
1Key Laboratory of Microbial Diversity Research and Application of Hebei Province, College of Life Science, Hebei University, Baoding, China.
Abstract:
Metformin is a kind of widely used antidiabetic drug that regulates glucose homeostasis by inhibiting liver glucose production and increasing muscle glucose uptake. Recently, some studies showed that metformin exhibits anticancer properties in a variety of cancers. Although several antitumor mechanisms have been proposed for metformin action, its mode of action in human liver cancer remains not elucidated. In our study, we investigated the underlying molecular mechanisms of metformin's antitumor effect on Huh-7 cells of hepatocellular carcinoma (HCC) in vitro. RNA sequencing was performed to explore the effect of metformin on the transcriptome of Huh-7 cells. The results revealed that 4,518 genes (with log2 fold change > 1 or < -1, adjusted p-value < 0.05) were differentially expressed in Huh-7 cells with treatment of 25-mM metformin compared with 0-mM metformin, including 1,812 upregulated and 2,706 downregulated genes. Gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses identified 54 classical pathways that were significantly enriched, and 16 pathways are closely associated with cancer, such as cell cycle, DNA replication, extracellular matrix-receptor interaction, and so on. We selected 11 differentially expressed genes, which are closely associated with HCC, to validate their differential expressions through a quantitative real-time reverse transcription-polymerase chain reaction. The result exhibited that the genes of fatty acid synthase, mini-chromosome maintenance complex components 6 and 5, myristoylated alanine-rich C-kinase substrate, fatty acid desaturase 2, C-X-C motif chemokine ligand 1, bone morphogenetic protein 4, S-phase kinase-associated protein 2, kininogen 1, and proliferating cell nuclear antigen were downregulated, and Dual-specificity phosphatase-1 is significantly upregulated in Huh-7 cells with treatment of 25-mM metformin. These differentially expressed genes and pathways might play a crucial part in the antitumor effect of metformin and might be potential targets of metformin treating HCC. Further investigations are required to evaluate the metformin mechanisms of anticancer action in vivo.
Insights
Metformin, an antidiabetic drug, shows anticancer effects on liver cancer cells. This study identified key gene expression changes and pathways affected by metformin, suggesting potential therapeutic targets for hepatocellular carcinoma (HCC).
Area of Science:
- Oncology
- Pharmacology
- Molecular Biology
Background:
- Metformin is a widely used antidiabetic medication with demonstrated anticancer properties across various cancer types.
- The precise molecular mechanisms underlying metformin's antitumor effects in human liver cancer, specifically hepatocellular carcinoma (HCC), remain largely unelucidated.
- Understanding these mechanisms is crucial for exploring metformin as a potential therapeutic agent for HCC.
Purpose of the Study:
- To investigate the molecular mechanisms of metformin's antitumor effect on hepatocellular carcinoma (HCC) Huh-7 cells *in vitro*.
- To identify differentially expressed genes and enriched pathways in HCC cells upon metformin treatment using RNA sequencing.
- To validate the expression of key genes potentially involved in metformin's anticancer action in HCC.
Main Methods:
- Utilized RNA sequencing to analyze the transcriptome of Huh-7 HCC cells treated with 25-mM metformin compared to untreated controls.
- Performed Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses to identify significantly enriched biological pathways.
- Validated the differential expression of 11 selected HCC-associated genes using quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR).
Main Results:
- Metformin treatment resulted in significant differential expression of 4,518 genes in Huh-7 cells (1,812 upregulated, 2,706 downregulated).
- Enrichment analysis identified 54 significantly enriched pathways, with 16 directly associated with cancer, including cell cycle and DNA replication.
- Quantitative real-time reverse transcription-polymerase chain reaction confirmed downregulation of genes such as fatty acid synthase and upregulation of Dual-specificity phosphatase-1, among others.
Conclusions:
- Metformin significantly alters the transcriptome of HCC cells, impacting numerous cancer-associated pathways.
- Specific differentially expressed genes and pathways identified in this study may mediate metformin's antitumor effects in HCC.
- These findings highlight potential molecular targets for metformin-based therapies in hepatocellular carcinoma, warranting further *in vivo* investigation.
