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HNF1? inhibition triggers epithelial-mesenchymal transition in human liver cancer cell lines.

ABSTRACT: BACKGROUND: Hepatocyte Nuclear Factor 1? (HNF1?) is an atypical homeodomain-containing transcription factor that transactivates liver-specific genes including albumin, ?-1-antitrypsin and ?- and ?-fibrinogen. Biallelic inactivating mutations of HNF1A have been frequently identified in hepatocellular adenomas (HCA), rare benign liver tumors usually developed in women under oral contraceptives, and in rare cases of hepatocellular carcinomas developed in non-cirrhotic liver. HNF1?-mutated HCA (H-HCA) are characterized by a marked steatosis and show activation of glycolysis, lipogenesis, translational machinery and mTOR pathway. We studied the consequences of HNF1? silencing in hepatic cell lines, HepG2 and Hep3B and we reproduced most of the deregulations identified in H-HCA. METHODS: We transfected hepatoma cell lines HepG2 and Hep3B with siRNA targeting HNF1? and obtained a strong inhibition of HNF1? expression. We then looked at the phenotypic changes by microscopy and studied changes in gene expression using qRT-PCR and Western Blot. RESULTS: Hepatocytes transfected with HNF1? siRNA underwent severe phenotypic changes with loss of cell-cell contacts and development of migration structures. In HNF1?-inhibited cells, hepatocyte and epithelial markers were diminished and mesenchymal markers were over-expressed. This epithelial-mesenchymal transition (EMT) was related to the up regulation of several EMT transcription factors, in particular SNAIL and SLUG. We also found an overexpression of TGF?1, an EMT initiator, in both cells transfected with HNF1? siRNA and H-HCA. Moreover, TGF?1 expression is strongly correlated to HNF1? expression in cell models, suggesting regulation of TGF?1 expression by HNF1?. CONCLUSION: Our results suggest that HNF1? is not only important for hepatocyte differentiation, but has also a role in the maintenance of epithelial phenotype in hepatocytes.

SUBMITTER: Pelletier L

PROVIDER: S-EPMC3203860 | biostudies-other | 2011

REPOSITORIES: biostudies-other

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HNF1α inhibition triggers epithelial-mesenchymal transition in human liver cancer cell lines.

Pelletier Laura L Rebouissou Sandra S Vignjevic Danijela D Bioulac-Sage Paulette P Zucman-Rossi Jessica J

BMC cancer 20111005

<h4>Background</h4>Hepatocyte Nuclear Factor 1α (HNF1α) is an atypical homeodomain-containing transcription factor that transactivates liver-specific genes including albumin, α-1-antitrypsin and α- and β-fibrinogen. Biallelic inactivating mutations of HNF1A have been frequently identified in hepatocellular adenomas (HCA), rare benign liver tumors usually developed in women under oral contraceptives, and in rare cases of hepatocellular carcinomas developed in non-cirrhotic liver. HNF1α-mutated HC ...[more]

PMID: 21975049

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Project description:Mitochondrial dysfunction promotes cancer aggressiveness, metastasis, and resistance to therapy. Similar traits are associated with epithelial mesenchymal transition (EMT). We questioned whether mitochondrial dysfunction induces EMT in head and neck cancer (HNC) cell lines. We induced mitochondrial dysfunction in four HNC cell lines with carbonyl cyanide-4(trifluoromethoxy)phenylhydrazone (FCCP), a mitochondrial electron transport chain uncoupling agent, and oligomycin, a mitochondrial ATP synthase inhibitor. Extracellular flux analyses and expression of the cystine/glutamate antiporter system xc (xCT) served to confirm mitochondrial dysfunction. Expression of the EMT-related transcription factor SNAI2, the mesenchymal marker vimentin and vimentin/cytokeratin double positivity served to detect EMT. In addition, holotomographic microscopy was used to search for morphological features of EMT. Extracellular flux analysis and xCT expression confirmed that FCCP/oligomycin induced mitochondrial dysfunction in all cell lines. Across the four cell lines, mitochondrial dysfunction resulted in an increase in relative SNAI2 expression from 8.5 ± 0.8 to 12.0 ± 1.1 (mean ± SEM; p = 0.007). This effect was predominantly caused by the CAL 27 cell line (increase from 2.2 ± 0.4 to 5.5 ± 1.0; p < 0.001). Similarly, only in CAL 27 cells vimentin expression increased from 2.2 ± 0.5 × 10-3 to 33.2 ± 10.2 × 10-3 (p = 0.002) and vimentin/cytokeratin double positive cells increased from 34.7 ± 5.1 to 67.5 ± 9.8% (p = 0.003), while the other 3 cell lines did not respond with EMT (all p > 0.1). Across all cell lines, FCCP/oligomycin had no effect on EMT characteristics in holotomographic microscopy. Mitochondrial dysfunction induced EMT in 1 of 4 HNC cell lines. Given the heterogeneity of HNC, mitochondrial dysfunction may be sporadically induced by EMT, but EMT does not explain the tumor promoting effects of mitochondrial dysfunction in general.

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HNF1? inhibition triggers epithelial-mesenchymal transition in human liver cancer cell lines.

Publications

HNF1α inhibition triggers epithelial-mesenchymal transition in human liver cancer cell lines.

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