C/EBPalpha is up-regulated in a subset of hepatocellular carcinomas and plays a role in cell growth and proliferation.

Abstract

BACKGROUND & AIMS:

C/EBPalpha (cebpa) is a putative tumor suppressor. However, initial results indicated that cebpa was up-regulated in a subset of human hepatocellular carcinomas (HCCs). The regulation and function of C/EBPalpha was investigated in HCC cell lines to clarify its role in liver carcinogenesis.

METHODS:

The regulation of C/EBPalpha expression was studied by quantitative reverse transcription-polymerase chain reaction (qRT-PCR), Western blotting, immunohistochemistry, methylation-specific PCR, and chromatin immunoprecipitation assays. C/EBPalpha expression was knocked-down by small interfering RNA or short hairpin RNA. Functional assays included colony formation, methylthiotetrazole, bromodeoxyuridine incorporation, and luciferase-reporter assays.

RESULTS:

Cebpa was up-regulated at least 2-fold in a subset (approximately 55%) of human HCCs compared with adjacent nontumor tissues. None of the up-regulated samples were positive for hepatitis C infection. The HCC cell lines Hep3B and Huh7 expressed high, PLC/PRF/5 intermediate, HepG2 and HCC-M low levels of C/EBPalpha, recapitulating the pattern of expression observed in HCCs. No mutations were detected in the CEBPA gene in HCCs and cell lines. C/EBPalpha was localized to the nucleus and functional in Hep3B and Huh7 cells; knocking-down its expression reduced target-gene expression, colony formation, and cell growth, associated with a decrease in cyclin A and CDK4 concentrations and E2F transcriptional activity. Epigenetic mechanisms including DNA methylation, and the binding of acetylated histone H3 to the CEBPA promoter-regulated cebpa expression in the HCC cells.

CONCLUSIONS:

C/EBPalpha is up-regulated in a subset of HCCs and has growth-promoting activities in HCC cells. Novel oncogenic mechanisms involving C/EBPalpha may be amenable to epigenetic regulation to improve treatment outcomes.

• Authors: Guo-Dong Lu, Carol Ho-Wing Leung, Benedict Yan, Colyn Mui-Yeong Tan, Sie Yieh Low, Myat Oo Aung, Manuel Salto-Tellez, Seng Gee Lim, Shing Chuan Hooi
• Journal: Gastroenterology (Gastroenterology) Vol. 139 Issue 2 Pg. 632-43, 643.e1-4 (Aug 2010) ISSN: 1528-0012 [Electronic] United States
• PMID: 20347819 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright (c) 2010 AGA Institute. Published by Elsevier Inc. All rights reserved.

Chemical References

• CCAAT-Enhancer-Binding Proteins
• CEBPA protein, human
• Cyclin A
• Histones
• CDK4 protein, human
• Cyclin-Dependent Kinase 4

Topics

• Blotting, Western
• CCAAT-Enhancer-Binding Proteins (genetics, metabolism)
• Carcinoma, Hepatocellular (genetics, metabolism, pathology)
• Cell Nucleus (metabolism)
• Cell Proliferation
• Chromatin Immunoprecipitation
• Cyclin A (metabolism)
• Cyclin-Dependent Kinase 4 (metabolism)
• DNA Methylation
• Epigenesis, Genetic
• Gene Expression Regulation, Neoplastic
• Hep G2 Cells
• Histones (metabolism)
• Humans
• Immunohistochemistry
• Liver Neoplasms (genetics, metabolism, pathology)
• RNA Interference
• Reverse Transcriptase Polymerase Chain Reaction
• Up-Regulation