Liver Cancer
Copyright ©The Author(s) 2005. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Sep 28, 2005; 11(36): 5627-5632
Published online Sep 28, 2005. doi: 10.3748/wjg.v11.i36.5627
Upregulation of human telomerase reverse transcriptase mRNA expression by in vitro transfection of hepatitis B virus X gene into human hepatocarcinoma and cholangiocarcinoma cells
Zhen-Liang Qu, Sheng-Quan Zou, Nai-Qiang Cui, Xian-Zhong Wu, Ming-Fang Qin, Di Kong, Zhen-Li Zhou
Zhen-Liang Qu, Nai-Qiang Cui, Xian-Zhong Wu, Ming-Fang Qin, Di Kong, Zhen-Li Zhou, Department of Surgery, Tianjin Nankai Hospital, Tianjin 300100, China
Sheng-Quan Zou, Department of Surgery, Tongji Hospital, Wuhan 430030, Hubei Province, China
Author contributions: All authors contributed equally to the work.
Correspondence to: Zhen-Liang Qu, MD, Department of Surgery, Tianjin Nankai Hospital, Tianjin 300100, China. zlqu2002@hotmail.com
Telephone: +86-22-27022268
Received: January 14, 2005
Revised: February 13, 2005
Accepted: February 18, 2005
Published online: September 28, 2005
Abstract

AIM: To study the changes of human telomerase reverse transcriptase (hTERT) mRNA expression in human hepatocarcinoma cell lines (HepG2) and cholangiocarcinoma cell lines (QBC939) after HBx gene transfection and to illustrate the significance of transcriptional regulation of hTERT gene by HBx gene in the carcinogenesis.

METHODS: HepG2 and QBC939 cell lines were cultured and co-transfected with eukaryotic expression vector containing the HBx coding region and cloning vector containing enhanced green fluorescent protein (EGFP) coding sequence using lipid-mediated gene transduction technique. Thirty-six hours after transfection, EGFP expression in cells was used as the indicator of successful transfection. Flow cytometry was performed to determine the transfection efficiency. Cells were harvested and total RNA was extracted using TRIzol® reagent. The expression of hTERT mRNA in HepG2 and QBC939 cell lines was assayed by reverse transcription-polymerase chain reaction. The expression of HBx protein in both cell lines was detected by immunocytochemical staining and Western blotting.

RESULTS: Flow cytometry showed that the transfection efficiency was 46.4% in HepG2 cells and 29.6% in QBC939 cells for both HBx gene expression vector and blank vector. The expression of hTERT mRNA was meaningfully increased in HepG2 and QBC939 cell lines when transfected with HBx gene expression vector compared to those transfected with OPTI-MEM medium and blank vector. Immunocytochemical staining and Western blotting revealed HBx protein expression in HepG2 and QBC939 cells only when transfected with HBx gene.

CONCLUSION: HBx gene transfection can upregulate the transcriptional expression of hTERT mRNA. The transactiv-ation of hTERT gene by HBx gene is a newfound mechanism for pathogenesis of hepatocarcinomas and cholangioca-rcinomas after HBV infection.

Keywords: Hepatocholangiocarcinoma, Human telomerase reverse transcriptase, Gene expression, Hepatitis B virus, X protein