Original Article
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World J Gastroenterol. Feb 21, 2013; 19(7): 1068-1078
Published online Feb 21, 2013. doi: 10.3748/wjg.v19.i7.1068
Long interspersed nuclear element ORF-1 protein promotes proliferation and resistance to chemotherapy in hepatocellular carcinoma
Fan Feng, Yin-Ying Lu, Fan Zhang, Xu-Dong Gao, Chuan-Fu Zhang, Alex Meredith, Zhong-Xian Xu, Yu-Tao Yang, Xiu-Juan Chang, Hong Wang, Jian-Hui Qu, Zhen Zeng, Jun-Lan Yang, Chun-Ping Wang, Yun-Feng Zhu, Jia-Jun Cui, Yong-Ping Yang
Fan Feng, Yin-Ying Lu, Xu-Dong Gao, Zhong-Xian Xu, Xiu-Juan Chang, Hong Wang, Jian-Hui Qu, Zhen Zeng, Chun-Ping Wang, Yong-Ping Yang, Clinical Research Center of Hepatocarcinoma, 302 Military Hospital, Beijing 100039, China
Fan Feng, Institute of Toxicology and Pharmacology, Chinese Academy of Military Medical Sciences, Beijing 100850, China
Fan Zhang, Jun-Lan Yang, Yun-Feng Zhu, Tumor Center, PLA General Hospital, Beijing 100853, China
Chuan-Fu Zhang, Jia-Jun Cui, Institute of Disease Control and Prevention, Chinese Academy of Military Medical Sciences, Beijing 100071, China
Alex Meredith, Jia-Jun Cui, Department of Cancer and Cell Biology, University of Cincinnati College of Medicine, Cincinnati, OH 45267, United States
Yu-Tao Yang, Beijing Institute for Neuroscience, Capital Medical University, Beijing 100069, China
Author contributions: Feng F, Lu YY and Zhang F contributed equally to this work; Lu YY, Zhang F, Gao XD, Xu ZX, Yang YT, Qu JH, Zeng Z and Chang XJ performed the majority of experiments; Zhang CF, Wang H, Yang JL, Wang CP and Zhu YF provided some reagents and analytical tools and were also involved in revising the manuscript; Feng F, Cui JJ and Yang YP designed the study; Feng F and Cui JJ wrote the manuscript; Meredith A refined the manuscript and the experiment.
Supported by The Key Scientific and Technological Research Foundation of the National Special Purpose Program, No. 2008ZX10002-018
Correspondence to: Yong-Ping Yang, Professor, Clinical Research Center of Hepatocarcinoma, 302 Military Hospital, No. 100, the 4th West Ring Road, Beijing 100039, China. yongpingyang@hotmail.com
Telephone: +86-10-63879193 Fax: +86-10-63879193
Received: July 20, 2012
Revised: December 4, 2012
Accepted: December 27, 2012
Published online: February 21, 2013
Abstract

AIM: To clarify the specific roles and mechanisms of long interspersed nuclear element-1 ORF-1 protein [human long interspersed nuclear element-1 (LINE-1), ORF-1p] in chemotherapeutic drug resistance and cell proliferation regulation in hepatocellular carcinoma (HCC) cells.

METHODS: MTT assays were performed to identify the effect of the chemotherapeutic drug toxicity on HepG2 cells. Cell proliferation inhibition and the IC50 were calculated by the Origin 8.0 software. Western blotting assays were performed to investigate whether LINE-1 ORF-1p modulates the expression of some important genes, including p53, p27, p15, Bcl-2, mdr, and p-gp. To corroborate the proliferation and anchor-independent growth results, the HepG2 cells were analyzed by flow cytometry to investigate the effect of LINE-1 ORF-1p on the apoptosis regulation.

RESULTS: LINE-1 ORF-1p contributed to the resistance to several chemotherapeutic drugs (cisplatin and epirubicin) in HepG2 cells. The IC50 of the epirubicin and cisplatin increased from 36.04 nmol/L to 59.11 nmol/L or from 37.94 nmol/L to 119.32 nmol/L. Repression of LINE-1 ORF-1p expression by the siRNA could markedly enhance the response of HepG2 cells to the epirubicin and cisplatin. The IC50 correspondingly decreased from 28.06 nmol/L to 3.83 nmol/L or from 32.04 nmol/L to 2.89 nmol/L. Interestingly, down-regulation of LINE-1 ORF-1p level by siRNA could promote the response of HepG2 cells to the paclitaxel. The IC50 decreased from 35.90 nmol/L to 7.36 nmol/L. However, overexpression of LINE-1 ORF-1p did not modulate the paclitaxel toxicity in HepG2 cells. Further Western blotting revealed that LINE-1 ORF-1p enhanced mdr and p-gp gene expression. As a protein arrested in the nucleus, LINE-1 ORF-1p may function through modulating transcriptional activity of some important transcription factors. Indeed, LINE-1 ORF-1p promoted HepG2 cell proliferation, anchor-independent growth and protected the cells against apoptosis through modulating the expression of p15, p21, p53, and Bcl-2 genes.

CONCLUSION: LINE-1 ORF-1p promotes HepG2 cell proliferation and plays an important role in the resistance of chemotherapeutic drugs. By establishing novel roles and defining the mechanisms of LINE-1 ORF-1p in HCC chemotherapeutic drug resistance and cell proliferation regulation, this study indicates that LINE-1 ORF-1p is a potential target for overcoming HCC chemotherapeutic resistance.

Keywords: Long interspersed nuclear element-1 ORF-1 protein, Hepatocellular carcinoma, Chemotherapeutic drugs, Multi-drug resistance