Int. J. Mol. Sci. 2014, 15(1), 525-544; doi:10.3390/ijms15010525
Article

Enhanced Antitumor Effects of Adenoviral-Mediated siRNA against GRP78 Gene on Adenosine-Induced Apoptosis in Human Hepatoma HepG2 Cells

1,* email, 1email, 1email, 1email, 1email, 1email, 1email, 1email and 2email
Received: 13 November 2013; in revised form: 19 December 2013 / Accepted: 23 December 2013 / Published: 3 January 2014
(This article belongs to the Section Molecular Pathology)
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract: Our previous studies show that adenosine-induced apoptosis is involved in endoplasmic reticulum stress in HepG2 cells. In this study, we have investigated whether knockdown of GRP78 by short hairpin RNA (shRNA) increases the cytotoxic effects of adenosine in HepG2 cells. The adenovirus vector-delivered shRNA targeting GRP78 (Ad-shGRP78) was constructed and transfected into HepG2 cells. RT-PCR assay was used to determine RNA interference efficiency. Effects of knockdown of GRP78 on adenosine-induced cell viabilities, cell-cycle distribution and apoptosis, as well as relative protein expressions were determined by flow cytometry and/or Western blot analysis. The intracellular Ca2+ concentration was detected by laser scanning confocal microscope. Mitochondrial membrane potential (ΔΨm) was measured by a fluorospectrophotometer. The results revealed that GRP78 mRNA was significantly downregulated by Ad-shGRP78 transfection. Knockdown of GRP78 enhanced HepG2 cell sensitivity to adenosine by modulating G0/G1 arrest and stimulating Bax, Bak, m-calpain, caspase-4 and CHOP protein levels. Knockdown of GRP78 worsened cytosolic Ca2+ overload and ΔΨm loss. Knockdown of caspase-4 by shRNA decreased caspase-3 mRNA expression and cell apoptosis. These findings indicate that GRP 78 plays a protective role in ER stress-induced apoptosis and show that the combination of chemotherapy drug and RNA interference adenoviruses provides a new treatment strategy against malignant tumors.
Keywords: ER stress; glucose-regulated protein 78 (GRP78); adenosine; RNA interference; caspase-4
PDF Full-text Download PDF Full-Text [2973 KB, uploaded 19 June 2014 05:22 CEST]

Export to BibTeX |
EndNote


MDPI and ACS Style

Wu, L.-F.; Guo, Y.-T.; Zhang, Q.-H.; Xiang, M.-Q.; Deng, W.; Ye, Y.-Q.; Pu, Z.-J.; Feng, J.-L.; Huang, G.-Y. Enhanced Antitumor Effects of Adenoviral-Mediated siRNA against GRP78 Gene on Adenosine-Induced Apoptosis in Human Hepatoma HepG2 Cells. Int. J. Mol. Sci. 2014, 15, 525-544.

AMA Style

Wu L-F, Guo Y-T, Zhang Q-H, Xiang M-Q, Deng W, Ye Y-Q, Pu Z-J, Feng J-L, Huang G-Y. Enhanced Antitumor Effects of Adenoviral-Mediated siRNA against GRP78 Gene on Adenosine-Induced Apoptosis in Human Hepatoma HepG2 Cells. International Journal of Molecular Sciences. 2014; 15(1):525-544.

Chicago/Turabian Style

Wu, Ling-Fei; Guo, Yi-Tian; Zhang, Qing-Hua; Xiang, Meng-Qi; Deng, Wei; Ye, Yan-Qing; Pu, Ze-Jin; Feng, Jia-Lin; Huang, Guan-You. 2014. "Enhanced Antitumor Effects of Adenoviral-Mediated siRNA against GRP78 Gene on Adenosine-Induced Apoptosis in Human Hepatoma HepG2 Cells." Int. J. Mol. Sci. 15, no. 1: 525-544.

Int. J. Mol. Sci. EISSN 1422-0067 Published by MDPI AG, Basel, Switzerland RSS E-Mail Table of Contents Alert