Basic Study
Copyright ©The Author(s) 2015. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Dec 21, 2015; 21(47): 13225-13239
Published online Dec 21, 2015. doi: 10.3748/wjg.v21.i47.13225
Hepatitis B and C virus-induced hepatitis: Apoptosis, autophagy, and unfolded protein response
Behzad Yeganeh, Adel Rezaei Moghadam, Javad Alizadeh, Emilia Wiechec, Seyed Moayed Alavian, Mohammad Hashemi, Bita Geramizadeh, Afshin Samali, Kamran Bagheri Lankarani, Martin Post, Payam Peymani, Kevin M Coombs, Saeid Ghavami
Behzad Yeganeh, Martin Post, Department of Physiology and Experimental Medicine, Hospital for Sick Children Research Institute, University of Toronto, Toronto M5G 0A4, Canada
Adel Rezaei Moghadam, Young Researchers and Elite Club, Ardabil Branch, Islamic Azad University, Ardabil 156131-56491, Iran
Javad Alizadeh, Saeid Ghavami, Department of Human Anatomy and Cell Science, College of Medicine, Faculty of Health Sciences, University of Manitoba, Winnipeg R3E 0J9, Canada
Javad Alizadeh, Kevin M Coombs, Saeid Ghavami, The Children's Hospital Research Institute of Manitoba, University of Manitoba, Winnipeg R3E 0J9, Canada
Emilia Wiechec, Department of Clinical and Experimental Medicine, Division of Otorhinolaryngology, Linköping University, 581-85 Linköping, Sweden
Seyed Moayed Alavian, Baqiyatallah Research Center for Gastroenterology and Liver Diseases (BRCGL), 1417-613151 Tehran, Iran
Mohammad Hashemi, Department of Clinical Biochemistry, School of Medicine, Zahedan University of Medical Sciences, Zahedan 43181-98167, Iran
Bita Geramizadeh, Department of Pathology and Organ Transplant Research Center, Shiraz University of Medical Sciences, Shiraz 71348, Iran
Afshin Samali, Department of Biochemistry, National University of Ireland Galway, 000 Galway, Ireland
Kamran Bagheri Lankarani, Payam Peymani, Saeid Ghavami, Health Policy Research Centre, Shiraz University of Medical Sciences, Shiraz 71348-45794, Iran
Kevin M Coombs, Manitoba Center for Proteomics and Systems Biology, University of Manitoba, Winnipeg R3E 0J9, Canada
Kevin M Coombs, Department of Medical Microbiology and Infectious Diseases, The Children's Hospital Research Institute of Manitoba, University of Manitoba, Winnipeg R3E 0J9, Canada
Author contributions: Yeganeh B and Post M performed fluorescence immunohistochemistry, tissue array; Yeganeh B interpreted the data, drafted the whole primary version of manuscript; Rezaei Moghadam A and Alizadeh J analyzed the fluorescence immunohistochemistry results, participated in preparing the apoptosis and autophagy parts of manuscript, respectively; Wiechec E analyzed the fluorescence immunohistochemistry and tissue array results, drafted of the autophagy and apoptosis part of manuscript; Alavian SM diagnosed and got liver biopsy of HBV patients; Hashemi M contributed PCR or samples for HBV and HCV virus titer, and analyzed the fluorescence immunohistochemistry results; Geramizadeh B contributed pathology diagnosis, scoring, and graded of HBV and HCV biopsies; Samali A contributed unfolded protein response, and proof final versions of the manuscript; Bagheri Lankarani K diagnosed and got liver biopsy of HCV patients; Peymani P prepared liver biopsy sample, biochemical liver function, statistical analysis; Coombs KM and Ghavami S designed the project, and proofed final versions of the manuscript; Ghavami S prepared primary draft of UPR part of manuscript; Peymani P, Coombs KM and Ghavami S had equal senior authorship.
Supported by University of Manitoba Start-up funds and an award from the Manitoba Medical Service Foundation to Ghavami S; University of Manitoba Start-up Funds to Alizadeh J.
Institutional review board statement: This retrospective study was approved by local research ethics committee of Health policy research Center (Protocol number HP-101-91). All patients were informed about the study and gave verbal informed consent prior to enrollment.
Conflict-of-interest statement: All authors do not have any conflict of interest.
Data sharing statement: This is an open access work and the researcher can use the data for any further investigations.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See:
Correspondence to: Saeid Ghavami, PhD, Assistant Professor, Department of Human Anatomy and Cell Science, College of Medicine, Faculty of Health Sciences, University of Manitoba, Winnipeg R3E 0J9, Room 104, Canada.
Telephone: +1-204-2723061 Fax: +1-204-7893920
Received: August 29, 2015
Peer-review started: August 30, 2015
First decision: September 29, 2015
Revised: October 14, 2015
Accepted: November 13, 2015
Article in press: November 13, 2015
Published online: December 21, 2015

AIM: To investigate the co-incidence of apoptosis, autophagy, and unfolded protein response (UPR) in hepatitis B (HBV) and C (HCV) infected hepatocytes.

METHODS: We performed immunofluorescence confocal microscopy on 10 liver biopsies from HBV and HCV patients and tissue microarrays of HBV positive liver samples. We used specific antibodies for LC3β, cleaved caspase-3, BIP (GRP78), and XBP1 to detect autophagy, apoptosis and UPR, respectively. Anti-HCV NS3 and anti-HBs antibodies were also used to confirm infection. We performed triple blind counting of events to determine the co-incidence of autophagy (LC3β punctuate), apoptosis (cleaved caspase-3), and unfolded protein response (GRP78) with HBV and HCV infection in hepatocytes. All statistical analyses were performed using SPSS software for Windows (Version 16 SPSS Inc, Chicago, IL, United States). P-values < 0.05 were considered statistically significant. Statistical analyses were performed with Mann-Whitney test to compare incidence rates for autophagy, apoptosis, and UPR in HBV- and HCV-infected cells and adjacent non-infected cells.

RESULTS: Our results showed that infection of hepatocytes with either HBV and HCV induces significant increase (P < 0.001) in apoptosis (cleavage of caspase-3), autophagy (LC3β punctate), and UPR (increase in GRP78 expression) in the HCV- and HBV-infected cells, as compared to non-infected cells of the same biopsy sections. Our tissue microarray immunohistochemical expression analysis of LC3β in HBVNeg and HBVPos revealed that majority of HBV-infected hepatocytes display strong positive staining for LC3β. Interestingly, although XBP splicing in HBV-infected cells was significantly higher (P < 0.05), our analyses show a slight increase of XBP splicing was in HCV-infected cells (P > 0.05). Furthermore, our evaluation of patients with HBV and HCV infection based on stage and grade of the liver diseases revealed no correlation between these pathological findings and induction of apoptosis, autophagy, and UPR.

CONCLUSION: The results of this study indicate that HCV and HBV infection activates apoptosis, autophagy and UPR, but slightly differently by each virus. Further studies are warranted to elucidate the interconnections between these pathways in relation to pathology of HCV and HBV in the liver tissue.

Keywords: Cell fate, Cell death, Hepatocyte, Viral infection, Endoplasmic reticulum stress

Core tip: For the first time, the current study has addressed the co-incidence of apoptosis, autophagy, and unfolded protein response in the liver tissue of hepatitis B and hepatitis C (HBV, and HCV) infected patients. The results showed that all of these events are activated at the same time by HBV and HCV infection in the liver. All of these pathways probably are involved in replication and pathogenesis of HBV and HCV infection; therefore their modulation would probably be beneficial for new therapeutic approaches in these diseases.