Basic Study
Copyright ©The Author(s) 2015. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Jul 14, 2015; 21(26): 8032-8042
Published online Jul 14, 2015. doi: 10.3748/wjg.v21.i26.8032
Changes in gene expression in liver tissue from patients with fulminant hepatitis E
Anshu Naik, Amit Goel, Vinita Agrawal, Aditya N Sarangi, Nanda Chhavi, Vineeta Singh, Shahid Jameel, Rakesh Aggarwal
Anshu Naik, Amit Goel, Aditya N Sarangi, Nanda Chhavi, Rakesh Aggarwal, Department of Gastroenterology, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow 226014, India
Vinita Agrawal, Department of Pathology, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow 226014, India
Vineeta Singh, Sandor Proteomics Pvt Ltd, Hyderabad 500034, India
Shahid Jameel, Virology Group, International Centre for Genetic Engineering and Biotechnology, New Delhi 110067, India
Author contributions: Naik A carried out the laboratory work and data analysis, and prepared the first draft of the manuscript; Goel A carried out initial screening and identification of patients and liver biopsies, and helped in drafting and critical revision of the manuscript; Agrawal V supervised the histological and immunohistochemical studies; Sarangi AN did the microarray data analysis and helped in drafting the related segments of the manuscript, and in critical revision of the manuscript; Chhavi N carried out initial screening and identification of patients and liver biopsies; Singh V did the laboratory work related to microarray analysis; Jameel S conceived the idea, participated in design of the study and interpretation of data, and helped revise the manuscript; Aggarwal R conceived the idea, participated in design of the study, clinical identification of patients, supervision of laboratory work, analysis of microarray data and overall supervision of the work, and helped in drafting and revision of the manuscript; all authors read and approved the final manuscript.
Supported by National Institutes of Health, Bethesda, United States, No. 5R01AI076192.
Institutional review board statement: The study was reviewed and approved by Sanjay Gandhi Postgraduate Institute of Medical Sciences institutional review board. Written, informed consent was obtained from and all patient or their families, as appropriate.
Institutional animal care and use committee statement: Exempt, since this study did not include the use of any animals.
Conflict-of-interest statement: None of the authors listed on the above manuscript had any conflict of interest, either financial or non-financial in relation to this paper.
Data sharing statement: Participants gave informed consent for data sharing. The corresponding author would be willing to share the data as required for the above manuscript with the editors.
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: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Dr. Rakesh Aggarwal, Department of Gastroenterology, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow 226014, India. aggarwal.ra@gmail.com
Telephone: +91-522-2494431 Fax: +91-522-2668017
Received: January 29, 2015
Peer-review started: January 30, 2015
First decision: March 10, 2015
Revised: March 27, 2015
Accepted: May 2, 2015
Article in press: May 4, 2015
Published online: July 14, 2015
Processing time: 165 Days and 18.7 Hours
Abstract

AIM: To study host gene expression and number of immune cells in liver tissues from patients with fulminant hepatitis E (FH-E).

METHODS: Microarray-based expression profiling was done using Illumina Human WG-6_v3_BeadChip arrays on post-mortem liver tissue from 5 patients with FH-E, and compared with similar tissue from 6 patients with fulminant hepatitis B (FH-B; disease controls) and normal liver tissue from 6 persons. Differential expression was defined as ≥ 2.0-fold change with Benjamini-Hochberg false discovery rate below 0.05 using t-test in liver tissue from FH-B and FH-E, than healthy liver tissue. For some genes that showed differential expression in FH-E, microarray data were validated using quantitative reverse transcription PCR. Differentially expressed gene lists were then subjected to “Gene Ontology” analysis for biological processes, and pathway analysis using BioCarta database on the DAVID server. In addition, tissue sections were stained for CD4+, CD8+ and CD56+ cells using indirect immunohistochemistry; cells staining positive for each of these markers were counted and compared between groups.

RESULTS: Compared to normal livers, those from patients with FH-E and FH-B showed differential expression of 3377 entities (up-regulated 1703, downregulated 1674) and 2572 entities (up 1164, down 1408), respectively. This included 2142 (up 896, down 1246) entities that were common between the two sets; most of these belonged to metabolic, hemostatic and complement pathways, which are active in normal livers. Gene expression data from livers of patients with FH-E but not those of FH-B showed activation of several immune response pathways, particularly those involving cytotoxic T cells. The fold-change values of mRNA for selected genes in livers from FH-E than in normal liver tissue determined using quantitative reverse transcription PCR showed excellent concordance with microarray analysis. At immunohistochemistry, CD8+ T cells showed an increase in liver biopsies from both FH-E [median 53.4 per arbitrary unit area (range 31.2-99.9)] and FH-B [median 49.3 (19.3-51.0); P = 0.005] compared to control liver tissue [median 6.9 (3.1-14.9)].

CONCLUSION: FH-E patients show CD8+ T cell infiltration and increased gene expression of cytotoxic T cell pathways in liver, suggesting a possible pathogenetic role for these cells.

Keywords: Cytotoxic T cells; Gene expression; Hepatitis E; Hepatitis E virus; Immune response; Liver biopsy; Microarray; Natural killer cells; Pathogenesis

Core tip: Data on pathogenesis of hepatitis E virus (HEV) infection, which is a common cause of acute hepatitis in several developing countries, are quite limited. This manuscript reports our data on microarray-based gene expression analysis and immunohistochemistry in liver tissue from patients with HEV infection, as compared to liver tissue from patients with hepatitis B virus infection and normal liver tissue. These data advance the current knowledge about the pathogenesis of HEV infection.