Variability of BCP and Precore Region Among the Occult HBV Infected Voluntary Blood Donors from Eastern India

Abstract

© 2011, INASL 2 PBMCs/well were stimulated with ORF-2 peptide (452–617 a.a) and subjected to MTT assay. Proliferation index (PI) at 570 nm was calculated. All the samples were subjected for anti-HEV IgM, IgG serology and HEV RNA detection by RT-PCR. Results: Patients with hepatitis were diagnosed as acute hepatitis E based on anti-HEV IgM/RNA positivity. PBMCs of acute hepatitis patients showed reactivity to the ORF-2 peptide in comparison to control subjects in terms of blast cell transformation. The proliferation indices of patient and control groups were 3.0398 ± 0.666 and 1.21 ± 0.077, respectively. Thus significant antigenic reactivity (p = 0.0126) was observed in hepatitis E patients. Anti-HEV IgM was positive in 15 (93.75%), IgG in 10 (62.5%) and RNA in 7 (43.75%) patients. None of the parameters were positive in control group. There was no significant difference in PI values between patient groups with and without HEV RNA positivity (2.99 ± 0.664, 3.077 ± 0.705; p = 0.359). Conclusion: HEV specific ORF 2 peptide (452–617 a.a) effectively stimulates the PBMCs in the patient group as compared to healthy controls. Thus, the tested peptide was found to be antigenic and specific for HEV and can be used in understanding the pathogenesis of HEV infection in vitro with regards to gene modulation and various cytokine profiles. Conflict of Interest: None Gene Expression Analysis in Peripheral Blood Mononuclear Cells in Patients with Acute Hepatitis E A Naik*, A Goel*, D Goel*, A Pandey**, S Rastogi†, R Aggarwal* *Department of Gastroenterology, Sanjay Gandhi Postgraduate Institute of Medical Sciences, Lucknow **Department of Obstetrics and Gynecology, King George’s Medical College, Lucknow †Command Hospital, Lucknow Background: Hepatitis E is the commonest cause of acute viral hepatitis in India and several developing countries. Pathogenesis of liver injury and immune events during this disease are not well understood. To better define immune events during hepatitis E, we undertook a gene expression study of peripheral blood mononuclear cells in patients with this disease. Methods: PBMCs were obtained from 12 patients with acute hepatitis E (typical clinical and biochemical findings, and positive IgM anti-HEV [Genelabs, Singapore]), 4 patients with acute hepatitis B (HBsAg positive, IgM anti-HBc positive, IgM anti-HEV negative) and 10 healthy controls, using Ficoll-Hypaque density gradient centrifugation. Total RNA was isolated from PBMCs using Qiagen RNeasy Protect Mini Kit, and cDNA synthesis and microarray analysis were done using Illumina HumanWG-6 v3.0 Expression BeadChips. Genes showing differential expression in hepatitis E, but not in hepatitis B, were identified and downstream pathway analysis was done. Results: PBMCs from patients with hepatitis E showed differential expression of 170 genes (upregulated 140, downregulated 30). PBMCs from patients with hepatitis B showed differential expression of 80 genes (up 62, down 18). Of these, 24 differentially-expressed genes were common. Genes differentially expressed in PBMCs from patients with hepatitis E but not in hepatitis B belonged to immune pathways that included cell-mediated immune response, immune cell trafficking and antigen presentation. The genes that were overexpressed in PBMCs from patients included CDCA5, CCNA2, CCNB2, PCNA, IGLL1 and TNFRSF17; most of these genes are involved in cell division and cellular function. In comparison, most of the genes that were underexpressed in PBMCs from hepatitis E (CCL20, TNF, IL1RN, CCL4L1, CCL3L3, CCL3, CCL3L1, IL1RN and IL1B) belonged to chemokine and pro-inflammatory cytokine families. Conclusion: Downregulation of chemokines and pro-inflammatory cytokines in PBMCs from patients with hepatitis E may contribute to disease pathogenesis by allowing enhanced viral replication. Conflict of Interest: None Variability of BCP and Precore Region Among the Occult HBV Infected Voluntary Blood Donors from Eastern India A Biswas*, R Panigrahi*, A Pal*, M Bandopadhyay*, BK De**, S Chakrabarti†, R Chakravarty* *ICMR Virus Unit, Kolkata **Department of Medicine, Calcutta Medical College, Kolkata †National Institute of Cholera and Enteric Diseases, Kolkata Background: In India prevalence of occult hepatitis B virus (HBV) infection in the general population is widespread. Data about the variability of the basal core promoter region (BCP) and precore (PC) region of the occult HBV strains is lacking. Objective: Mutations in the BCP and PC regions are associated with persistent/intermittent HBV replication. We evaluated the variability BCP and PC region of HBV in asymptomatic carriers and aimed to characterize the variability with different viral strains. Methods: From voluntary blood donation camps initially 1400 blood samples were collected. The anti-HBc and HBsAg status of the samples were confirmed by ELISA. PCR amplification to test the presence of HBV DNA was done from anti-HBc positive but HBsAg 03_JCEH-Abstract.indd 2 3/18/2011 11:13:03 AM