title: Role of TLR gene expression and cytokine profiling in the immunopathogenesis of viral hepatitis E.
abstract: BACKGROUND
  The clinical manifestations of Hepatitis E virus (HEV) range from self-limiting acute viral hepatitis (AVH) to acute liver failure (ALF). The varied clinical course is thought to be immune-mediated. Toll-like receptors (TLRs) play a central role in sensing and initiating innate antiviral-response and downstream signaling of TLRs modulates cytokine production, thereby playing an important role in determining the disease course.
  
  
  OBJECTIVE
  The present study was designed to elucidate the role of TLRs and cytokine production in the immunopathogenesis of HEV.
  
  
  METHODS
  Peripheral blood mono-nuclear cells were separated from 50 AVH-HEV, 30 ALF-HEV patients and 50 healthy-controls. One-part of the PBMC was processed for RNA-extraction another pulsed with HEV-ORF2-peptide. Gene-expression levels of TLR (2-4, 7, and 8) were checked using semi-quantitative Real-time-PCR. Cytokine levels were analyzed using Cytokine-Bead-Array. TLR3-silencing experiments were performed and post-silencing cytokine levels were estimated.
  
  
  RESULTS
  TLR3 gene-expression in AVH was significantly higher than ALF (202.4±36.36 Vs 13.71±5.01; p<0.0001). Higher amount of both anti-and pro-inflammatory cytokines; IFNγ, TNF-α, IL10 and TGF-β were detected in the PBMC culture-supernatant of AVH Vs ALF (p<0.0001, p=0.0008, p=0.0002, p<0.0001 respectively). Post-silencing TLR3, significant decrease in IFNγ level was observed in the PBMC culture-supernatant (4.08±1.06 Vs 23.20±12.51; p=N0.0213).
  
  
  CONCLUSIONS
  TLR3 and IFNγ were found to play an important role in HEV disease pathogenesis. Patients capable of expressing high levels of TLR 3 and robust IFNγ response are able to limit the disease and recover uneventfully; while the patients with lower expression of TLR3 and IFNγ progress to ALF.
authors.affiliation: Department of Virology, Postgraduate Institute of Medical Education & Research, Chandigarh 160012, India. Electronic address: manasivirol@gmail.com.
authors.forename: Manasi
authors.initials: M
authors.lastname: Majumdar
authors.affiliation: Department of Virology, Postgraduate Institute of Medical Education & Research, Chandigarh 160012, India. Electronic address: virology_pgi@yahoo.com.
authors.forename: R K
authors.initials: RK
authors.lastname: Ratho
authors.affiliation: Department of Hepatology, Postgraduate Institute of Medical Education & Research, Chandigarh 160012, India. Electronic address: ykchawla@gmail.com.
authors.forename: Yogesh
authors.initials: Y
authors.lastname: Chawla
authors.affiliation: Department of Virology, Postgraduate Institute of Medical Education & Research, Chandigarh 160012, India. Electronic address: minipsingh@gmail.com.
authors.forename: Mini P
authors.initials: MP
authors.lastname: Singh
mesh_terms: D000293:Adolescent; D000328:Adult; D018450:Disease Progression; D005260:Female; D020869:Gene Expression Profiling / Q000379:methods; D016751:Hepatitis E / Q000235:genetics* / Q000276:immunology / Q000821:virology; D016752:Hepatitis E virus / Q000276:immunology* / Q000472:pathogenicity; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism*; D017114:Liver Failure, Acute / Q000235:genetics / Q000276:immunology / Q000821:virology*; D008297:Male; D051196:Toll-Like Receptor 3 / Q000235:genetics*; D051193:Toll-Like Receptors / Q000235:genetics; D015854:Up-Regulation; D055815:Young Adult
chemical_list: C573415:IFNG protein, human; C495344:TLR3 protein, human; D051196:Toll-Like Receptor 3; D051193:Toll-Like Receptors; D007371:Interferon-gamma
keywords: Cytokine; HEV-ORF2 peptide; Hepatitis E virus; Peripheral blood mono-nuclear cell; Toll like receptor