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postgraduate thesis: Comparison of influenza A virus induced apoptosis in human respiratoryepithelial cells: an in vitro and ex vivostudy

TitleComparison of influenza A virus induced apoptosis in human respiratoryepithelial cells: an in vitro and ex vivostudy
Authors
Advisors
Issue Date2011
PublisherThe University of Hong Kong (Pokfulam, Hong Kong)
Citation
Yuen, K. [阮潔雯]. (2011). Comparison of influenza A virus induced apoptosis in human respiratory epithelial cells : an in vitro and ex vivo study. (Thesis). University of Hong Kong, Pokfulam, Hong Kong SAR. Retrieved from http://dx.doi.org/10.5353/th_b4717701
AbstractHighly pathogenic avian influenza H5N1, which is panzootic in poultry, continues to spread and becomes endemic in Asia, Africa, and Europe. It causes human disease with high fatality (about 60%) and continues to pose a pandemic threat. The pathological lesions associated with human H5N1 disease is Acute Respiratory Distress Syndrome (ARDS). The biological basis underlying the development of ARDS in human H5N1 disease remains controversial. Clinical, animal and in vitro studies suggested that the differences between H5N1 influenza viruss and low pathogenic influenza viruses in regard to viral replication, tissue tropism and cytokine dysregulation may contribute to disease pathogenesis. We previously found delayed onset of apoptosis in influenza H5N1 virus infected human peripheral blood monocyte-derived macrophages. This may allow a longer survival time for the virus in target cells for prolonged viral replication, which may contribute to the pathogenesis of H5N1 disease. As human bronchial and alveolar epithelial cells are target cells of influenza virus, I explored if influenza H5N1 and H1N1 virus infected human respiratory epithelial cells displayed differential apoptotic response and dissected the apoptotic pathways triggered by influenza virus infection. In this study, the apoptotic response in highly pathogenic influenza H5N1 viruses, A/HK/483/97 and A/Vietnam/1203/04, their precursor avian influenza H9N2 virus, A/Quail/HK/G1/97, and seasonal H1N1 virus, A/HK/54/98 infected primary human alveolar and bronchial epithelial cells was compared by TUNEL. A delayed onset of apoptosis in influenza H5N1 viruses and avian H9N2 virus infected alveolar epithelial cells was observed; the pattern was similar in bronchial epithelial cells. Concomitantly, by Western blotting, a delay in caspase 3 activation in H5N1 virus (A/HK/483/97) infected alveolar epithelial cells compared to H1N1 virus (A/HK/54/98) infected cells was shown. Also, influenza H5N1 and H1N1 virus induced apoptosis through both intrinsic and extrinsic pathways in human alveolar epithelial cells. Chemokine IP-10 was differentially up-regulated in influenza H5N1 virus infected alveolar epithelial cells, but its relationship to the delayed onset of apoptosis requires further studies. TRAIL, an upstream signaling molecule of extrinsic apoptotic pathway, mRNA was up-regulated in influenza H5N1 infected alveolar epithelial cells but not in influenza H1N1 infected cells. Using recombinant viruses, I showed that the 627 amino acid residue on PB2 of H5N1 virus and mutation of amino acids on 253 and 591 residues on PB2 of H9N2 virus contribute to the TRAIL upregulation. Immunohistochemical staining of physiologically relevant ex vivo model of human bronchus showed that influenza H5N1 (A/Vietnam/3046/04) and H9N2 (A/Quail/HK/G1/97) virus did not infect human bronchi as well as human H1N1 (A/HK/54/98) virus. Profiling of apoptosis related genes showed that TRAIL tends to be up-regulated in H5N1 virus infected bronchi ex vivo. This study demonstrated the delayed onset of apoptosis by H5N1 virus infected respiratory epithelial cells may be a mean for influenza virus to have prolonged replication within the human respiratory tract and contribute to disease severity. The results generated provide a robust research agenda, yielding critical information that elucidate molecular mechanisms, such as TRAIL up-regulation, that may contribute to the virulence and pathogenesis in human H5N1 disease.
DegreeMaster of Philosophy
SubjectInfluenza A virus.
Avian influenza A virus.
Apoptosis.
Epithelial cells.
Respiratory organs - Diseases.
Dept/ProgramPathology
AwardHKU 3 Minute Thesis Award, 2rd Runner-up (2011)

 

DC FieldValueLanguage
dc.contributor.advisorNicholls, JM-
dc.contributor.advisorPeiris, JSM-
dc.contributor.advisorChan, MCW-
dc.contributor.authorYuen, Kit-man.-
dc.contributor.author阮潔雯.-
dc.date.issued2011-
dc.identifier.citationYuen, K. [阮潔雯]. (2011). Comparison of influenza A virus induced apoptosis in human respiratory epithelial cells : an in vitro and ex vivo study. (Thesis). University of Hong Kong, Pokfulam, Hong Kong SAR. Retrieved from http://dx.doi.org/10.5353/th_b4717701-
dc.description.abstractHighly pathogenic avian influenza H5N1, which is panzootic in poultry, continues to spread and becomes endemic in Asia, Africa, and Europe. It causes human disease with high fatality (about 60%) and continues to pose a pandemic threat. The pathological lesions associated with human H5N1 disease is Acute Respiratory Distress Syndrome (ARDS). The biological basis underlying the development of ARDS in human H5N1 disease remains controversial. Clinical, animal and in vitro studies suggested that the differences between H5N1 influenza viruss and low pathogenic influenza viruses in regard to viral replication, tissue tropism and cytokine dysregulation may contribute to disease pathogenesis. We previously found delayed onset of apoptosis in influenza H5N1 virus infected human peripheral blood monocyte-derived macrophages. This may allow a longer survival time for the virus in target cells for prolonged viral replication, which may contribute to the pathogenesis of H5N1 disease. As human bronchial and alveolar epithelial cells are target cells of influenza virus, I explored if influenza H5N1 and H1N1 virus infected human respiratory epithelial cells displayed differential apoptotic response and dissected the apoptotic pathways triggered by influenza virus infection. In this study, the apoptotic response in highly pathogenic influenza H5N1 viruses, A/HK/483/97 and A/Vietnam/1203/04, their precursor avian influenza H9N2 virus, A/Quail/HK/G1/97, and seasonal H1N1 virus, A/HK/54/98 infected primary human alveolar and bronchial epithelial cells was compared by TUNEL. A delayed onset of apoptosis in influenza H5N1 viruses and avian H9N2 virus infected alveolar epithelial cells was observed; the pattern was similar in bronchial epithelial cells. Concomitantly, by Western blotting, a delay in caspase 3 activation in H5N1 virus (A/HK/483/97) infected alveolar epithelial cells compared to H1N1 virus (A/HK/54/98) infected cells was shown. Also, influenza H5N1 and H1N1 virus induced apoptosis through both intrinsic and extrinsic pathways in human alveolar epithelial cells. Chemokine IP-10 was differentially up-regulated in influenza H5N1 virus infected alveolar epithelial cells, but its relationship to the delayed onset of apoptosis requires further studies. TRAIL, an upstream signaling molecule of extrinsic apoptotic pathway, mRNA was up-regulated in influenza H5N1 infected alveolar epithelial cells but not in influenza H1N1 infected cells. Using recombinant viruses, I showed that the 627 amino acid residue on PB2 of H5N1 virus and mutation of amino acids on 253 and 591 residues on PB2 of H9N2 virus contribute to the TRAIL upregulation. Immunohistochemical staining of physiologically relevant ex vivo model of human bronchus showed that influenza H5N1 (A/Vietnam/3046/04) and H9N2 (A/Quail/HK/G1/97) virus did not infect human bronchi as well as human H1N1 (A/HK/54/98) virus. Profiling of apoptosis related genes showed that TRAIL tends to be up-regulated in H5N1 virus infected bronchi ex vivo. This study demonstrated the delayed onset of apoptosis by H5N1 virus infected respiratory epithelial cells may be a mean for influenza virus to have prolonged replication within the human respiratory tract and contribute to disease severity. The results generated provide a robust research agenda, yielding critical information that elucidate molecular mechanisms, such as TRAIL up-regulation, that may contribute to the virulence and pathogenesis in human H5N1 disease.-
dc.languageeng-
dc.publisherThe University of Hong Kong (Pokfulam, Hong Kong)-
dc.relation.ispartofHKU Theses Online (HKUTO)-
dc.rightsThe author retains all proprietary rights, (such as patent rights) and the right to use in future works.-
dc.rightsCreative Commons: Attribution 3.0 Hong Kong License-
dc.source.urihttp://hub.hku.hk/bib/B47177019-
dc.subject.lcshInfluenza A virus.-
dc.subject.lcshAvian influenza A virus.-
dc.subject.lcshApoptosis.-
dc.subject.lcshEpithelial cells.-
dc.subject.lcshRespiratory organs - Diseases.-
dc.titleComparison of influenza A virus induced apoptosis in human respiratoryepithelial cells: an in vitro and ex vivostudy-
dc.typePG_Thesis-
dc.identifier.hkulb4717701-
dc.description.thesisnameMaster of Philosophy-
dc.description.thesislevelMaster-
dc.description.thesisdisciplinePathology-
dc.description.naturepublished_or_final_version-
dc.identifier.doi10.5353/th_b4717701-
dc.date.hkucongregation2012-
dc.description.awardHKU 3 Minute Thesis Award, 2rd Runner-up (2011)-

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