eprintid: 39277
rev_number: 10
userid: 9222
dir: disk0/00/03/92/77
datestamp: 2020-07-31 02:30:15
lastmod: 2020-07-31 02:30:15
status_changed: 2020-07-31 02:30:15
type: thesis_degree
metadata_visibility: show
contact_email: gwenkettenburg3@gmail.com
eprint_status: archive
creators_name: Kettenburg, Gwenddolen
creators_email: gwk5@pitt.edu
creators_id: gwk5
title: Developing a Model of H5N1 Influenza Pathogenesis in Precision-Cut Human Lung Slices
ispublished: unpub
divisions: sch_gsph_infectiousdiseasesmicrobiology
full_text_status: public
keywords: Influenza, cell death, pyroptosis, gasdermin-D, caspase-1, VX-765, precision cut lung slices, H5N1, disease, pathogenesis, apoptosis
abstract: Highly pathogenic avian H5N1, known as HPAI H5N1, is a strain of influenza that is highly contagious in poultry and is occasionally spread to people that have had close contact with infected poultry. In humans, the fatality rate is 60%, though person-to-person transmission rarely occurs. HPAI H5N1 causes severe acute respiratory distress syndrome (ARDS), fluid buildup in the lung alveoli, making it hard for the lungs to get adequate oxygen due to a severe inflammatory response, followed by an inflammatory response and subsequent epithelial cell death in the lungs. The mechanism for ARDS is poorly understood. While models of infection exist in lab animals, a representative ex vivo model of infection in humans is needed to study the severe outcome of disease. An emerging alternative to animal models and immortalized cell culture models is precision-cut tissue slices of the organ of interest, using human donors as a source of tissue.  In this study, I developed a model of H5N1-infected precision-cut lung porcine then human slices and utilized said model to elucidate the mechanism of ARDS by selectively inhibiting members of cell death pathways and observe changes in downstream cytokines. Fluorescent immunohistochemistry was used to visualize and quantify markers of cell death and infection using image quantification. Changes in IL-1B were observed by enzyme-linked immunosorbent assay (ELISA). Here, I demonstrate that pyroptotic cell death is induced in response to H5N1 infection ex vivo as demonstrated by reduced IL-1B levels in response to a caspase-1/4 inhibitor and a gasdermin-D (GSDMD) inhibitor. This approach could prove public health relevance in developing novel, host-directed therapies to treat severe influenza infection in humans by providing a model to easily test human responses.
date: 2020-07-30
date_type: published
pages: 103
institution: University of Pittsburgh
refereed: TRUE
etdcommittee_type: committee_chair
etdcommittee_type: committee_member
etdcommittee_type: committee_member
etdcommittee_name: Barratt-Boyes, Simon
etdcommittee_name: Mattila, Joshua
etdcommittee_name: Watkins, Simon
etdcommittee_email: smbb@pitt.edu
etdcommittee_email: jmattila@pitt.edu
etdcommittee_email: simon.watkins@pitt.edu
etdcommittee_id: smbb
etdcommittee_id: jmattila
etdcommittee_id: simon.watkins
etd_defense_date: 2020-05-27
etd_approval_date: 2020-07-30
etd_submission_date: 2020-06-24
etd_release_date: 2020-07-30
etd_access_restriction: immediate
etd_patent_pending: FALSE
thesis_type: thesis
degree: MS
citation:   Kettenburg, Gwenddolen  (2020) Developing a Model of H5N1 Influenza Pathogenesis in Precision-Cut Human Lung Slices.  Master's Thesis, University of Pittsburgh.    (Unpublished)  
document_url: http://d-scholarship-dev.library.pitt.edu/39277/1/KettenburgG_MSthesis_6_2020.pdf