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Developing a Model of H5N1 Influenza Pathogenesis in Precision-Cut Human Lung Slices

Kettenburg, Gwenddolen (2020) Developing a Model of H5N1 Influenza Pathogenesis in Precision-Cut Human Lung Slices. Master's Thesis, University of Pittsburgh. (Unpublished)

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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.


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Details

Item Type: University of Pittsburgh ETD
Status: Unpublished
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Kettenburg, Gwenddolengwk5@pitt.edugwk5
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Committee ChairBarratt-Boyes, Simonsmbb@pitt.edusmbb
Committee MemberMattila, Joshuajmattila@pitt.edujmattila
Committee MemberWatkins, Simonsimon.watkins@pitt.edusimon.watkins
Date: 30 July 2020
Date Type: Publication
Defense Date: 27 May 2020
Approval Date: 30 July 2020
Submission Date: 24 June 2020
Access Restriction: No restriction; Release the ETD for access worldwide immediately.
Number of Pages: 103
Institution: University of Pittsburgh
Schools and Programs: Graduate School of Public Health > Infectious Diseases and Microbiology
Degree: MS - Master of Science
Thesis Type: Master's Thesis
Refereed: Yes
Uncontrolled Keywords: Influenza, cell death, pyroptosis, gasdermin-D, caspase-1, VX-765, precision cut lung slices, H5N1, disease, pathogenesis, apoptosis
Date Deposited: 31 Jul 2020 02:30
Last Modified: 31 Jul 2020 02:30
URI: http://d-scholarship.pitt.edu/id/eprint/39277

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