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Heme-induced acute lung injury by inflammasome activation in vascular endothelium

Flage, Bethany/J (2018) Heme-induced acute lung injury by inflammasome activation in vascular endothelium. Master's Thesis, University of Pittsburgh. (Unpublished)

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Abstract

Acute lung injury (ALI) is a form of acute respiratory failure, having an estimated annual incidence of 200,000 with a 40% mortality rate in the United States alone. Patients suffering from hemolytic diseases such as sepsis, malaria, and sickle cell disease are placed at a greater risk of developing ALI because of the excess amount of heme release that occurs during the pathogenesis of these diseases. Free heme is known to be a danger associated molecular pattern (DAMP) molecule and previous reports from our lab have identified particularly toxic effects apply to the lung endothelial barrier, ultimately contributing to ALI. However, the cellular mechanism of how heme is able to cause endothelial damage remains largely unknown. Recent studies have revealed that heme is able to induce the activation of nod-like receptor 3 (NLRP3) inflammasome in monocytes, which is followed by the maturation and release of inflammatory molecules Interleukin-1β (IL-1β) and Interleukin-18 (IL-18). Therefore, in this study we hypothesized that heme induces NLRP3 in pulmonary microvascular endothelial cells, resulting in destructive inflammatory activity that leads to the development of ALI. To test this hypothesis, we challenged human primary microvascular endothelial cells (HMVEC-Ls) with heme and assessed cell death, barrier integrity, cytokine release, and NLRP3 activation. Heme challenge of HMVEC-Ls produced a 75% decrease in barrier resistance as compared to vehicle, measured via electrical cell-substrate impedance sensing (ECIS). This barrier instability was concomitant with a significant 3-fold increase in IL-18 (p=0.046) release and an increase in NLRP3 protein, measured through extracellular supernatant and western blot respectively. Importantly, inhibition of NLRP3 resulted in a 35% increase in barrier integrity, which corresponded to a 20% decrease in lung wet-to-dry weight ratio in mice that were injected with heme. These results demonstrate that heme mediated endothelial damage involves an inflammatory response associated with NLRP3 activation, which is a potential target for therapeutic intervention. Future studies are warranted to confirm heme-mediated inflammasome formation and activation. Identifying specific molecular and cellular mechanisms by which heme damages the vascular endothelium in the pathogenesis of ALI is important for decreasing morbidity among patients and improving the overall health of the public.


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Details

Item Type: University of Pittsburgh ETD
Status: Unpublished
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Flage, Bethany/Jbef52@pitt.edubef520000-0002-0319-6489
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Thesis AdvisorMailliard, Robbierbm19@pitt.edu
Committee MemberBility, Mosesmtbility@pitt.edu
Committee MemberRobbie, Mailliardrbm19@pitt.edu
Date: 28 June 2018
Date Type: Publication
Defense Date: 17 April 2018
Approval Date: 28 June 2018
Submission Date: 4 April 2018
Access Restriction: 1 year -- Restrict access to University of Pittsburgh for a period of 1 year.
Number of Pages: 68
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: acute lung injury, NLRP3, vascular endothelium, heme
Date Deposited: 28 Jun 2018 20:06
Last Modified: 28 Jun 2018 20:06
URI: http://d-scholarship.pitt.edu/id/eprint/34076

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