Date of Award
2007
Degree Type
Thesis
Degree Name
Doctor of Philosophy
Program
Pharmacology and Toxicology
Supervisor
Dr. David McCormack
Second Advisor
Dr. Sanjay Mehta
Abstract
This thesis has examined the role of nitric oxide (NO) in human pulmonary microvascular endothelial cell (PMVEC) barrier dysfunction under septic conditions. Initially, establishing an in vitro model to study sepsis induced human acute lung injury (ALI) was essential to focus our studies. The results of these investigations showed that HUVEC are perhaps a less suitable model than HPMVEC for research into the pathophysiologic mechanisms of septic EC injury and neutrophil-induced injury of relevance to human ALI. Subsequently, the pathophysiologic implications of NO, specifically in reference to PMVEC injury under septic conditions, including barrier dysfunction and neutrophil-PMVEC interactions was assessed. NO is thought to be toxic in inflammatory conditions such as sepsis and ALI/ARDS. Experiments described in this thesis show that inducible nitric oxide synthase (iNOS)-derived NO from human neutrophils, and not human PMVECs, contributes to septic PMVEC barrier dysfunction. Moreover, the studies suggests that peroxynitrite, the product of NO and O2-, and not simply NO by itself, mediated PMVEC barrier dysfunction. Furthermore, exogenous NO treatment decreased both cytomix induced neutrophil adhesion and trans-PMVEC neutrophil migration. In contrast, exogenous peroxynitrite increased both neutrophil- PMVEC adhesion and trans-PMVEC neutrophil migration, which was reversible with the addition of a peroxynitrite scavenger. Moreover, inhibiting neutrophil-PMVEC adhesion completely inhibited septic-induced trans-PMVEC albumin leak. In contrast, inhibition of trans-PMVEC neutrophil migration significantly increased albumin leak and human PMVEC barrier dysfunction. Directly stimulating trans-PMVEC neutrophil migration had no effect on trans-PMVEC albumin leak, suggesting that trans-PMVEC neutrophil migration itself does not directly lead to endothelial cell barrier dysfunction. This body of work has discovered NO can be detrimental in EC injury depending on the cell source of production. Additionally its damaging effects are likely due to the oxidation products of NO, such as peroxynitrite, rather than NO itself.
Recommended Citation
Shelton, Jennifer L,, "Human Pulmonary Microvascular Endothelial Cell Dysfunction under Septic Conditions" (2007). Digitized Theses. 5138.
https://ir.lib.uwo.ca/digitizedtheses/5138