PROJECT SUIVIMARY (See instructions): Francisella tularensis (Ft) is an intracellular Gram-negative bacterium that is highly pathogenic when aerosolized. We reason that the remarkable pathogenicity of aerosolized Ft reflects unique microbial features that render local innate host defenses in the lung quantitatively or qualitatively ineffective and hypothesize that specific surface features of Ft, including extracellular capsule-like material (CLM), subvert local pulmonary innate defenses, resulting in blunted host response in the lung and consequently the observed virulence of aerosolized Ft. As part of our comprehensive assessment of Ft-ain/vay innate host defense interaction and the contribution of CLM, Project 2 will examine alveolar epithelial cells (AEC), pulmonary microvascular endothelial cells (PMVEC) and neutrophils that traverse Ft-stimulated AEC and PMVEC (rPMN) - three interconnected elements of early pulmonary innate host response. We will pursue the following two aims:
Aim 1 : To characterize the mechanisms and functional consequences of Ft resistance to endogenous pulmonary alveolar epithelial and endothelial cell defenses Do invading Ft or its CLM alter innate antimicrobial and proinflammatory responses of AEC and PMVEC? Do invading Ft alter PECAM-CD99 interactions involved in transmigration? Does Ft infection of AEC and PMVEC activate the inflammasome? Do interactions of AEC or PMVEC with Ft alter microbial phenotype and do such changes influence virulence? How do structural changes in CLM influence interactions between Ft and AEC or PMVEC and thereby alter PMN transmigration? How do ainway dendritic cells influence Ft invasion of AEC and AEC responses to invasion? Aim 2: To define the functional alterations In transmigrated PMN recruited by Ft that contribute to Ft virulence Does development of the rPMN phenotype require contact with Ft, shed bacterial components such as CLM, PMVEC-specific factor(s), or a combination of these elements? Does a similar phenotype develop in PMN that migrate across AEC? What is the functional phenotype of rPMN? What mechanism(s) underlie the depressed NADPH oxidase of rPMN? Do structural changes in Ft CLM alter interactions of Ft with PMVEC/AEC with subsequent effects on rPMN? Do rPMN that ingest Ft progress normally to undergo caspase-3-mediated apoptosis or are they redirected to a caspase-1-mediated proinflammatory cell death?
As part of our PPG, these studies will provide important new insights regarding innate immune function in the lung, in general, and in the pathogenesis of the severe pneumonia caused by aerosolized Ft. Such new information is a prerequisite for the development of novel interventions to prevent and to treat infections with this virulent organism.
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