Infectious agents, allergens and other noxious agents enter through exposed mucosal surfaces, such as the respiratory, gastrointestinal and genitourinary tracts. In response to these pathological agents inflammatory and immune cells are recruited and cross the epithelial barrier in response to a chemotactic gradient. To maintain their function as a barrier to infection, an adequate number of inflammatory cells must cross into the lumenal spaces. In addition, if the mucosal epithelium is compromised, the defense against infection is lost. The epithelial barrier musttherefore be restored as quickly as possible, to minimize the opportunity for entry of infectious agents. Matrix metalloproteinases (MMPs) are upregulated during lung injury, repair and inflammation. Transgenic mouse models with altered proteolytic potential present unique opportunities to elucidate important cellular and genetic pathways by which extracellular proteolysis participates in these epithelial defenses a mechanism. The overall goal of this proposal is to identify the pathways impacting epithelial defense (inflammatory cell recruitment, epithelial repair) regulated by MMPs and to identify their ma.ior tissue substrates. We will address the characteristics and functional significance of MMP interactions by combined in vivo and in vitro approaches. We propose to determine the role of MMPs in regulating chemoattractants for inflammatory cells in the bronchioalveolar spaces. We hypothesize that dysregulated airway MMP activity modifies the synthesis or release of chemoattractants and/or antagonists of chemoattractant action, into the airspaces, and thus chemoattractant gradients needed to attract the cells out of the parenchymal space into the airspaces are not formed or maintained. We will study this in vivo using micro-organisms or their products in models of infection or allergic sensitization in mice that have genetic modifications in MMP expression or activity, or treat mice with MMP inhibitors. We will evaluate cell recruitment as well as analyse the production of chemoattractants in the spaces. We then propose to determine the mechanism of action and molecular targets of MMPs that regulate inflammatory cell recruitment into the airspaces using normal and genetically modified epithelial cell in culture in collaboration with Project 3 and Cores B and C. We use both candidate and unbiased biochemical approaches. In collaboration with Projects 1 and 2 we will also investigate the role of MMPs in epithelial wound healing using MMP mutant mice, and cells and MMP inhibitors. The immediate implications of this work are in its applications to use of instilled chemoattractant agents and other molecules that modify epithelial defense and will provide proof-of-principle that these critical MMP substrates are potentially efficacious in intervention in epithelial defense mechanisms. Only a thorough understanding of the actions and effects of MMPs and their major substrates will help mitigate the defense against infection so that recruitment of adequate numbers of inflammatory cells into the spaces beyond the epithelial barrier and the ability of the epithelial to repair itself can be accelerated.
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