To understand how microbial colonization results in tissue destruction, a study is proposed of the molecular and cellular mechanisms of normal and microbially-induced collagen breakdown by resident human gingival cells (fibroblasts, endothelial cells, epithelial cells) and by migrant cells of the immune system (PMN leukocytes, macrophages). 1. The component proteases of the fibroblast collagenolytic cascade, including procollagenase activator, collagenase, gelatinase, and, to a limited extent, plasminogen activators will be isolated from culture media by conventional and HPLC methods and used to produce murine monoclonal and rabbit polyclonal antibodies of unequivocal specificity. The homology of collagenolytic cascades in diverse cell types and relationship of isozymes of individual cascade components will be examined by determination of immunologic cross reactivity. Rate limiting plasmin-dependent and independent cascade pathways for procollagenase activation will be examined and the role of individual cascade components in extracellular collagent breakdown determined. 2. The localization and codistribution of collagenolytic proteases in single cells, colonies of cells and tissue specimens will be studied by fluorescent and ultrastructural immunohistochemical methods, and the role of cellular activation in expression of degradative functions will be examined. In addition, the role of collagenolytic cascade proteases in phagocytic collagen breakdown, and the relationship of extracellular and phagocytic collagen breakdown mechanisms will be studied. 3. The ability of suspected periodontopathogens (prototype strains and wild type isolates) and of unknown cultivatable microorganisms from periodontal disease sites to elicit cytostatic or cytocidal responses in host cells will be investigated. Moreover, the ability of selected prototype, wildtype and unknown strains to generate mitogenic and general activated responses in host cells resulting in induction of collagen degradation will be identified. The responsible microbial mediators will be isolated and characterized and the in site of action in the cascade investigated.
