IL1 AND GINGIVAL COLLAGENASE GENE REGULATION

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Interleukin-1 (IL-1) stimulates production of gingival fibroblast metalloproteinases (e . g. collagenase, stromelysin (proteoglycanase)) and thereby has been linked to gingival soft tissue destruction associated with periodontitis. The intracellular biochemical events that link IL-1 gingival fibroblast receptor activation to metalloproteinase gene activation are poorly understood. It is likely that a number of proteins (e.g. pretranscriptional/post-transcriptional regulatory factors, enzymes involved in signal transduction) play key roles in the modulation of these metalloproteinases. Recent studies with synovial and gingival fibroblasts in culture have provided new insights on the molecular events associated with the IL-1 induction of cellular metalloproteinases, there is much less information available on the induction of these metalloproteinases in gingival fibroblasts. Based on this, the objective of the present proposal involves a series of experiments designed to begin to identify six gingival fibroblast genes and products induced by IL-1 that could be involved in the modulation of metalloproteinases likely associated with periodontitis. The specific aims are to: (l) To obtain full length cDNA sequences for four IL- 1 induced genes (previously unidentified) obtained from human gingival fibroblasts, characterize them and express their proteins, (2) To determine if/how protein products derived from these three IL-1 induced genes and two other known genes (C/EBP and NFkB) upregulated by IL-1, functionally bind to the promoter region or modulate metalloproteinase transcription and (3) To determine if these selected genes functionally participate in the IL-1 induction of metalloproteinases. These aims will be accomplished by using: (a) gingival fibroblast cell lines and primary cultures derived from gingival tissue obtained from normal and patients with periodontitis; (b) full length cDNA clone isolation, characterization and sequencing; (c) computerized sequence analysis; (d) in vitro protein translation; (e) co- transfection, overexpression and underexpression (anti sense) studies; and (f) gel shift binding and foot printing assays. These proposed in-vitro studies with human gingival fibroblasts offer a model system to provide fundamental molecular information on gene activation induced by IL-1, and thereby identifying events which are likely involved in the pathogenesis of periodontitis. Identification of such genes holds promise for better defining the molecular basis of connective tissue destruction in periodontitis. Consequently, an important therapeutic implication of the present proposal is that blockers of these IL-1 induced biochemical events could be useful clinically for the future treatment of periodontitis.