Stricture formation is a common complication of Crohn's disease resulting in severe morbidity, intestinal obstruction and surgical intervention. The overall objective of this proposal is to determine the pathologic mechanism of this stricturing process with the ultimate goal of providing modalities for therapeutic intervention. We have demonstrated that these strictures result from increases in collagen and muscle in the bowel wall and that the accumulating collagen is synthesized by smooth muscle cells in response to chronic inflation. Using a human intestinal smooth muscle (HISM) cell culture model we have demonstrated that these cells make large amounts of collagen, particularly when stimulated by TGF-beta, and that their proliferation is stimulated by PDGF and IL-I. The current proposal will determine: 1) which inflammatory mediators are present in strictured intestine in vivo - immunohistochemical studies of strictured bowel, inflamed bowel without strictures and normal resection margins will be performed with antibodies to TGF-beta, PDGF and TNF; 2) what is the mechanism of regulation of HISM cell collagen production by inflammatory mediators - the now well-characterized in vitro smooth muscle cell model will be utilized to correlate effects of these mediators on net collagen synthesis with effects on collagen turnover, collagen mRNA levels, collagenase production and cyclic nucleotide production; 3) whether heparin, protamine or corticosteroids inhibit the stimulation of HISM cells by inflammatory mediators; and 4) whether smooth muscle cells isolated from strictured bowel are phenotypically different from cells isolated from normal bowel. The growth, collagen production and response to inflammatory mediators of the two groups of cells will be compared in vitro. Current technology available to the collagen biochemist and cell biologist has not been previously applied to the fibrosis complicating Crohn's disease. This proposal will provide information that will contribute significantly to an understanding of the mechanism of stricture formation in this devastating disease and forms the basis for future therapies to be used for its control and amelioration.
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