There is compelling evidence to suggest that fibroblast populations are heterogeneous in all tissues that have been examined. Preliminary and published data indicate increased production of macrophage-derived growth factor (MDGF) and the fibroblast inhibitory factor prostaglandin E2 (PGE2) by alveolar macrophages from bleomycin-induced fibrotic lung, which in conjunction with the suggestion of heterogeneity in the lung fibroblast population have created an environment conducive to a selection process and cause an alteration in the composition of the fibroblast population. Two hypotheses will be tested in this project. First, the heterogeneity of the normal lung fibroblast population is altered by growth and other fibrogenic factors in the fibrotic milieu by positive selection for cells with increased proliferative and collagen synthetic capacity. Second, a similar result is also obtained by a negative selection process in which subpopulations responsive to growth inhibitory mediators are depleted, leaving behind those subpopulations which are not subject to control by normal inhibitory influences. These hypotheses will be tested by examining the extent of heterogeneity of lung fibroblasts and comparing it to that in fibrotic lung as induced by endotracheal injection of bleomycin into mice. Clones and subpopulations will be obtained by limiting dilution techniques and flow sorting using the flow cytometer. The parameters which distinguish the subpopulations will be determined both in the absence, and in the presence of fibrogenic and anti-fibrogenic mediators. Two primary parameters will be used throughout these studies, namely, growth rate and collagen synthesis. When heterogeneity in responsiveness to a given mediator is detected, more detailed analysis will be undertaken to uncover the basis for the heterogeneity, such as by analysis of cell surface receptor density and effects on transcriptional regulation of collagen synthesis. The ability to induce alterations in the heterogeneity of normal lung fibroblast populations in vitro by treatment of these cells by fibrogenic and anti-fibrogenic mediators will be examined to determine if the in vivo fibrotic process can be reproduced in vitro. Finally, preliminary attempts will be made to evaluate the usefulness of bronchoalveolar lavage-derived fibroblast heterogeneity for evaluation and staging of the fibrotic process. The results of these studies should provide new insights as to the important mechanisms governing changes in lung fibroblast populations critical towards the development of fibrosis.
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