The macrophage may contribute to pulmonary fibrosis by releasing fibrogenic growth factors, but the mechanisms regulating the macrophage secretory repertoire in inflammation and repair have not been elucidated. this proposal addresses the hypothesis that the macrophage participates in inflammation and repair by the sequential expression of two distinct macrophage phenotypes; a """"""""degradative"""""""" phenotype, defined by the increased synthesis of lysosomal enzymes, and a """"""""reparative"""""""" phenotype, defined by the increased expression of fibrogenic growth factors, including IGF-1, fibronectin, and TGFbeta. We further hypothesize that these phenotypes are exclusive, and regulated by cytokines present at sites of inflammation (PDGF and TNF), and extracellular matrix (ECM) components generated during the inflammatory response. A corollary of the phenotype hypothesis is that fibrosis would result from the macrophage becoming """"""""locked"""""""" in the reparative state, rather than growth factor expression being """"""""turned-off"""""""" as repair is completed. Preliminary studies showing that macrophages both respond to PDGF by synthesizing lysosomal enzymes and express PDGF transcripts in response to an induction of the """"""""degradative"""""""" phenotype by the inflammatory stimulus beta1,3 glucan, suggest a novel role for PDGF in the autocrine regulation of the """"""""degradative"""""""" phenotype. In Phase I of this proposal, we will examine the validity of this hypothesis of autocrine regulation in an in vitro system of mouse bone-marrow derived macrophages by determining (1) the isoform specificity of the PDGF stimulated increase in lysosomal enzyme synthesis, (2) the kinetics and specificity of isoform expression of PDGF at the mRNA and protein levels in response to beta1,3 glucan, (3) the effect of blocking PDGF production on induction of the degradative phenotype by beta1,3 glucan, and (4) the sub-types and level of expression of PDGF receptors. Further preliminary studies suggest that TNF regulates the transition from the degradative to the reparative phenotype by down-regulating lysosomal enzyme synthesis while up-regulating fibrogenic gene expression. It will be determined if TNF acts by antagonizing PDGF expression. Preliminary studies suggest that specific receptor mediated interactions between macrophages and ECM components generated during the inflammatory process may stimulate fibrogenic gene expression. In Phase II of the proposal the hypothesis that ECM components regulate fibrogenic gene expression will be investigated by (1) determining if up-regulation and activation of ECM receptors occurs in vivo during inflammation and repair by examining the level of expression of the fibronectin receptor and the hyaluronic acid receptor (CD44) on wound macrophages and resident cells; and (2) determining the effect of ECM receptor activation by both soluble and cross-linked receptor ligation on fibrogenic gene and protein expression in elicited macrophages. The results of these studies will form the basis for future investigation of the role of macrophage phenotypes in the pathogenesis of pulmonary fibrosis in humans.
|Horton, M R; Shapiro, S; Bao, C et al. (1999) Induction and regulation of macrophage metalloelastase by hyaluronan fragments in mouse macrophages. J Immunol 162:4171-6|
|Horton, M R; Burdick, M D; Strieter, R M et al. (1998) Regulation of hyaluronan-induced chemokine gene expression by IL-10 and IFN-gamma in mouse macrophages. J Immunol 160:3023-30|
|Horton, M R; McKee, C M; Bao, C et al. (1998) Hyaluronan fragments synergize with interferon-gamma to induce the C-X-C chemokines mig and interferon-inducible protein-10 in mouse macrophages. J Biol Chem 273:35088-94|
|McKee, C M; Lowenstein, C J; Horton, M R et al. (1997) Hyaluronan fragments induce nitric-oxide synthase in murine macrophages through a nuclear factor kappaB-dependent mechanism. J Biol Chem 272:8013-8|
|Noble, P W; McKee, C M; Cowman, M et al. (1996) Hyaluronan fragments activate an NF-kappa B/I-kappa B alpha autoregulatory loop in murine macrophages. J Exp Med 183:2373-8|
|McKee, C M; Penno, M B; Cowman, M et al. (1996) Hyaluronan (HA) fragments induce chemokine gene expression in alveolar macrophages. The role of HA size and CD44. J Clin Invest 98:2403-13|
|Shin, H S; Drysdale, B E; Shin, M L et al. (1994) Definition of a lipopolysaccharide-responsive element in the 5'-flanking regions of MuRantes and crg-2. Mol Cell Biol 14:2914-25|