Bleomycin-induced pulmonary fibrosis in hamsters. An alveolar macrophage product increases fibroblast prostaglandin E2 and cyclic adenosine monophosphate and suppresses fibroblast proliferation and collagen production.

Bleomycin-induced pulmonary fibrosis in hamsters is associated with collagen accumulation that results from increased lung collagen synthesis rates. However, 1-2 wk after intratracheal instillation of bleomycin, lung collagen synthesis rates decline toward control values. To evaluate the potential role of the bronchoalveolar macrophage in regulating lung collagen production, we studied the effects of macrophages from normal and bleomycin-treated hamsters upon fibroblasts in vitro. We observed: (a) Medium from macrophage cultures decreased fibroblast [3H]thymidine incorporation and nondialyzable [3H]hydroxyproline production in a dose-dependent manner. Fibroblast cell counts were decreased in exposed cultures, and fibroblast viability was unchanged. Procollagen prolyl hydroxylation and prolyl-transfer RNA-specific activity were not altered by macrophage medium; this indicates that [3H]hydroxyproline reflects collagen production rate under the experimental conditions. (b) The suppressive effect of macrophage medium was selective for collagen since collagen production decreased more than noncollagen protein production. (c) Medium from bleomycin-treated hamster macrophages suppressed fibroblast proliferation and collagen production to a greater degree than medium from normal hamster macrophages. (d) Fibroblast suppression by macrophage medium was associated with increased fibroblast endogenous prostaglandin E2 production and intracellular cyclic AMP (cAMP). (e) Incubation of fibroblasts with indomethacin before exposure completely inhibited prostaglandin E2 production and increases in cAMP, and eliminated suppression of fibroblast proliferation and collagen production. The macrophage-derived suppressive factor has an apparent molecular weight of 20,000-30,000 and is heat stable. It does not appear to be species restricted since both hamster and human lung fibroblasts are similarly suppressed. It is at least in part preformed in macrophages obtained by lavage, but its production can also be stimulated in vitro. We concluded that alveolar macrophages release a product that stimulates endogenous fibroblast prostaglandin E2 production and cAMP formation with resultant suppression of fibroblast proliferation and collagen production. Enhanced release of suppressive factor by macrophages during a time when lung collagen production is declining in bleomycin-induced pulmonary fibrosis suggests that macrophages may limit collagen accumulation in pulmonary fibrosis.

Modulation of collagen production following bleomycin-induced pulmonary fibrosis in hamsters. Presence of a factor in lung that increases fibroblast prostaglandin E2 and cAMP and suppresses fibroblast proliferation and collagen production.

To elucidate mechanisms involved in the regulation of lung collagen content we studied hamsters with bleomycin-induced pulmonary fibrosis. Lung collagen in this model is increased as the result of greatly increased lung collagen synthesis rates. However, collagen synthesis rates are subsequently restored to normal. Hamster lung explants from both normal and bleomycin-exposed hamsters were cultured, and the effects of explant conditioned medium (CM) on lung fibroblast (IMR-90) proliferation and collagen production in vitro were determined. Lung explant CM increased fibroblast prostaglandin (PG)E2 production and intracellular cAMP, and decreased both fibroblast proliferation and collagen production in a dose-dependent manner. Greater activity was observed with lung explant CM from bleomycin-exposed lungs. Incubation of fibroblasts with indomethacin prior to addition of CM blocked CM-mediated changes in PGE2 and cAMP and inhibited changes in fibroblast proliferation and collagen production. Exogenous PGE2 or dibutyryl cAMP also suppressed fibroblast proliferation and collagen production. The suppressive activity in lung-conditioned medium is nondialyzable, has an apparent molecular weight of 15,000-20,000 by gel filtration, and is heat-stable. It is not species-restricted since CM from hamster lung affected human and hamster lung fibroblasts similarly. Activity is present preformed in lung and bronchoalveolar lavage fluid, although bronchoalveolar macrophages produce a nondialyzable factor in culture which suppresses fibroblast proliferation. The suppressive activity identified in fibrotic lung may represent a means for limiting collagen accumulation following tumor injury.

Collagen biosynthesis in bleomycin-induced pulmonary fibrosis in hamsters.

Bleomycin results in pulmonary interstitial fibrosis characterized by accumulation of collagen. We studied the mechanisms of this accumulation in hamsters 4 to 24 days after intratracheal injection of 1 U of bleomycin. Lung collagen was significantly increased within 11 days. Collagen synthesis was increased 10-fold in explant cultures obtained 8 days after injection. Prolyl hydroxylase activity and the degradation of newly synthesized collagen were also increased proportionally to collagen synthesis. Total protein synthesis also increased but not as much as collagen synthesis. We conclude that the accumulation of collagen in the lung following proceeds from an increased rate of collagen synthesis that is maximal early after injection and persists for at least 24 days after exposure.