M Kawamoto, DJ Romberger, Y Nakamura, Y Adachi, L Tate, RF Ertl, JR Spurzem and SI Rennard
University of Nebraska Medical Center, Omaha 68198-5300, USA.

To elucidate bronchial epithelial cell (BEC)-fibroblast interactions with regard to extracellular matrix production, bovine BECs were cultured, and conditioned media were assayed for their effects on fibroblast extracellular matrix production. Bovine BECs were prepared by protease digestion and grown in serum-free medium. When confluent, 48-h conditioned medium (CM) was collected and added to confluent human fetal lung fibroblasts (HFL-1) with 10 micrograms/ml ascorbic acid every other day. Type I collagen and fibronectin production from fibroblasts were quantified by enzyme-linked immunosorbent assay. Fifty percent BEC CM caused an increase in type I collagen (582 +/- 92 pg/10(3) cells/h versus 220 +/- 42, P < 0.001) and in fibronectin (1,422 +/- 60 pg/10(3) cells/h versus 360 +/- 24, P < 0.001) production after 4 days. These observations were confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by autoradiography in HFL-1 cultures labeled with [14C]proline. Dose-dependent stimulation was observed in response to BEC CM. Stimulation of macromolecule release was accompanied by increased steady-state fibronectin and alpha 1 (I) collagen mRNA levels. Sephadex G-150 column chromatography of BEC CM revealed two distinct peaks of activity at approximate molecular weights of 25 kD and > 66 kD. Transforming growth factor beta (TGF- beta)-neutralizing antibody blocked the activity of both peaks, suggesting that TGF-beta produced by the epithelial cells may drive fibroblast matrix production and that a TGF-beta binding substance may be present, or that TGF-beta aggregation may occur. Because some partially purified preparations had increased stimulatory activity compared to crude supernatants, potential inhibitors of matrix production were also sought.(ABSTRACT TRUNCATED AT 250 WORDS)

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