ENDOTHELIAL-CELL EFFECT ON SMOOTH-MUSCLE CELL COLLAGEN-SYNTHESIS

Extracellular matrix (ECM) constitutes the bulk of mature intimal hype rplastic lesions. To determine if endothelial cells (ECs) inhibit smoo th muscle cell (SMC) collagen synthesis, bovine aortic SMCs were cultu red on plastic semipermeable membranes either alone or opposite ECs. S MCs were pulsed with 4 mu Ci/mi [H-3]proline for 3 days and collagen s ynthesis was measured as trichloroacetic acid-precipitable counts in c onditioned media and in cell lysate fractions. Each was standardized t o SMC DNA (cpm/mu g DNA). EC effect on SMC gene expression of type I c ollagen was studied using Northern analysis. To determine whether TGF- beta 1 activation is involved in collagen synthesis regulation, parall el studies were performed in the presence of a neutralizing antibody t o TGF-beta 1 (25 mu g/ml) or aprotinin (a plasmin inhibitor, 200 mu g/ ml). SMCs cocultured with ECs were either untreated or treated with 5 ng/ml TGF-beta 1. The effect of culture substrate on EC-SMC interactio n was studied by repeating experiments on type I collagen (CI), matrig el (MG), fibronectin (FN), and type IV collagen (CIV). Results: Compar ed to SMCs cultured alone, ECs inhibited SMC collagen synthesis by 40 +/- 5% (P < 0.01) and gene expression of type I collagen by 60 +/- 4% (P < 0.01). The addition of neutralizing TGF-beta 1 Ab or aprotinin to SMCs cultured alone had no effect on collagen synthesis. Collagen syn thesis in SMCs cultured alone on MG was reduced by 49% and by 16% on C IV when compared to SMCs cultured on plastic (plastic: 100 +/- 12% vs MG: 51 +/- 15% vs CIV: 84 +/- 18%, P = 0.10). Collagen synthesis was i ncreased in SMCs cultured alone on FN (195 +/- 14% vs plastic: 100 +/- 12, P < 0.001) and CI (207 +/- 17 vs plastic: 100 +/- 12, P < 0.001). ECs decreased SMC collagen synthesis by 18 +/- 4% when cocultured on CI (P < 0.05) and by 22 +/- 2% when cocultured on MG (P < 0.05). FN pr evented EC inhibition of SMC collagen synthesis. Conclusions: ECs inhi bit SMC collagen synthesis and downregulate SMC type I collagen gene e xpression. TGF-beta 1 does not appear to be involved in this process. Culture substrate composition can affect SMC collagen synthesis and EC modulation of this process. (C) 1997 Academic Press.