Endothelial Cell Effect on Smooth Muscle Cell Collagen Synthesis

• Published in: The Journal of surgical research 1997-04, Vol.69 (1), p.113-118
• Main Authors: Powell, Richard J., Hydowski, James, Frank, Oleg, Bhargava, Jaya, Sumpio, Bauer E.
• Format: Article
• Language: English
• Subjects: Animals; Biological and medical sciences; Cattle; Cell interactions, adhesion; Collagen - biosynthesis; Collagen - genetics; Endothelium, Vascular - cytology; Endothelium, Vascular - physiology; Extracellular Matrix Proteins - physiology; Fundamental and applied biological sciences. Psychology; Gene Expression; Molecular and cellular biology; Muscle, Smooth, Vascular - cytology; Muscle, Smooth, Vascular - metabolism; RNA - metabolism; Transforming Growth Factor beta - physiology
• ISSN: 0022-4804; 1095-8673
• DOI: 10.1006/jsre.1997.5045

Extracellular matrix (ECM) constitutes the bulk of mature intimal hyperplastic lesions. To determine if endothelial cells (ECs) inhibit smooth muscle cell (SMC) collagen synthesis, bovine aortic SMCs were cultured on plastic semipermeable membranes either alone or opposite ECs. SMCs were pulsed with 4 μCi/ml [3H]proline for 3 days and collagen synthesis was measured as trichloroacetic acid-precipitable counts in conditioned media and in cell lysate fractions. Each was standardized to SMC DNA (cpm/μg DNA). EC effect on SMC gene expression of type I collagen was studied using Northern analysis. To determine whether TGF-β1 activation is involved in collagen synthesis regulation, parallel studies were performed in the presence of a neutralizing antibody to TGF-β1 (25 μg/ml) or aprotinin (a plasmin inhibitor, 200 μg/ml). SMCs cocultured with ECs were either untreated or treated with 5 ng/ml TGF-β1. The effect of culture substrate on EC–SMC interaction was studied by repeating experiments on type I collagen (CI), matrigel (MG), fibronectin (FN), and type IV collagen (CIV). Results: Compared 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-β1 Ab or aprotinin to SMCs cultured alone had no effect on collagen synthesis. Collagen synthesis in SMCs cultured alone on MG was reduced by 49% and by 16% on CIV when compared to SMCs cultured on plastic (plastic: 100 ± 12% vs MG: 51 ± 15% vs CIV: 84 ± 18%,P= 0.10). Collagen synthesis was increased 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 prevented EC inhibition of SMC collagen synthesis. Conclusions: ECs inhibit SMC collagen synthesis and downregulate SMC type I collagen gene expression. TGF-β1 does not appear to be involved in this process. Culture substrate composition can affect SMC collagen synthesis and EC modulation of this process.