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Differential expression of genes related to contraction and inflammation in human aortic smooth muscle cells cultured on plastic or 3D‐Matrigel

We have found that human aortic smooth muscle cells grown in a monolayer on plastic plates (SMC‐P) assume a synthetic phenotype in appearance as well as in expression of contractile proteins while those cultured in 3D‐Matrigel (SMC‐M) assume the phenotype and morphology of a contractile muscle cell. To test the hypothesis that SMC‐M have a phenotype closer to a smooth muscle cell than SMC‐P we performed real time PCR for SM‐myosin mRNA and a gene array for extracellular matrix and adhesion molecules. Human aortic SMC were obtained from Cambrex and cells from passages 3–9 were seeded on 6 well plates, directly on plastic or on a growth factor depleted 3D‐Matrigel gel. SMC‐P were serum starved for 48 hrs prior to harvesting. RNA was isolated using the Trizol method, reverse transcribed and 1 μl used for the PCR reaction in a Roche LightCycler. Cells for the array were harvested in dispase and the total RNA was isolated and labeled following manufacturer’s instructions (SuperArray, Frederick, MD). Labeled cRNA was hybridized overnight on the OHS‐013 array. The gene array showed elevated expression in SMC‐P for connective tissue growth factor (CTGF), fibronectin, ICAM‐1, osteopontin, and tenascin C. A follow up RT‐PCR showed that CTGF was 21 fold greater in SMC‐P. Expression of SM‐1 myosin was 37 fold greater in SMC‐M than in SMC‐P. These results show that SMC‐P have a high expression of genes related to inflammation and growth and may be representative of the synthetic phenotype found in the sub‐intimal space in atherosclerosis or restenosis. Also, the increased expression of contractile proteins in SMC‐M makes the latter a better model for in vitro studies of SMC resembling the contractile cells of the media of the vascular wall.