Analysis of abnormal differentiation of smooth muscles in arterial lesions and an in vitro trial to induce smooth muscle cell differentiation

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Both proliferation and dedifferentiation involve the formation of arterial lesions, like arterio- and atherosclerosis or restenosis occurring following coronary angioplasy. Vascular smooth muscles change their phenotype from the contractile (adult type) to the synthetic state (embryonic type) during proliferation. In this project we examined when and how the phenotypic modulation of smooth muscles occurs and participates in the formation of vascular lesions in both animal models and human from the standpoint of gene expression of contractile proteins. We first isolated cDNA clones for three types of smooth muscle myosin heavy chain (MHC) isoforms (SM1, SM2 and SMemb) and developed sensitive immunohistology. In the ballooning-injured rabbit aortas, neointimal cells were found to be composed of smooth muscles with the embryonic phentotype at 2-3 weeks after injury, although they maintained the ability to re-differentiate to the adult phenotype in 4 weeks. In human coronary arteries, inti … More mal thickening started in the first decade and could reach 3-5 times thicker than the media by the fourth decades. The phenotypic modulation of smooth muscles in human coronary arteries was disappearance of SM2 and SM1 in the intima-media border regions, followed by accumulation of macrophages, thus indicating that human coronary atherosclerosis develops in a completely different fashion from those found in animal models. However, in the restenotic lesions of coronary arteries, we found smooth muscle cells with the embryonic phenotype vigorously proliferated. These results indicate that immunohistochemistry using anti-smooth muscle MHC isoforms is highly useful to study the process of athero- and arteriosclerosis.The promoter regions of SMemb and SM1/2 were also characterized in this study. The 5'-promoter region of the SMemb gene does not have a TATA box. However, a novel cis-element at around -100 bp seemed to play a key role in the activation of this gene. SM1 gene, on the other hand, has a TATA box as well as several cis-elements involved in muscle development and differentiation.Finally we tried to induce smooth muscle cell differentiation in vitro using a P19 mouse embryonic carcinoma cell line. We obtained evidence showing that P19 could be induced to smooth muscle cell with retinoic acid because SM1 MHC were positive under this regimen. Less