Green tea polyphenol epigallocatechin-3 gallate induces apoptosis of proliferating vascular smooth muscle cells via activation of p53.

Green tea polyphenols (GTPs), which possess antioxidant properties, have been shown to inhibit the development of atherosclerotic lesions. Epigallocatechin-3-gallate (EGCG), the most abundant GTP, displays antiproliferative effects in a variety of cell types. Here, we examined the effects of GTPs on aortic smooth muscle cell (SMC) proliferation. Treatment with a GTP mixture or EGCG at a dose of 40 to 50 microg/ml slowed SMC growth, while at a higher dose of 80 microg/ml EGCG also induced cell death as judged by TUNEL assay. Apoptosis was mainly observed in proliferating SMCs in subconfluent cultures; whereas at higher confluency, cell viability was largely unaffected. Treatment with 80 microg/ml EGCG induced the tumor suppressor p53, which was functional as judged by activation of the target cyclin-dependent kinase inhibitor p21CIP1. Inhibition of p53 activity with a dominant negative mutant reduced cell death. The increase in p53 protein was due to increased stability. EGCG also induced functional nuclear factor-kappaB (NF-kappaB) complexes, and inhibition of this activity reduced the extent of cell death. Thus, EGCG inhibits growth and induces death of SMCs in a p53- and NF-kappaB-dependent manner. These results provide evidence for a new molecular mechanism whereby green tea polyphenols inhibit SMC proliferation and function to prevent the development of atherosclerosis.

B-Myb represses vascular smooth muscle cell collagen gene expression and inhibits neointima formation after arterial injury.

OBJECTIVE: The function of B-Myb, a negative regulator of vascular smooth muscle cell (SMC) matrix gene transcription, was analyzed in the vasculature. METHODS AND RESULTS: Mice were generated in which the human B-myb gene was driven by the basal cytomegalovirus promoter, and 3 founders were identified. Mice appeared to develop normally, and human B-myb was expressed in the aortas. Total B-Myb levels were elevated in aortas of adult transgenic versus wild-type (WT) animals and varied inversely with alpha1(I) collagen mRNA expression. However, neonatal WT and transgenic aortas displayed comparable levels of alpha1(I) collagen mRNA, likely resulting from elevated levels of cyclin A, which ablated repression by B-Myb. Aortic SMCs from adult transgenic animals displayed decreased alpha1(I) collagen mRNA levels. To examine the role of B-Myb after vascular injury, animals were subjected to femoral artery denudation, which induces SMC-rich lesion formation. A dramatic reduction in neointima formation and lumenal narrowing was observed in arteries of B-myb transgenic versus WT mice 4 weeks after injury. CONCLUSIONS: Data indicate that B-Myb, which inhibits matrix gene expression in the adult vessel wall, reduces neointima formation after vascular injury. To analyze B-Myb function in the vasculature, mice overexpressing B-myb were generated. Neonates displayed normal alpha1(I) collagen mRNA levels, whereas adults expressed decreased collagen mRNA in aortas and isolated vascular SMCs. On femoral artery denudation, neointima formation was dramatically reduced in B-myb transgenic mice.

B-Myb represses elastin gene expression in aortic smooth muscle cells.

B-Myb represses collagen gene transcription in vascular smooth muscle cells (SMCs) in vitro and in vivo. Here we sought to determine whether elastin is similarly repressed by B-Myb. Levels of tropoelastin mRNA and protein were lower in aortas and isolated SMCs of adult transgenic mice expressing the human B-myb gene, driven by the basal cytomegalovirus promoter, compared with age-matched wild type (WT) animals. However, the vessel wall architecture and levels of insoluble elastin revealed no differences. Since elastin deposition occurs early in development, microarray analysis was performed using nontransgenic mice. Aortic levels of tropoelastin mRNA were low during embryonal growth and increased substantially in neonates, whereas B-myb levels varied inversely. Tropoelastin mRNA expression in aortas of 6-day-old neonatal transgenic and WT animals was comparable. Recently, we demonstrated that cyclin A-Cdk2 prevents B-Myb-mediated repression of collagen promoter activity. Cyclin A2 levels were higher in neonatal versus adult WT or transgenic mouse aortas. Ectopic cyclin A expression reversed the ability of B-Myb to repress elastin gene promoter activity in adult SMCs. These results demonstrate for the first time that B-Myb represses SMC elastin gene expression and that cyclin A plays a role in the developmental regulation of elastin gene expression in the aorta. Furthermore, the findings provide additional insight into the mechanism of B-myb-mediated resistance to femoral artery injury.