Expression of Cyclin-Dependent Kinase Inhibitors in Vascular Disease

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Abstract
—Arterial lesions in cardiovascular diseases are characterized by proliferation and migration of smooth muscle cells as well as deposition of connective tissue matrix. Factors that stimulate vascular smooth muscle cell (VSMC) proliferation are well described; however, the role of proteins that limit intimal hyperplasia is not well understood. To examine the function of Kip/Cip and INK cyclin-dependent kinase inhibitors (CKIs) in vascular diseases, the expression of p27Kip1 and p16INK was examined in VSMCs in vitro and in porcine arteries and human atherosclerosis in vivo. Western blot and fluorescence activated cell-sorting analysis demonstrated that levels of p27Kip1, but not p16INK, increased during serum deprivation of primary VSMC cultures and caused G1 arrest. p27Kip1 inhibited Cdk2 activity, suggesting that Kip CKIs promote G1 arrest in VSMCs by binding cyclin E/Cdk2. In porcine arteries, p27Kip1, but not p16INK, was constitutively expressed at low levels. Immediately after balloon injury, cell proliferation increased as p27Kip1 levels declined. Three weeks after injury, p27Kip1 was strongly expressed in intimal VSMCs when VSMC proliferation was Kip1 functions as an inhibitor of cell proliferation in injured arteries. In contrast, p16INK expression was detected only transiently early after injury. CKI expression was examined in 35 human coronary arteries, ranging from normal to advanced atherosclerosis. p27Kip1 expression was abundant in nonproliferating VSMCs and macrophages within normal (7 of 8) and atherosclerotic (25 of 27) arteries. p21Cip1 levels were undetectable in normal arteries but were elevated in atherosclerotic (19 of 27) arteries. p16INK could not be detected in normal or atherosclerotic arteries (0 of 35). Thus, the Kip/Cip and INK CKIs have different temporal patterns of expression in VSMCs in vitro and in injured arteries and atherosclerotic lesions in vivo. In contrast to p16INK, p27Kip1 likely contributes to the remodeling process in vascular diseases by the arrest of VSMCs in the G1 phase of the cell cycle.