title: Adenovirus-mediated overexpression of a cyclin-dependent kinase inhibitor, p57Kip2, suppressed vascular smooth muscle cell proliferation.
abstract: Vascular smooth muscle cell (VSMC) proliferation after arterial injury is playing a pivotal role in the pathogenesis of a number of vascular proliferative disorders including atherosclerosis and restenosis after balloon-mediated angioplasty. Thus, a better understanding of the molecular mechanisms underlying VSMC proliferation in response to injury would have important therapeutic implications. Cell proliferation is controlled by an intricate network of extracellular and intracellular signaling pathways which are processing various growth regulatory signals and integrating them into the basic cell-cycle regulatory machinery through control of cyclin dependent kinases (CDK). CDK are positively regulated by cyclins and negatively regulated by CDK inhibitory proteins (CKI). To dissect the role of CKI in VSMC proliferation, we prepared the replication-deficient adenovirus constructs expressing p21 family members (Ad-CKI), p21Waf1, p27Kip1 and p57Kip2, respectively, and investigated the effect of CKI overexpression on the proliferation of vascular smooth muscle cells. The overexpression of each CKI protein in cultured VSMC was confirmed by western blot analysis. Flowcytometric analysis revealed that the Ad-CKI infected VSMC were largely retained in G1 phase, and had significantly less G2/M fraction than control cells. The extent of DNA synthesis in VSMC was assessed by [3H]-thymidine uptake, and shown to be inhibited by Ad-CKI dose dependently. Among three CKIs tested in this study, p57Kip2 showed the most significant suppression of DNA synthesis. In order to investigate in vivo effect of p57Kip2 overexpression, Ad-p57 was locally delivered to the luminal wall of rabbit carotid arteries after balloon angioplasty. Histological examinations revealed that the local infection of Ad-p57 significantly suppressed the neointimal formation at the site of vascular injury. These results clearly demonstrated the antiproliferative role of p57Kip2 in VSMC, and also proposed a possibility of gene therapy approach for vascular proliferative disorders.
authors.affiliation: Hokkaido University Graduate School of Medicine, Sapporo 060-8638, Japan.
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Takagi
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D002455:Cell Division / Q000502:physiology; D002478:Cells, Cultured; D050761:Cyclin-Dependent Kinase Inhibitor p57; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors*; D004247:DNA / Q000096:biosynthesis; D004791:Enzyme Inhibitors / Q000378:metabolism*; D005434:Flow Cytometry; D018014:Gene Transfer Techniques; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000166:cytology*; D009687:Nuclear Proteins / Q000096:biosynthesis*; D016133:Polymerase Chain Reaction; D011817:Rabbits
chemical_list: D050761:Cyclin-Dependent Kinase Inhibitor p57; D004791:Enzyme Inhibitors; D009687:Nuclear Proteins; D004247:DNA; D018844:Cyclin-Dependent Kinases