(Circulation. 2001;103:407.)
© 2001 American Heart Association, Inc.
Basic Science Reports |
Growth Suppression of Human Coronary Vascular Smooth Muscle Cells by Gene Transfer of the Transcription Factor E2F-1
From the Wafic Said Molecular Cardiology and Gene Therapy Research Laboratory, Texas Heart Institute (H.S.S., M.K.B., P.M.D., J.T.W., P.Z.); the Departments of Medicine (H.S.S., P.M.D., J.T.W., P.Z.), Pathology (D.L.H.-B.), and Integrative Biology (T.V.), University of Texas–Houston Medical School; and the Department of Neuro-Oncology, M.D. Anderson Cancer Center (T.-J.L.), Houston, Tex.
Correspondence to Pierre Zoldhelyi, Texas Heart Institute MC 2-255, 1101 Bates St, Houston, TX 77030. E-mail zoldhelyi{at}aol.com
Background—The transcription factor E2F-1 promotes S-phase entry and death in transformed cells and primary cardiomyocytes. We tested the hypothesis that overexpression of E2F-1 forces growth-arrested human coronary vascular smooth muscle cells (VSMCs) to enter the S phase, undergo apoptosis, and thereby regulate VSMC growth.
Methods and
Results—Early-passage (
5 passages) coronary
VSMCs were transduced at an MOI of 100 with a recombinant adenovirus
encoding human E2F-1. E2F-1 expression was observed by
immunohistochemistry as early as 6 to 8 hours after exposure of the
VSMCs to Ad.E2F-1 but not to the control vector Ad.RR. When cells were
kept in growth-arrest medium, 40% of Ad.E2F-1–treated VSMCs entered
the S phase by 96 hours, whereas the percentage remained <5% in
Ad.RR-treated cells. Transition to the S phase in the E2F-1–transduced
VSMCs was followed by apoptosis, as reflected by chromatin
condensation, membrane blebbing, cell detachment, and loss of
mitochondrial membrane integrity. E2F-1 overexpression resulted in
positive dUTP nick end-labeling mediated by terminal deoxynucleotidyl
transferase, associated with a robust increase in caspase 3–like
activity. Four days after infection with Ad.E2F-1, the fraction of
hypodiploid VSMCs in subG1 increased to 75%. At
7 days, gene transfer of E2F-1 had completely suppressed the growth of
VSMCs, whereas the number of Ad.RR-infected cells had increased >8
times.
Conclusions—Overexpression of the transcription factor E2F-1 regulates growth of human coronary VSMCs by forcing the cells to enter the S phase and then to die. Cell death appears to involve caspase 3–like activity, which, in the VSMCs, is markedly increased by overexpression of E2F-1.
Key Words: cells • genes • muscle, smooth • apoptosis • restenosis
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