title: Growth suppression of human coronary vascular smooth muscle cells by gene transfer of the transcription factor E2F-1.
abstract: 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.
  
  
  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 subG(1) 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.
authors.affiliation: Wafic Said Molecular Cardiology and Gene Therapy Research Laboratory, Texas Heart Institute, Houston, TX 77030, USA.
authors.forename: H S
authors.initials: HS
authors.lastname: Shelat
authors.forename: T J
authors.initials: TJ
authors.lastname: Liu
authors.forename: D L
authors.initials: DL
authors.lastname: Hickman-Bick
authors.forename: M K
authors.initials: MK
authors.lastname: Barnhart
authors.forename: T
authors.initials: T
authors.lastname: Vida
authors.forename: P M
authors.initials: PM
authors.lastname: Dillard
authors.forename: J T
authors.initials: JT
authors.lastname: Willerson
authors.forename: P
authors.initials: P
authors.lastname: Zoldhelyi
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D017209:Apoptosis; D002352:Carrier Proteins; D053148:Caspase 3; D020169:Caspases / Q000096:biosynthesis; D002452:Cell Count; D018797:Cell Cycle Proteins; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000166:cytology / Q000254:growth & development*; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; D004790:Enzyme Induction; D005434:Flow Cytometry; D005822:Genetic Vectors; D006801:Humans; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D024510:Muscle Development; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000254:growth & development*; D056465:Retinoblastoma-Binding Protein 1; D016196:S Phase; D013997:Time Factors; D050680:Transcription Factor DP1; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics* / Q000494:pharmacology; D014162:Transfection
chemical_list: D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; D056465:Retinoblastoma-Binding Protein 1; D050680:Transcription Factor DP1; D014157:Transcription Factors; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases