Forkhead transcription factor FOXO3a is a negative regulator of angiogenic immediate early gene CYR61, leading to inhibition of vascular smooth muscle cell proliferation and neointimal hyperplasia.
Cysteine-rich angiogenic protein 61 (CYR61, CCN1) is an immediate early gene expressed in vascular smooth muscle cells (VSMCs) on growth factor stimulation, and its expression has been suggested to be associated with postangioplasty restenosis. The forkhead transcription factors are reported to play various roles in cellular proliferation, apoptosis, and even adaptation to cellular stress. We hypothesized that the forkhead transcription factor FOXO3a may regulate CYR61 expression in VSMCs and investigated the CYR61-modulating effect of FOXO3a in the process of vascular response to vasoactive signals and vascular injury. To evaluate the effect of FOXO3a on CYR61 expression, rat VSMCs were infected with adenoviral vectors expressing constitutively active FOXO3a (Ad-TM-FOXO3a). Constitutively active FOXO3a gene transduction suppressed CYR61 expression. Luciferase assay with the deletion constructs of the forkhead factor binding motif in CYR61 promoter region, which resulted in a significant decrease in luciferase expression compared with the intact construct, and chromatin immunoprecipitation analysis confirmed transcriptional regulation of CYR61 by FOXO3a. Serum and angiotensin II rapidly induced CYR61 expression, which was significantly reduced by Ad-TM-FOXO3a. Reduction of VSMC proliferation and migration associated with FOXO3a activation was significantly reversed by cotransfection of adenoviral vector expressing CYR61, whereas apoptosis induction by FOXO3a was not influenced. In a rat balloon carotid arterial injury model, CYR61 was rapidly induced in VSMCs in the early stage of injury and remained elevated until 14 days, which was suppressed by Ad-TM-FOXO3a transfection. After 14 days, there was a significant reduction in neointima by FOXO3a transduction compared with the control group (0.06+/-0.02 versus 0.20+/-0.07 mm(2), P<0.01). Such reduction of neointimal hyperplasia by Ad-TM-FOXO3a was reversed by CYR61 replenishment. These data suggest that FOXO3a is a negative transcription factor of CYR61 and that suppression of CYR61 is among several mechanisms by which FOXO3a inhibits VSMC proliferation and neointimal hyperplasia.
Lee
Hae-Young
HY
National Research Laboratory for Cardiovascular Stem Cell, Seoul National University Hospital, Korea.
Chung
Jae-Woong
JW
Youn
Seock-Won
SW
Kim
Ju-Young
JY
Park
Kyung-Woo
KW
Koo
Bon-Kwon
BK
Oh
Byung-Hee
BH
Park
Young-Bae
YB
Chaqour
Brahim
B
Walsh
Kenneth
K
Kim
Hyo-Soo
HS
D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D001013:Aorta, Thoracic; D017209:Apoptosis / Q000187:drug effects; D050197:Atherosclerosis / Q000378:metabolism / Q000473:pathology; D020212:Carotid Artery Injuries / Q000209:etiology / Q000235:genetics / Q000378:metabolism; D002404:Catheterization / Q000009:adverse effects; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured / Q000187:drug effects; D003470:Culture Media / Q000494:pharmacology; D017077:Culture Media, Conditioned / Q000494:pharmacology; D055515:Cysteine-Rich Protein 61; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors / Q000235:genetics / Q000502:physiology*; D005786:Gene Expression Regulation / Q000502:physiology; D017930:Genes, Reporter; D006801:Humans; D006965:Hyperplasia; D017874:Immediate-Early Proteins / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D036341:Intercellular Signaling Peptides and Proteins / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology; D008297:Male; D051379:Mice; D009131:Muscle, Smooth, Vascular / Q000166:cytology*; D016376:Oligonucleotides, Antisense / Q000235:genetics; D011401:Promoter Regions, Genetic; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000502:physiology; D014158:Transcription, Genetic; D014161:Transduction, Genetic; D017539:Tunica Intima / Q000473:pathology
C501848:CCN1 protein, human; C491850:CCN1 protein, mouse; C501849:CCN1 protein, rat; D003470:Culture Media; D017077:Culture Media, Conditioned; D055515:Cysteine-Rich Protein 61; C499517:FOXO3 protein, human; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D016376:Oligonucleotides, Antisense; D011993:Recombinant Fusion Proteins; D000804:Angiotensin II