Activated forkhead transcription factor inhibits neointimal hyperplasia after angioplasty through induction of p27.
OBJECTIVE
We examined the effects of FKHRL1 (forkhead transcription factor in rhabdomyosarcoma like-1) overexpression on vascular smooth muscle cell (VSMC) proliferation, apoptosis, and cell cycle, in vitro, and the role of FKHRL1 and p27 in the pathophysiology of neointimal growth after balloon angioplasty, in vivo. Furthermore, we tested whether FKHRL1 overexpression can inhibit neointimal hyperplasia in a rat carotid artery model.


RESULTS
Adenovirus expressing the constitutively active FKHRL1 (FKHRL1-TM; triple mutant) with 3 Akt phosphorylation sites mutated was transfected to subconfluent VSMCs. FKHRL1 overexpression in cultured VSMCs increased p27 expression, leading to G1 phase cell-cycle arrest and increased apoptosis. In vivo, the phosphorylation of FKHRL1 increased significantly 3 hours after balloon injury and decreased thereafter, with the subsequent downregulation of p27. Although the phosphorylation of FKHRL1 was greatest at 3 hours, the downregulation of p27 showed a temporal delay, only slightly starting to decrease after 3 hours and reaching a nadir at 72 hours after balloon injury. Gene transfer of FKHRL1-TM increased p27, decreased proliferation, and increased apoptosis of VSMCs, which resulted in a marked reduction in neointima formation (intima-to-media ratio: 0.31+/-0.13 versus 1.17+/-0.28, for FKHRL1-TM versus Adv-GFP; P<0.001).


CONCLUSIONS
Balloon angioplasty leads to the phosphorylation of FKHRL1 and decreased expression of p27, thereby promoting a proliferative phenotype in VSMCs in vitro and in vivo. This study reveals the importance of FKHRL1 in proliferation and viability of VSMCs and suggests that it may serve as a molecular target for interventions to reduce neointima formation after angioplasty.
Park
Kyung-Woo
KW
Cardiovascular Laboratory, Clinical Research Institute, Seoul National University Hospital, Korea.
Kim
Dae-Hee
DH
You
Hyun-Jung
HJ
Sir
Jung-Ju
JJ
Jeon
Soo-In
SI
Youn
Seock-Won
SW
Yang
Han-Mo
HM
Skurk
Carsten
C
Park
Young-Bae
YB
Walsh
Kenneth
K
Kim
Hyo-Soo
HS
D000800:Angioplasty, Balloon / Q000009:adverse effects*; D000818:Animals; D001013:Aorta, Thoracic / Q000293:injuries / Q000473:pathology / Q000502:physiology; D017209:Apoptosis / Q000502:physiology; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002470:Cell Survival / Q000502:physiology; D002478:Cells, Cultured; D050760:Cyclin-Dependent Kinase Inhibitor p27; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D000071161:Forkhead Box Protein O1; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors; D015870:Gene Expression; D018014:Gene Transfer Techniques; D006801:Humans; D006965:Hyperplasia; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000293:injuries* / Q000473:pathology* / Q000502:physiology; D009419:Nerve Tissue Proteins; D010766:Phosphorylation; D051381:Rats; D017207:Rats, Sprague-Dawley; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism*; D025521:Tumor Suppressor Proteins / Q000378:metabolism*; D017539:Tunica Intima / Q000473:pathology
C493962:Cdkn1b protein, rat; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C085126:FOXO1 protein, human; C499517:FOXO3 protein, human; C499515:FOXO3 protein, rat; D000071161:Forkhead Box Protein O1; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors; D009419:Nerve Tissue Proteins; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C074639:Foxo1 protein, rat; D050760:Cyclin-Dependent Kinase Inhibitor p27