Lentivirus-mediated RNAi targeting CREB binding protein attenuates neointimal formation and promotes re-endothelialization in balloon injured rat carotid artery.
OBJECTIVE
Lentiviral vectors provide a promising strategy for the treatment of cardiovascular diseases, owing to their ability to govern efficient and durable gene transfer. However, relatively few studies have been addressed on restenosis after balloon or stent associated arterial injury. We previously found that CREB binding protein (CBP), a powerful transcriptional coactivator, regulated cell proliferation and apoptosis in vascular endothelial and smooth muscle cells. Therefore, we investigated whether inhibition of CBP by lentivirus-mediated small interfering RNA can reduce neointimal formation after arterial injury.


METHODS
The carotid arteries from Sprague-Dawley rats were injured by balloon catheter, followed by incubating with 100 microl lentivirus expressing CBP or negative control (NC)-specific short hairpin RNAs (shRNAs) or PBS solution for 30 minutes. The rats were euthanized for real-time PCR, Western blot, immunohistochemical staining, and morphometric analysis at 4 weeks after balloon injury and in vivo gene transfer.


RESULTS
Lentiviral shRNA targeting CBP markedly reduced CBP expression. Moreover, CBP siRNA showed potent inhibition on balloon injury-induced Nuclear factor kappaB (NF-kappaB) acetylation. Compared with controls, the significant decrease of neointimal formation by CBP siRNA was accompanied by reduced cell proliferation in the neointima of injured arteries. However, no changes in medial area were observed among these different groups. Interestingly, endothelial cell marker CD31 immunostaining and morphometric analysis both showed that CBP knockdown significantly accelerated re-endothelialization.


CONCLUSIONS
These findings suggest that CBP is involved in the control of neointimal formation and re-endothelialization via regulating NF-kappaB acetylation. Lentivirus-mediated CBP silencing may represent a novel therapeutic approach for the prevention of restenosis after vascular interventions.
Yang
Jian
J
Department of Cardiology, Renmin Hospital of Wuhan University, Wuhan, China.
Jiang
Hong
H
Chen
Si-Si
SS
Chen
Jing
J
Li
Wan-Qiang
WQ
Xu
Sheng-Kai
SK
Wang
Ji-Chun
JC
D000107:Acetylation; D000800:Angioplasty, Balloon / Q000009:adverse effects*; D000818:Animals; D050882:CREB-Binding Protein / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D020212:Carotid Artery Injuries / Q000628:therapy*; D049109:Cell Proliferation; D004730:Endothelium, Vascular / Q000254:growth & development* / Q000293:injuries / Q000378:metabolism; D005822:Genetic Vectors / Q000378:metabolism; D016086:Lentivirus / Q000235:genetics; D016328:NF-kappa B / Q000378:metabolism; D058426:Neointima / Q000628:therapy*; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D017539:Tunica Intima / Q000378:metabolism / Q000473:pathology
D016328:NF-kappa B; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D034741:RNA, Small Interfering; D050882:CREB-Binding Protein