Prevention of ischemically induced neointimal hyperplasia using ex vivo antisense oligodeoxynucleotides.
BACKGROUND
Chronic graft vascular disease in cardiac allografts results from coronary artery neointimal formation. Vascular ischemia has been shown to provoke the development of neointimal hyperplasia through endothelial damage. We used a rodent model of neointimal formation to test the in vivo effects of antisense oligodeoxynucleotides (AS ODN) specifically designed to block this process.


METHODS
Aortas from ACI rats were mock transfected or transfected with 18 base pair AS ODNs against the 5' start codon region of both CDC2 kinase, and proliferating cell nuclear antigen (PCNA) mRNA. Transfection was accomplished by placing the aorta in ODN solution (transfected group, 40 micromol/L of each sequence) or saline solution alone (mock-transfected group) and exposing to hydrostatic pressure (2 atm) for 24 hours at 4 degrees C. Vessels were then interposition-grafted into the abdominal aorta of untreated isogenic recipients and procured on postoperative days (POD) 1, 6, and 14.


RESULTS
Nuclear localization of fluorescein isothiocyanate ODN was observed in 81%+/-3% of medial smooth muscle cells at 24 hours after interposition grafting and reperfusion. Efficacy of AS ODNs at blocking CDC2 kinase and PCNA was verified on POD 6 by enzyme-linked immunosorbent assay. This blockade significantly reduced ischemically induced vascular narrowing on POD 14 as assessed by use of computerized image analysis (3.85%+/-2.45% luminal narrowing for transfected vs 7.11%+/-2.03% for control subjects, p < 0.03).


CONCLUSIONS
Our data show the efficacy of AS ODN at blocking rat PCNA and CDC2 kinase up-regulation provoked by ischemia. This ex vivo approach had beneficial effects against vascular narrowing in a rodent model of ischemically induced neointimal hyperplasia, an antigen-independent factor important in the development of subsequent chronic graft vascular disease.
Poston
R S
RS
Department of Cardiothoracic Surgery, Stanford University School of Medicine, Calif, USA.
Tran
K P
KP
Mann
M J
MJ
Hoyt
E G
EG
Dzau
V J
VJ
Robbins
R C
RC
D000818:Animals; D001012:Aorta, Abdominal / Q000637:transplantation*; D001018:Aortic Diseases / Q000517:prevention & control*; D016203:CDC2 Protein Kinase / Q000235:genetics; D002908:Chronic Disease; D003062:Codon / Q000235:genetics; D003327:Coronary Disease / Q000209:etiology; D004730:Endothelium, Vascular / Q000473:pathology; D016027:Heart Transplantation / Q000009:adverse effects; D006874:Hydrostatic Pressure; D006965:Hyperplasia; D007091:Image Processing, Computer-Assisted; D007511:Ischemia / Q000503:physiopathology; D008297:Male; D016376:Oligonucleotides, Antisense / Q000627:therapeutic use*; D018809:Proliferating Cell Nuclear Antigen / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D011912:Rats, Inbred ACI; D014162:Transfection; D014185:Transplantation, Isogeneic; D017539:Tunica Intima / Q000473:pathology*; D015854:Up-Regulation / Q000187:drug effects
D003062:Codon; D016376:Oligonucleotides, Antisense; D018809:Proliferating Cell Nuclear Antigen; D012333:RNA, Messenger; D016203:CDC2 Protein Kinase