Exogenous in vivo NO-donor treatment preserves p53 levels and protects vascular cells from apoptosis.
OBJECTIVE
Nitric oxide (NO) is critical in cardiovascular protection. However, NO production is impaired in atherosclerosis resulting in thrombosis, vasoconstriction, and restenosis. Exogenous NO-donors (NOd) have proven protection against ischemia and thrombosis. However, their effect on vascular cell function remains unknown. Our objective was to determine the effect of NOd on vascular smooth muscle cell (VSMC) survival.


METHODS
Pigs (N=16) were randomly distributed in the following treatment groups: (I) LA419 (nitratethiol; 10-day p.o. 0.9 mg/kg bid); (II) LA816 (nitrosothiol; 2h i.v. infusion, 6.6 nmol/(kg min)); (III) nitroglycerine (GTN, 2h i.v. infusion, 2.5mg/kg); and (IV) placebo-control. After sacrifice, pig aortic arch explants were either frozen or incubated with 20% homologous pig serum for different time periods (18h, 2 and 10 days). Bcl2/bax ratio, phosphorylated-p53, cyclin-D2, casp-3, casp-8, and bax mRNA levels and protein expression were evaluated by real-time PCR and western blot, respectively.


RESULTS
Within the first 2 days explants from NOd-treated animals had an increased pro-survival Bcl2/bax ratio compared with placebo-control (5x higher in LA816 and GTN i.v.-treated animals and 10x higher in 10-day orally treated LA419 animals; P<0.05). Phosphorylated-P53-protein levels were consistently increased by all NOd-treatments vs. placebo-control (P<0.05) and p53 mRNA levels showed an increase in placebo-controls at day 10 while they did not rise in NOd-treated animals (P<0.05). Cyclin-D2, casp-3, and bax mRNA content were 6x lower in NOd-treated animals than in control explants at day-10 (P<0.001). No changes were detected on caspase-8.


CONCLUSIONS
Taken together, our findings suggest that exogenous in vivo NO treatment seems to preserve VSMC from mitochondrial-dependent apoptosis and drive cells to quiescence through p53 increase.
Duran
Xavier
X
Cardiovascular Research Center, CSIC-ICCC, Barcelona, Spain.
Vilahur
Gemma
G
Badimon
Lina
L
D000818:Animals; D001011:Aorta / Q000473:pathology*; D017209:Apoptosis; D053181:Caspase 8 / Q000378:metabolism; D002470:Cell Survival; D007511:Ischemia / Q000473:pathology; D009131:Muscle, Smooth, Vascular / Q000473:pathology; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000737:chemistry*; D010919:Placebos; D012333:RNA, Messenger / Q000378:metabolism; D013552:Swine; D013927:Thrombosis / Q000473:pathology; D013997:Time Factors; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
D020030:Nitric Oxide Donors; D010919:Placebos; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; D009569:Nitric Oxide; D053181:Caspase 8