Oxidant generation predominates around calcifying foci and enhances progression of aortic valve calcification.
OBJECTIVE
We hypothesized that reactive oxygen species (ROS) contribute to progression of aortic valve (AV) calcification/stenosis.


RESULTS
We investigated ROS production and effects of antioxidants tempol and lipoic acid (LA) in calcification progression in rabbits given 0.5% cholesterol diet +10(4) IU/d Vit.D2 for 12 weeks. Superoxide and H2O2 microfluorotopography and 3-nitrotyrosine immunoreactivity showed increased signals not only in macrophages but preferentially around calcifying foci, in cells expressing osteoblast/osteoclast, but not macrophage markers. Such cells also showed increased expression of NAD(P)H oxidase subunits Nox2, p22phox, and protein disulfide isomerase. Nox4, but not Nox1 mRNA, was increased. Tempol augmented whereas LA decreased H2O2 signals. Importantly, AV calcification, assessed by echocardiography and histomorphometry, decreased 43% to 70% with LA, but increased with tempol (P < or = 0.05). Tempol further enhanced apoptosis and Nox4 expression. In human sclerotic or stenotic AV, we found analogous increases in ROS production and NAD(P)H oxidase expression around calcifying foci. An in vitro vascular smooth muscle cell (VSMC) calcification model also exhibited increased, catalase-inhibitable, calcium deposit with tempol, but not with LA.


CONCLUSIONS
Our data provide evidence that ROS, particularly hydrogen peroxide, potentiate AV calcification progression. However, tempol exhibited a paradoxical effect, exacerbating AV/vascular calcification, likely because of its induced increase in peroxide generation.
Liberman
Marcel
M
Vascular Biology Laboratory, Heart Institute, University of São Paulo School of Medicine, Av. Eneas Carvalho Aguiar, 44, Annex II, 9th floor, CEP 05403-000, São Paulo, Brazil.
Bassi
Estêvão
E
Martinatti
Marina Kamla
MK
Lario
Fábio Cerqueira
FC
Wosniak
João
J
Pomerantzeff
Pablo M A
PM
Laurindo
Francisco R M
FR
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D001024:Aortic Valve Stenosis / Q000000981:diagnostic imaging / Q000201:enzymology / Q000473:pathology*; D002114:Calcinosis / Q000473:pathology*; D002791:Cholesterol, Dietary / Q000008:administration & dosage; D004195:Disease Models, Animal; D018450:Disease Progression; D004452:Echocardiography; D005453:Fluorescence; D006801:Humans; D006861:Hydrogen Peroxide / Q000009:adverse effects / Q000378:metabolism*; D007150:Immunohistochemistry; D019255:NADPH Oxidases / Q000378:metabolism; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D011336:Probability; D011817:Rabbits; D011897:Random Allocation; D017382:Reactive Oxygen Species / Q000009:adverse effects / Q000378:metabolism*; D012016:Reference Values; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D046509:Tissue Culture Techniques
D000975:Antioxidants; D002791:Cholesterol, Dietary; D017382:Reactive Oxygen Species; D006861:Hydrogen Peroxide; D019255:NADPH Oxidases