Impaired endothelial nitric oxide synthase activity associated with enhanced caveolin binding in experimental cirrhosis in the rat.
OBJECTIVE
A reduction in nitric oxide (NO) has been implicated as a cause of intrahepatic vasoconstriction in cirrhosis, but the regulatory mechanisms remain undefined. The aim of this study was to examine a contributory role for caveolin-1, a putative negative regulator of endothelial NO synthase, in mediating deficient intrahepatic NO production in the intact cirrhotic liver.


METHODS
Cirrhosis was induced by carbon tetrachloride inhalation. Flow regulation of NO production and perfusion pressure was examined in the perfused rat liver. Protein expression of endothelial NO synthase (eNOS), caveolin, and calmodulin was examined by Western blotting and immunohistochemistry. NOS activity and NO production were assessed by citrulline generation and chemiluminescence, respectively. Protein-protein interactions were examined using whole tissue protein immunoprecipitation.


RESULTS
In response to incremental increases in flow, cirrhotic animals produced significantly less NO(x) than control animals. NOS activity was significantly reduced in liver tissue from cirrhotic animals compared with control animals in the presence of similar eNOS protein levels. Deficient eNOS activity was associated with a severalfold increase in binding of eNOS with caveolin. Protein levels of caveolin-1 were markedly increased in the cirrhotic liver.


CONCLUSIONS
These studies provide evidence that enhanced expression and interaction of caveolin with eNOS contribute to impaired NO production, reduced NOS activity, and vasoconstriction in the intact cirrhotic liver.
Shah
V
V
Department of Medicine, Yale University School of Medicine, New Haven, Connecticut, USA.
Toruner
M
M
Haddad
F
F
Cadelina
G
G
Papapetropoulos
A
A
Choo
K
K
Sessa
W C
WC
Groszmann
R J
RJ
D000818:Animals; D002147:Calmodulin / Q000378:metabolism; D051242:Caveolin 1; D022461:Caveolins; D066298:In Vitro Techniques; D008099:Liver / Q000201:enzymology; D008102:Liver Circulation / Q000502:physiology; D008106:Liver Cirrhosis, Experimental / Q000378:metabolism*; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III; D010477:Perfusion; D011312:Pressure; D051381:Rats; D017207:Rats, Sprague-Dawley; D012016:Reference Values; D014018:Tissue Distribution
D002147:Calmodulin; C495537:Cav1 protein, rat; D051242:Caveolin 1; D022461:Caveolins; D008565:Membrane Proteins; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D052250:Nitric Oxide Synthase Type III; C496316:Nos3 protein, rat