Differential expression and localization of nitric oxide synthases in cirrhotic livers of bile duct-ligated rats.
Increased vascular nitric oxide (NO) production has been implicated in the pathogenesis of the hyperdynamic circulation in liver cirrhosis. This study investigated the expression of three isoforms of NO synthase (NOS) in rat cirrhotic livers. Cirrhosis was induced by chronic bile duct ligation (BDL). NOS enzyme activity was assessed by L-citrulline generation. Competitive RT-PCR was performed to detect the mRNA levels of NOS. In situ hybridization was done to localize NOS mRNA. Protein expression of NOS was evaluated by Western blotting and immunohistochemistry. The L-citrulline assay showed that constitutive NOS (cNOS) enzymatic activity was decreased, while inducible NOS (iNOS) activity was increased in BDL livers. Both endothelial NOS (eNOS) and neuronal NOS (nNOS) mRNA were detected in BDL and sham rats, but with enhanced expression in BDL rats. eNOS protein was redistributed with less expression in sinusoidal endothelial cells, but the total levels in liver were not changed. nNOS was induced in hepatocytes of BDL rats, in contrast to only a weak signal observed around some blood vessels in sham livers. Intense mRNA and protein expression of iNOS was induced in livers of BDL rats and was localized in hepatocytes, with no or a negligible amount in control livers. In conclusion, iNOS was induced in cirrhotic liver with its activity increased. In contrast, cNOS activity was impaired, regardless of unchanged eNOS protein levels and enhanced nNOS expression. These results suggest that all three types of NOS have a role in cirrhosis, but their expression and regulation are different.
Wei
Chang-Li
CL
Department of Biochemistry, Faculty of Medicine, National University of Singapore, 10 Kent Ridge Crescent, Singapore 119260, Singapore.
Khoo
Hoon-Eng
HE
Lee
Kang-Hoe
KH
Hon
Wei-Min
WM
D000818:Animals; D001652:Bile Ducts / Q000502:physiology*; D015153:Blotting, Western; D015971:Gene Expression Regulation, Enzymologic; D007150:Immunohistochemistry; D008099:Liver / Q000201:enzymology / Q000378:metabolism; D008106:Liver Cirrhosis, Experimental / Q000139:chemically induced* / Q000201:enzymology* / Q000235:genetics; D008297:Male; D019001:Nitric Oxide Synthase / Q000032:analysis* / Q000235:genetics* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
D012333:RNA, Messenger; D019001:Nitric Oxide Synthase