Inhibition of inducible nitric oxide synthase activity prevents liver recovery in rat thioacetamide-induced fibrosis reversal.
BACKGROUND
Stimulation of nitric oxide (NO) synthesis similar to the application of NO donors could be of benefit in liver fibrosis. Many authors believe that activation of NO synthesis by pharmacological agents is promising in the treatment of liver fibrosis. However, there is considerable controversy in understanding the role of NO in fibrogenesis and fibrolysis. The aims of our study were to evaluate the effects of L-arginine, as an NO metabolic precursor, and those of NO synthase (NOS) inhibitors, L-nitroarginine methyl ester (L-NAME) and aminoguanidine (AG) in rats with thioacetamide (TAA)-induced liver fibrosis reversal.


METHODS
Male Wistar rats, 230-240 g, received TAA (200 mg kg(-1), intraperitoneally) twice a week for 3 months. Liver resolution was simulated by withdrawal of TAA administration. Thereafter the animals were subdivided into five groups and treated by intragastric intubation with: L-arginine (100 and 300 mg kg(-1)); L-NAME as an inhibitor of both constitutively expressed NOS (eNOS) and inducible NOS (iNOS) (20 mg kg(-1)), AG as a specific inhibitor of iNOS (100 mg kg(-1)) or placebo. The severity of liver fibrosis was assessed by morphometric evaluation of liver slides stained with Azan-Mallory, hydroxyproline (Hyp) determination and mRNA steady state levels of collagen I, transforming growth factor (TGF)-beta1, metalloproteinases (MMP)-13, -14, tissue inhibitor of MMP (TIMP)-1 and plasminogen activator inhibitor (PAI)-1 were quantified by real time PCR. The activities of serum marker enzyme, alanine aminotransferase, aspartate aminotransferase and alkaline phosphatase, were measured.


RESULTS
TAA treatment during 3 months induced micronodular liver fibrosis with a pronounced deposition of collagen fibres. L-Arginine did not affect this deposition nor did it affect both relative and total liver hydroxyproline content. Both NOS inhibitors significantly increased the square of the liver connective tissue stained by Azan-Mallory and the above parameters characterizing liver hydroxyproline content. Both NOS inhibitors up-regulated procollagen alpha1 (I), MMP-13, TIMP-1 and PAI-1 mRNA expression. The AG effects were more pronounced. than those of L-NAME. AG treatment also increased mRNA expression of TGF-beta1 and PAI-1.


CONCLUSIONS
Both NOS inhibitors developed a clear pro-fibrotic effect in the liver. Aminoguanidine was more fibrotic than L-NAME. Our data suggest a significant anti-fibrotic role for iNOS rather than for eNOS. L-Arginine did not show any anti-fibrotic properties in the TAA-model used.
Lukivskaya
O
O
National Academy of Sciences, Grodno, Belarus.
Patsenker
E
E
Lis
R
R
Buko
V U
VU
D000818:Animals; D001120:Arginine / Q000494:pharmacology; D004195:Disease Models, Animal; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D008099:Liver / Q000201:enzymology*; D008103:Liver Cirrhosis / Q000139:chemically induced / Q000201:enzymology* / Q000473:pathology; D008115:Liver Regeneration / Q000187:drug effects*; D008297:Male; D052247:Nitric Oxide Synthase Type II / Q000037:antagonists & inhibitors* / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013853:Thioacetamide / Q000494:pharmacology*
D004791:Enzyme Inhibitors; D013853:Thioacetamide; D001120:Arginine; D052247:Nitric Oxide Synthase Type II