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In hepatocytes the regulation of NOS-2 activity at physiological L-arginine levels suggests a close link to the urea cycle.
Nitric Oxide. 2006 Jun; 14(4):300-8.NO

Abstract

High-output synthesis of nitric oxide (NO) by the inducible isoform of NO-synthases (NOS-2) plays an important role in hepatic pathophysiological processes and may contribute to both organ protection and organ destruction during inflammatory reactions. As they compete for the same substrate, L-arginine, an interdependence of NOS-2 and arginase-1 has been repeatedly observed in cells where arginase-1 is cytokine-inducible. However, in hepatocytes, arginases are constitutively expressed and thus, their impact on hepatic NOS-2-derived NO synthesis as well as the influence of L-arginine influx via cationic amino acid transporters during inflammatory reactions are still under debate. Freshly isolated rat hepatocytes were cultured for 24h in the presence of various L-arginine concentrations with or without cytokine addition and nitrite and urea accumulation in culture supernatants was measured. We find that both, cytokine-induced NOS-2 and arginase activities strongly depend on extracellular L-arginine concentrations. When we competed for L-arginine influx via the cationic amino acid transporters by addition of L-lysine, we find a 60-70% inhibition of arginase activity without significant loss of NOS-2 activity. Addition of L-valine, as an arginase inhibitor, leads to a 25% increase in NO formation and an 80-90% decrease in arginase activity. Interestingly, product inhibition of arginase and competitive inhibition of CATs through the addition of L-ornithine leads to a highly significant increase in hepatocytic NOS-2 activity with a concomitant and complete abolishment of its dependence on extracellular L-arginine concentrations. In conclusion, hepatocytic NOS-2 activity shows a surprising pattern of dependence on exogenous L-arginine concentrations. Inhibition and competition experiments suggest a relatively tight link of NOS-2 and urea cycle activities. These data stress the hypothesis of a metabolon-like organization of the urea cycle together with NOS-2 in hepatocytes as excess L-ornithine will be metabolized to l-arginine and thereby increases NO production.

Authors+Show Affiliations

Research Group Immunobiology, Heinrich-Heine-University Düsseldorf, P.O. Box 101007, 40001 Düsseldorf, Germany.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

16410053

Citation

Lerzynski, Guido, et al. "In Hepatocytes the Regulation of NOS-2 Activity at Physiological L-arginine Levels Suggests a Close Link to the Urea Cycle." Nitric Oxide : Biology and Chemistry, vol. 14, no. 4, 2006, pp. 300-8.
Lerzynski G, Suschek CV, Kolb-Bachofen V. In hepatocytes the regulation of NOS-2 activity at physiological L-arginine levels suggests a close link to the urea cycle. Nitric Oxide. 2006;14(4):300-8.
Lerzynski, G., Suschek, C. V., & Kolb-Bachofen, V. (2006). In hepatocytes the regulation of NOS-2 activity at physiological L-arginine levels suggests a close link to the urea cycle. Nitric Oxide : Biology and Chemistry, 14(4), 300-8.
Lerzynski G, Suschek CV, Kolb-Bachofen V. In Hepatocytes the Regulation of NOS-2 Activity at Physiological L-arginine Levels Suggests a Close Link to the Urea Cycle. Nitric Oxide. 2006;14(4):300-8. PubMed PMID: 16410053.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - In hepatocytes the regulation of NOS-2 activity at physiological L-arginine levels suggests a close link to the urea cycle. AU - Lerzynski,Guido, AU - Suschek,Christoph V, AU - Kolb-Bachofen,Victoria, Y1 - 2006/01/10/ PY - 2005/06/17/received PY - 2005/10/14/revised PY - 2005/11/29/accepted PY - 2006/1/18/pubmed PY - 2006/8/11/medline PY - 2006/1/18/entrez SP - 300 EP - 8 JF - Nitric oxide : biology and chemistry JO - Nitric Oxide VL - 14 IS - 4 N2 - High-output synthesis of nitric oxide (NO) by the inducible isoform of NO-synthases (NOS-2) plays an important role in hepatic pathophysiological processes and may contribute to both organ protection and organ destruction during inflammatory reactions. As they compete for the same substrate, L-arginine, an interdependence of NOS-2 and arginase-1 has been repeatedly observed in cells where arginase-1 is cytokine-inducible. However, in hepatocytes, arginases are constitutively expressed and thus, their impact on hepatic NOS-2-derived NO synthesis as well as the influence of L-arginine influx via cationic amino acid transporters during inflammatory reactions are still under debate. Freshly isolated rat hepatocytes were cultured for 24h in the presence of various L-arginine concentrations with or without cytokine addition and nitrite and urea accumulation in culture supernatants was measured. We find that both, cytokine-induced NOS-2 and arginase activities strongly depend on extracellular L-arginine concentrations. When we competed for L-arginine influx via the cationic amino acid transporters by addition of L-lysine, we find a 60-70% inhibition of arginase activity without significant loss of NOS-2 activity. Addition of L-valine, as an arginase inhibitor, leads to a 25% increase in NO formation and an 80-90% decrease in arginase activity. Interestingly, product inhibition of arginase and competitive inhibition of CATs through the addition of L-ornithine leads to a highly significant increase in hepatocytic NOS-2 activity with a concomitant and complete abolishment of its dependence on extracellular L-arginine concentrations. In conclusion, hepatocytic NOS-2 activity shows a surprising pattern of dependence on exogenous L-arginine concentrations. Inhibition and competition experiments suggest a relatively tight link of NOS-2 and urea cycle activities. These data stress the hypothesis of a metabolon-like organization of the urea cycle together with NOS-2 in hepatocytes as excess L-ornithine will be metabolized to l-arginine and thereby increases NO production. SN - 1089-8603 UR - https://www.unboundmedicine.com/medline/citation/16410053/In_hepatocytes_the_regulation_of_NOS_2_activity_at_physiological_L_arginine_levels_suggests_a_close_link_to_the_urea_cycle_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1089-8603(05)00177-1 DB - PRIME DP - Unbound Medicine ER -