title: Cytokines and lipopolysaccharides induce inducible nitric oxide synthase but not enzyme activity in adult rat cardiomyocytes.
abstract: There is evidence that nitric oxide (NO) formation in adult cardiomyocytes stimulated with lipopolysaccharide (LPS) is not commensurate with iNOS levels. Tetrahydrobiopterin (BH(4)) is a key factor in the stabilization and NO production by iNOS homodimer. Thus we hypothesized that BH(4) is a limiting factor for NO production in adult cardiomyocytes in response to LPS and cytokines (TNF-alpha, IL-1, IFN-gamma alone, or mixed). It was verified that LPS and cytokines induced iNOS expression which did not translate into increased nitrite or [(14)C]citrulline production. This response coincided with defective BH(4) synthesis and low GTP cyclohydrolase activity. Furthermore, supplementation with BH(4) and ascorbate failed to increase iNOS activity. This effect was related to preferential accumulation of BH(2) rather than BH(4) in these cells. Uncoupled iNOS activity in stimulated cells was examined using mitochondrial aconitase activity as an endogenous marker of superoxide anion radical (O(2)(-)) formation, and found not to be significantly inhibited. 2-Hydroxyethidium also was not significantly increased. We conclude that adult cardiomyocytes are an unlikely source of NO and O(2)(-) in inflammatory conditions. This finding adds a new and unexpected layer of complexity to our understanding of the responses of the adult heart to inflammation.
authors.affiliation: Department of Biophysics, Medical College of Wisconsin, Milwaukee, WI 53226, USA. jvvivar@mcw.edu
authors.forename: Jeannette
authors.initials: J
authors.lastname: Vásquez-Vivar
authors.forename: Jennifer
authors.initials: J
authors.lastname: Whitsett
authors.forename: Irina
authors.initials: I
authors.lastname: Ionova
authors.forename: Eugene
authors.initials: E
authors.lastname: Konorev
authors.forename: Jacek
authors.initials: J
authors.lastname: Zielonka
authors.forename: Balaraman
authors.initials: B
authors.lastname: Kalyanaraman
authors.forename: Yang
authors.initials: Y
authors.lastname: Shi
authors.forename: Galen M
authors.initials: GM
authors.lastname: Pieper
mesh_terms: D000818:Animals; D001708:Biopterins / Q000031:analogs & derivatives / Q000378:metabolism; D015153:Blotting, Western; D016207:Cytokines / Q000378:metabolism*; D015870:Gene Expression; D008070:Lipopolysaccharides / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D016207:Cytokines; D008070:Lipopolysaccharides; D017382:Reactive Oxygen Species; D001708:Biopterins; D009569:Nitric Oxide; D052247:Nitric Oxide Synthase Type II; C003402:sapropterin