Induction of Nitric Oxide Synthase Activity by Cytokines in Ventricular Myocytes Is Necessary but Not Sufficient to Decrease Contractile Responsiveness to β-Adrenergic Agonists

Abstract

Recent evidence has documented that increased activity of an inducible nitric oxide synthase (iNOS; type 2 NO synthase) in primary isolates of adult rat ventricular myocytes after exposure to soluble mediators in medium conditioned by lipopolysaccharide-activated macrophages is associated with a decrease in their contractile responsiveness to β-adrenergic agonists. It remained unclear which specific inflammatory cytokines in this medium contribute to the induction of iNOS activity in myocytes and whether induction of iNOS would result in an obligatory decline in contractile function. Interleukin (IL)-1β and tumor necrosis factor-α (TNF-α) were both present in the lipopolysaccharide-activated macrophage-conditioned medium. However, only IL-1 receptor antagonist and not an anti–rat TNF-α antiserum diminished the extent of iNOS induction in myocytes exposed to this medium and prevented a decline in contractile responsiveness to isoproterenol. When recombinant cytokines were used, IL-1β, TNF-α, and IFN-γ each induced iNOS activity in cardiac myocytes at 24 hours. However, only the combination of IL-1β and IFN-γ reproducibly caused contractile dysfunction in cardiac myocytes. Among the constituents of the defined medium routinely used for maintenance of adult rat ventricular myocytes in primary culture, it was noted that insulin (10⁻⁷ mol/L) was required for NO production, as detected by nitrite release in cytokine-pretreated myocytes, although insulin had no effect on the extent of induction of iNOS mRNA or maximal enzyme activity in myocyte cell lysates. Insulin was also required for the decrease in contractile responsiveness to isoproterenol to be manifest. Thus, induction of iNOS is necessary but not sufficient to cause inflammatory cytokine-induced contractile dysfunction in cardiac myocytes.