Interactions between cytokines and neurohormonal systems in the failing heart.

The prognosis for patients with congestive heart failure (CHF) has been improved as a result of the use of angiotensin converting enzyme inhibitors and beta-adrenergic receptor blockers. The success of these therapies underscores the pathogenic role of neurohormonal activation in CHF. Clinical and experimental evidence supports a pathophysiologic role for pro-inflammatory cytokines and nitric oxide (NO) in the effects of angiotensin II and norepinephrine in CHF. Potential mechanism(s) responsible for the effects of these immunomodulators can be explained on the basis of established principles of myocardial excitation contraction coupling (E-C). A novel hypothesis is proposed that cytokines and NO-mediated alterations in E-C coupling contribute to the reversible myocardial depression and beta-adrenergic desensitization observed in a diverse group of clinical conditions that activate host inflammatory responses, including CHF. Basic studies into cytokine signaling pathways in cardiac myocytes have the potential to provide important new insights relevant to the design of new management strategies for the treatment of congestive heart failure patients.

HIV gp120 enhances NO production by cardiac myocytes through p38 MAP kinase-mediated NF-kappaB activation.

Human immunodeficiency virus (HIV) infection is associated with a surprisingly high frequency of myocardial dysfunction. Potential mechanisms include direct effects of HIV, indirect effects mediated by cytokines, or a combination. We have previously reported that interleukin-1beta (IL-1beta) (500 U/ml) alone induced nitric oxide (NO) production by neonatal rat cardiac myocytes (CM). Effects of the HIV-1 envelope, glycoprotein120 (gp120), on inducible NO synthase (iNOS) in CM have not been previously reported. Unlike IL-1beta, recombinant HIV-gp120 (1 microgram/ml) alone failed to enhance NO production in CM (0.5 +/- 0.4 vs. 0.4 +/- 0.5 micromol/1.25 x 10(5) cells/48 h, gp120 vs. control, respectively; n = 12, P = not significant). However, the addition of gp120 to IL-1beta significantly enhanced iNOS mRNA expression (70 +/- 1.5 vs. 26 +/- 2.4 optical units, IL-1beta + gp120 vs. IL-1beta, respectively; n = 3), iNOS protein synthesis (42 +/- 1.4 vs. 18 +/- 0.8 optical units, IL-1beta + gp120 vs. IL-1beta, respectively; n = 3), and NO production (NO(2)(-)) (6.6 +/- 0.6 vs. 4.1 +/- 0.8 micromol/1.25 x 10(5) cells/48 h, IL-1beta + gp120 vs. IL-1beta, respectively; n = 12, P

TNF-alpha enhances cardiac myocyte NO production through MAP kinase-mediated NF-kappaB activation.

We have previously reported that interleukin-1beta (IL-1beta) alone induced nitric oxide (NO) production by neonatal rat cardiac myocytes (CM). The effects of tumor necrosis factor-alpha (TNF-alpha) on inducible NO synthase (iNOS) were not characterized. Unlike IL-1beta, TNF-alpha alone failed to enhance NO production in CM. However, the addition of TNF-alpha to IL-1beta significantly enhanced iNOS mRNA expression, iNOS protein synthesis, and NO production (NO(-)(2)). TNF-alpha enhancement of IL-1beta-induced NO(-)(2) production was blocked by PD-98059, a selective mitogen-activated protein (MAP) kinase kinase inhibitor, but not calphostin C (Cal C), a protein kinase C inhibitor. TNF-alpha-enhanced MAP kinase activity was associated with an increase in IL-1beta-stimulated NF-kappaB activity. PD-98059, but not Cal C, inhibited both TNF-alpha-enhanced MAP kinase and NF-kappaB activities. Thus TNF-alpha enhancement of IL-1beta-induced NO production is associated with MAP kinase-mediated activation of NF-kappaB.

Norepinephrine-stimulated MAP kinase activity enhances cytokine-induced NO production by rat cardiac myocytes.

The effect of norepinephrine (NE) on cytokine-stimulated nitric oxide (NO) production by cardiac myocytes has not been previously reported. NE alone caused no significant increase in NO-2 levels over vehicle. Addition of NE to interleukin-1beta (IL-1beta) significantly increased inducible NO synthase (iNOS) mRNA expression, iNOS protein, and NO-2 production vs. IL-1beta alone. Addition of the alpha-adrenergic blocker prazosin or the beta-adrenergic blocker propranolol partially reduced the NE-mediated increase in iNOS mRNA expression and NO-2 production. Addition of prazosin and propranolol together completely abolished the NE-induced increase in iNOS mRNA expression and NO-2 production. NE significantly enhanced mitogen-activated protein (MAP) kinase activity that was reduced by prazosin, propranolol, and PD-98059, a selective MAP kinase kinase inhibitor. Addition of PD-98059 reduced the NE-mediated increase in iNOS mRNA expression and NO-2 production. We report for the first time that NE enhances IL-1beta-stimulated NO production by activation of alpha- and beta-adrenergic receptors through a novel MAP kinase mechanism.

Effects of cytokines and nitric oxide on myocardial E-C coupling.

Compelling evidence now exists that pro-inflammatory cytokines and nitric oxide (NO) are newly identified endogenous regulators of myocardial contractility. The mechanism(s) responsible for the inotropic and chronotropic effects of these novel mediators can be explained on the basis of recently established principles of myocardial excitation contraction coupling (E-C). A novel hypothesis is proposed that cytokines and NO-mediated alterations in E-C coupling contribute to the reversible myocardial depression and beta-adrenergic desensitization observed in a diverse group of clinical conditions that activate host inflammatory responses, including congestive heart failure. The results of in vitro studies indicate that cytokines and NO have both immediate, short-term, as well as long-term effects on cardiac performance. Basic studies into these cytokine signaling pathways in cardiac myocytes have the potential to provide important new insights relevant to the design of new management strategies for the treatment of congestive heart failure patients.

Comparison of paroxetine and nortriptyline in depressed patients with ischemic heart disease.

CONTEXT: Depression and ischemic heart disease often are comorbid conditions and, in patients who have had a myocardial infarction, the presence of depression is associated with increased mortality. Patients with heart disease need a safe and effective treatment for depression. OBJECTIVE: To compare the efficacy, cardiovascular effects, and safety of a specific serotonin reuptake inhibitor, paroxetine, with a tricyclic antidepressant, nortriptyline hydrochloride, in depressed patients with ischemic heart disease. DESIGN: Two-week placebo lead-in followed by a double-blind randomized 6-week medication trial. SETTING: Research clinics in 4 university centers. PATIENTS: Eighty-one outpatients meeting Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition criteria for major depressive disorder and with documented ischemic heart disease. INTERVENTIONS: Treatment with either paroxetine, 20 to 30 mg/d, or nortriptyline targeted to a therapeutic plasma level, 190 to 570 nmol/L (50-150 ng/mL), for 6 weeks. MAIN OUTCOME MEASURES: For effectiveness of treatment, a decline in the score of the Hamilton Rating Scale for Depression by 50% and final score of 8 or less; for cardiovascular safety, heart rate and rhythm, supine and standing systolic and diastolic blood pressures, electrocardiogram conduction intervals, indexes of heart rate variability, and rate of adverse events. RESULTS: By intent-to-treat analysis, 25 (61%) of 41 patients improved during treatment with paroxetine and 22 (55%) of 40 improved with nortriptyline. Neither drug significantly affected blood pressure or conduction intervals. Paroxetine had no sustained effects on heart rate or rhythm or indexes of heart rate variability, whereas patients treated with nortriptyline had a sustained 11% increase in heart rate from a mean of 75 to 83 beats per minute (P<.001) and a reduction in heart rate variability, as measured by the SD of all normal R-R intervals over a 24-hour period, from 112 to 96 (P<.01). Adverse cardiac events occurred in 1 (2%) of 41 patients treated with paroxetine and 7 (18%) of 40 patients treated with nortriptyline (P<.03). CONCLUSIONS: Paroxetine and nortriptyline are effective treatments for depressed patients with ischemic heart disease. Nortriptyline treatment was associated with a significantly higher rate of serious adverse cardiac events compared with paroxetine.

Beta-adrenergic regulation of constitutive nitric oxide synthase in cardiac myocytes.

Nitric oxide (NO) has been implicated in endogenous control of myocardial contractility. However, NO release has not yet been demonstrated in cardiac myocytes. Accordingly, endogenous NO production was measured with a porphyrinic microsensor positioned on the surface of individual neonatal or adult rat ventricular myocytes (n > 6 neonatal and adult cells per experiment). In beating neonatal myocytes, there was no detectable spontaneous NO release with each contraction. However, norepinephrine (NE; 0.25-1 microM) elicited transient NO release from beating neonatal (149 +/- 11 to 767 +/- 83 nM NO) and noncontracting adult (157 +/- 13 to 791 +/- 89 nM NO) cells. NO was released by adrenergic agonists with the following rank order of potency: isoproterenol (beta1beta2) > NE (alpha/beta1) > dobutamine (beta1) approximately epinephrine (alpha/beta1beta2) > tertbutylene (beta2); NO was not released by phenylephrine (alpha). NE-evoked NO release was reversibly blocked by N(G)-monomethyl-L-arginine, trifluoperazine, guanosine 5'-O-(2-thiodiphosphate), and nifedipine but was enhanced by 3-isobutyl-1-methylxanthine (0.5 mM = 14.5 +/- 1.6%) and BAY K 8644 (10 microM = 11.9 +/- 1%). NO was also released by A-23187 (10 microM = 884 +/- 88 nM NO), guanosine 5'-O-(3-thiotriphosphate) (1 microM = 334 +/- 56 nM NO), and dibutyryl adenosine 3',5'-cyclic monophosphate (10-100 microM = 35 +/- 9 to 284 +/- 49 nM NO) but not by ATP, bradykinin, carbachol, 8-bromoguanosine 3',5'-cyclic monophosphate, or shear stress. This first functional demonstration of a constitutive NO synthase in cardiac myocytes suggests its regulation by a beta-adrenergic signaling pathway and may provide a novel mechanism for the coronary artery vasodilatation and enhanced diastolic relaxation observed with adrenergic stimulation.

Elevated platelet factor 4 and beta-thromboglobulin plasma levels in depressed patients with ischemic heart disease.

Clinical depression has recently been recognized as an independent risk factor for cardiac mortality in patients after myocardial infarction. The underlying mechanisms of this increased mortality remain unclear. This study investigated the hypothesis that patients suffering from ischemic heart disease (IHD) and depression concurrently may have abnormal platelet activation resulting in an increased risk of thrombosis. Platelet factor 4 (PF4) and beta-thromboglobulin (beta-TG) were measured in young healthy control subjects, in nondepressed patients with IHD, and in depressed patients with IHD. Mean PF4 and beta-TG plasma levels in the IHD group with depression were found to be significantly higher than those of the control and IHD groups. This increase was not related to age, gender, racial difference, aspirin use, or severity of cardiac disease. This finding suggests that in depressed patients with IHD there is greater platelet activation, and may indicate an increased risk of thrombotic complications.

Cytokines and nitric oxide synthase inhibitor as mediators of adrenergic refractoriness in cardiac myocytes.

We have previously proposed that pro-inflammatory cytokines and nitric oxide (NO) contributed to reversible myocardial depression in patients with sepsis and congestive heart failure. Sepsis and heart failure are also associated with refractoriness to beta-adrenoceptor agonists. Therefore, the chronotropic effects of cytokines and the NO synthase inhibitor, NG-methyl-L-arginine (NMA), on beta-adrenoceptor stimulation of neonatal cardiac myocytes were studied. Tumor necrosis factor alpha, interleukin-1 beta and interleukin-6 but not interleukin-4 or interleukin-5 significantly enhanced spontaneous beating rates compared to untreated myocytes in serum-free media for 48 h (P < 0.01; n = 12 for each). NMA also significantly enhanced spontaneous beating rates (P < 0.01; n = 12 for each). Only interleukin-1 beta treatment resulted in significant nitrite production, immunohistochemical staining for inducible nitric oxide synthase and detection of inducible NO synthase messenger RNA by reverse transcriptase-polymerase chain reaction (RT-PCR). However, tumor necrosis factor alpha, interleukin-1 beta, interleukin-6, and NMA each completely blocked the positive chronotropic effects of the beta-adrenoceptor agonist, isoproterenol (P < 0.01; n = 12 for each). These findings are most consistent with an inducible NO synthase-independent effect of cytokines and NMA on the chronotropic responses of neonatal cardiac myocytes to beta-adrenoceptor stimulation. This effect of cytokines and NMA on adrenergic signaling may involve a myocardial constitutive NO synthase or an NO-independent mechanism.

Glucose and pyruvate regulate cytokine-induced nitric oxide production by cardiac myocytes.

Metabolic requirements for the production of nitric oxide (NO) by cytokine-stimulated neonatal rat cardiac myocytes (CM) were studied. CM were cultured for 48 h in media containing interleukin-1 beta (IL-1 beta) and free fatty acids. Removal of glucose from the media partially inhibited IL-1 beta-stimulated nitrite (NO2-) production [8.1 +/- 0.3 vs. 4.4 +/- 0.6 nmol.(1.25 X 10(5) cells)-1.48 h-1; P < 0.01; n = 12]. The glycolytic inhibitor 2-deoxy-D-glucose (2-DG) completely inhibited IL-1 beta-stimulated NO2- production [0.7 +/- 0.5 nmol.(1.25 X 10(5) cells)-1.48 h-1; P < 0.01; n = 12]. The addition of the glycolytic end product, pyruvate, completely blocked the 2-DG inhibition of IL-1 beta-stimulated NO2- production [7.4 +/- 0.4 nmol.(1.25 X 10(5) cells)-1.48 h-1; P < 0.01; n = 12]. Pyruvate alone did not significantly enhance NO2- production in the presence or absence of glucose (n = 12). The inactive analogue 3-O-methylglucose had no effect on NO2- production (n = 12). Reverse transcription-polymerase chain reaction revealed that pyruvate blocked 2-DG inhibition of inducible NO synthase mRNA expression. Neither 2-DG nor pyruvate had any effect on GTP-cyclohydrolase I mRNA expression in CM. We report for the first time that optimal IL-1 beta-stimulated NO production by CM requires both glucose and the glycolytic end product pyruvate.

Protein kinase A activation is required for IL-1-induced nitric oxide production by cardiac myocytes.

We have previously reported that interleukin-1 beta (IL-1) alone induced the transcription of inducible nitric oxide synthase (iNOS) mRNA and nitric oxide (NO) production by isolated neonatal rat cardiac myocytes (CM). The present studies were undertaken to explore the signal transduction pathways involved in IL-1-induced NO production by CM. The addition of IL-1 to CM resulted in a peak rise in both adenosine 3',5'-cyclic monophosphate (cAMP) and protein kinase A (PKA) activities by 10 min followed by rapid declines and return to basal levels within 60 min. The PKA inhibitor KT-5720 completely blocked NO-2 production by IL-1-stimulated CM (P < 0.01; n = 12). The protein kinase C (PKC) inhibitor, calphostin C, had no effect on NO2- production by IL-1 stimulated CM [P = not significant (NS); n = 12]. The addition of PKA+cAMP to cytosols derived from IL-1-treated CM did not directly enhance iNOS enzyme activity (P = NS; n = 3). CM treated with IL-1 alone stained positively for iNOS protein by immunohistochemistry. iNOS staining was absent in CM treated with IL-1+KT-5720. KT-5720 resulted in an earlier disappearance of iNOS mRNA from IL-1-treated CM, as detected by semiquantitative reverse transcriptase-polymerase chain reaction. We report for the first time that PKA (but not PKC) activation is required for IL-1-induced NO production by CM.

NF-kappa B and GTP cyclohydrolase regulate cytokine-induced nitric oxide production by cardiac myocytes.

We previously reported that interleukin-1 beta (IL-1) alone stimulated nitric oxide (NO) production by neonatal rat cardiac myocytes (CM) in culture. The present studies were undertaken to explore the signal transduction pathways involved in IL-1-induced NO production by CM. Translocation from the cytosol to the nucleus of nuclear factor-kappa B (NF-kappa B) and activation of guanosine 5'-triphosphate (GTP) cyclohydrolase [rate-limiting enzyme in tetrahydrobiopterin (BH4) synthesis] have been implicated in IL-1 signaling. Accordingly, the effects of the NF-kappa B inhibitor pyrolidine dithiocarbamate (PDTC) and the GTP cyclohydrolase inhibitor 2,4-diamino-6-hydroxypyrimidine (DAHP) on IL-1-induced NO production by CM were studied. PDTC and DAHP inhibited IL-1-induced NO2-production by CM (6.7 +/- 0.6 vs. 0.9 +/- 0.3 and 0.3 +/- 0.1 nmol. 1.25 x 10(5) cells(-1).48 h-1, respectively, P < 0.01, n = 12 for each). Immunohistochemical staining revealed that PDTC blocked IL-1-stimulated nuclear translocation of NF-kappa B. The membrane-permeable analogue of the NO synthase cofactor BH4, methyl-BH4 (mBH4), only partially reversed DAHP inhibition of NO2- formation (6.7 +/- 0.6 vs. 2.4 +/- 0.3 nmol. 1.25 x 10(5) cells-1.48 h-1, P < 0.01, n = 12). Semiquantitative reverse transcription polymerase chain reaction revealed no inducible NO synthase (iNOS) mRNA production in cells treated with IL-1 + PDTC.CM treated with IL-1 + DAHP did express iNOS mRNA. We report for the first time that nuclear translocation of NF-kappa B is essential for II-1-induced iNOS mRNA expression and GTP cyclohydrolase activity is required in addition in addition to BH4 for optimal NO production by CM.

Inotropic, chronotropic, and radioligand binding characteristics of leukotriene B4 in cardiac myocyte, papillary muscle, and membrane preparations.

Cardioprotective effects of dietary fish oil consumption have been attributed to the substitution of eicosapentaenoic acid (EPA) for arachidonic acid (AA) in cellular membranes. This substitution blocks the formation of leukotriene B(4) (LTB(4)) which is elaborated locally in the area of ischemic myocardium. We studied the in vitro inotropic, chronotropic, and radioligand binding characteristics of LTB(4) to gain potentially important insights into the mechanisms involved in the cardioprotective effect of EPA. LTB(4) was a concentration-dependent and reversible negative inotrope in isolated papillary muscle preparations. LTB(4) significantly blocked the positive inotropic effect of Bay K 8644 (BK) (calcium channel regulator) on papillary muscles and the positive chronotropic effect of BK on spontaneously beating neonatal cardiac myocytes. LTB(4) had no direct effect on [(3)H]PN200-110 binding to cardiac sarcolemmal L-type calcium channels or [3H]ryanodine binding to cardiac SR calcium release channels. These studies support a potentially important indirect effect of LTB(4) on myocardial E-C coupling.