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

1996 ◽  
Vol 271 (1) ◽  
pp. C429-C434 ◽  
Author(s):  
C. V. Oddis ◽  
R. L. Simmons ◽  
B. G. Hattler ◽  
M. S. Finkel
Keyword(s):  
Nitric Oxide ◽  
Protein Kinase ◽  
Protein Kinase A ◽  
Cardiac Myocytes ◽  
Interleukin 1 ◽  
Neonatal Rat ◽  
Inos Mrna ◽  
No Production ◽  
Neonatal Rat Cardiac Myocytes ◽  
Kinase A

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.

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

1996 ◽  
Vol 271 (4) ◽  
pp. C1244-C1249 ◽  
Author(s):  
C. V. Oddis ◽  
M. S. Finkel
Keyword(s):  
Nitric Oxide ◽  
Mrna Expression ◽  
Cardiac Myocytes ◽  
Interleukin 1 ◽  
Neonatal Rat ◽  
No Production ◽  
Neonatal Rat Cardiac Myocytes ◽  
The Media ◽  
Glycolytic Inhibitor ◽  
First Time

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.

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

1996 ◽  
Vol 270 (5) ◽  
pp. H1864-H1868 ◽  
Author(s):  
C. V. Oddis ◽  
M. S. Finkel
Keyword(s):  
Nitric Oxide ◽  
Cardiac Myocytes ◽  
Nuclear Translocation ◽  
Interleukin 1 ◽  
No Synthase ◽  
Neonatal Rat ◽  
Inos Mrna ◽  
No Production ◽  
Gtp Cyclohydrolase ◽  
Nf Kappa B

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.

cAMP enhances inducible nitric oxide synthase mRNA stability in cardiac myocytes

1995 ◽  
Vol 269 (6) ◽  
pp. H2044-H2050 ◽  
Author(s):  
C. V. Oddis ◽  
R. L. Simmons ◽  
B. G. Hattler ◽  
M. S. Finkel
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Cardiac Myocytes ◽  
Inducible Nitric Oxide Synthase ◽  
Mrna Stability ◽  
Neonatal Rat ◽  
Inos Mrna ◽  
No Production ◽  
Oxide Synthase ◽  
Inducible Nitric Oxide

The effects of adenosine 3',5'-cyclic monophosphate (cAMP) on cardiac myocyte nitric oxide (NO) production were studied. Maximal nitrite (NO2(-)) production by cultured neonatal rat cardiac myocytes was achieved with 500 U/ml interleukin-1 beta (IL-1 beta) for 48 h (4.6 +/- 0.3 nmol/1.25 x 10(5) cells; n = 12). Cardiac myocytes exposed to 500 U/ml IL-1 beta for 48 h stained positively for inducible nitric oxide synthase (iNOS) by immunohistochemistry. Forskolin (FSK; adenylate cyclase stimulator) or dibutyryl cAMP (DBcAMP; membrane-permeable cAMP analogue) administration alone had no effect on NO2(-) production. The addition of FSK or DBcAMP to IL-1 beta significantly increased NO2-) levels vs. IL-1 beta alone (9.7 +/- 0.6 and 10.9 +/- 0.8 vs. 4.6 +/- 0.3 nmol/1.25 x 10(5) cells per 48 h, respectively; P < 0.01; n = 12). Semiquantitative reverse transcriptase-polymerase chain reaction revealed increased iNOS mRNA in myocytes treated with FSK+IL-1 beta or DBcAMP+IL-1 beta vs. those treated with IL-1 beta alone. The addition of FSK or DBcAMP to IL-1 beta increased iNOS mRNA half-life over IL-1 beta treatment alone (10.6, 11.7 vs. 2.4 h, respectively). Cardiac myocytes do not express iNOS in response to cAMP alone. Rather, cAMP enhances iNOS mRNA stability following cytokine exposure.

Leptin-induced nitric oxide production in white adipocytes is mediated through PKA and MAP kinase activation

2005 ◽  
Vol 289 (2) ◽  
pp. C379-C387 ◽  
Author(s):  
Nadia Mehebik ◽  
Anne-Marie Jaubert ◽  
Dominique Sabourault ◽  
Yves Giudicelli ◽  
Catherine Ribière
Keyword(s):  
Nitric Oxide ◽  
Adipose Tissue ◽  
Protein Kinase ◽  
Protein Kinase A ◽  
Protein Phosphatase ◽  
Plasma Levels ◽  
Mitogen Activated Protein Kinase ◽  
No Production ◽  
Tyrosine Kinase 2 ◽  
Kinase A

Leptin injection increases plasma levels of nitrites and/or nitrates, an index of nitric oxide (NO) production. Because plasma levels of NO are correlated with fat mass and because adipose tissue is the main source of leptin, it seems that adipose tissue plays a major role in NO release induced by leptin. Adipocytes express both leptin receptors and nitric oxide synthase (NOS; including the endothelial isoform, NOS III, and the inducible isoform, NOS II). In this study, we have demonstrated that physiological concentrations of leptin stimulate NOS activity in adipocytes. This effect of leptin is abolished by 1) AG490, an inhibitor of Janus tyrosine kinase 2/signal transducer and activator of transcription 3; 2) U0126, an inhibitor of mitogen-activated protein kinase kinase/extracellular signal-regulated kinase (p42/p44 MAPK); and 3) N-[2-( p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide (H-89) or Rp diastereomer of adenosine 3′,5′-cyclic phosphorothioate, two inhibitors of protein kinase A, but not by wortmannin, an inhibitor of phosphatidylinositol 3-kinase. Immunoblotting studies have shown that leptin fails to activate Akt but increases p42/p44 MAPK phosphorylation, an effect that is prevented by U0126 but not by H-89. Furthermore, leptin induces NOS III phosphorylation at Ser1179and Thr497, but not when adipocytes are pretreated with H-89 or U0126. Finally, stimulation of adipocyte NOS activity by leptin is either unaltered when protein phosphatase 2A is inhibited by 1 nM okadaic acid or completely abolished when protein phosphatase 1 (PP1) activity is inhibited by 3 nM tautomycin, which supports a crucial role for PP1 in mediating this effect of leptin. On the whole, these experiments demonstrate that NOS activity is a novel target for leptin in adipocytes and that the leptin-induced NOS activity is at least in part the result of NOS III phosphorylations via both protein kinase A and p42/p44 MAPK activation. More generally, this study also leads to the hypothesis of NO as a potentially important factor for leptin signaling in adipocytes.

scholarly journals Protein kinase A mediates glucagon-like peptide 1-induced nitric oxide production and muscle microvascular recruitment

2013 ◽  
Vol 304 (2) ◽  
pp. E222-E228 ◽  
Author(s):  
Zhenhua Dong ◽  
Weidong Chai ◽  
Wenhui Wang ◽  
Lina Zhao ◽  
Zhuo Fu ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Blood Flow ◽  
Protein Kinase ◽  
Protein Kinase A ◽  
Microvascular Blood Flow ◽  
No Production ◽  
Glucagon Like Peptide ◽  
Glucagon Like Peptide 1 ◽  
Microvascular Recruitment ◽  
Kinase A

Glucagon-like peptide-1 (GLP-1) causes vasodilation and increases muscle glucose uptake independent of insulin. Recently, we have shown that GLP-1 recruits muscle microvasculature and increases muscle glucose use via a nitric oxide (NO)-dependent mechanism. Protein kinase A (PKA) is a major signaling intermediate downstream of GLP-1 receptors. To examine whether PKA mediates GLP-1's microvascular action in muscle, GLP-1 was infused to overnight-fasted male rats for 120 min in the presence or absence of H89, a PKA inhibitor. Hindleg muscle microvascular recruitment and glucose use were determined. GLP-1 infusion acutely increased muscle microvascular blood volume within 30 min without altering microvascular blood flow velocity or blood pressure. This effect persisted throughout the 120-min infusion period, leading to a significant increase in muscle microvascular blood flow. These changes were paralleled with an approximately twofold increase in plasma NO levels and hindleg glucose extraction. Systemic infusion of H89 completely blocked GLP-1-mediated muscle microvascular recruitment and increases in NO production and muscle glucose extraction. In cultured endothelial cells, GLP-1 acutely increased PKA activity and stimulated endothelial NO synthase phosphorylation at Ser1177 and NO production. PKA inhibition abolished these effects. In ex vivo studies, perfusion of the distal saphenous artery with GLP-1 induced significant vasorelaxation that was also abolished by pretreatment of the vessels with PKA inhibitor H89. We conclude that GLP-1 recruits muscle microvasculature by expanding microvascular volume and increases glucose extraction in muscle via a PKA/NO-dependent pathway in the vascular endothelium. This may contribute to postprandial glycemic control and complication prevention in diabetes.

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

2000 ◽  
Vol 279 (6) ◽  
pp. H3138-H3143 ◽  
Author(s):  
Hong Kan ◽  
Zirong Xie ◽  
Mitchell S. Finkel
Keyword(s):  
Protein Kinase ◽  
Cardiac Myocytes ◽  
Myocardial Dysfunction ◽  
Neonatal Rat ◽  
Inos Mrna ◽  
Protein Kinase Activity ◽  
No Production ◽  
Direct Effects ◽  
Hiv Gp120 ◽  
Hiv 1

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-1β (IL-1β) (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-1β, recombinant HIV-gp120 (1 μg/ml) alone failed to enhance NO production in CM (0.5 ± 0.4 vs. 0.4 ± 0.5 μmol/1.25 × 105 cells/48 h, gp120 vs. control, respectively; n = 12, P = not significant). However, the addition of gp120 to IL-1β significantly enhanced iNOS mRNA expression (70 ± 1.5 vs. 26 ± 2.4 optical units, IL-1β + gp120 vs. IL-1β, respectively; n = 3), iNOS protein synthesis (42 ± 1.4 vs. 18 ± 0.8 optical units, IL-1β + gp120 vs. IL-1β, respectively; n = 3), and NO production (NO2 −) (6.6 ± 0.6 vs. 4.1 ± 0.8 μmol/1.25 × 105 cells/48 h, IL-1β + gp120 vs. IL-1β, respectively; n = 12, P ≤ 0.5). HIV-gp120 enhancement of IL-1β-induced NO2 −production was blocked by 10 μM of SB-203580 (SB), a selective p38 protein kinase inhibitor (3.6 ± 0.2 vs. 6.6 ± 0.6 μmol/1.25 × 105 cells/48 h, IL-1β + gp120 + SB vs. IL-1β + gp120, respectively; n = 12, P ≤ 0.5). HIV-gp120-enhanced p38 protein kinase activity was associated with an increase in IL-1β-stimulated NF-κB activity (184 ± 12.7 vs. 92 ± 10.7 optical units, IL-1β + gp120 vs. IL-1β, respectively; n = 3). None of these effects was seen with another recombinant HIV-1 protein, Tat. Thus HIV-gp120 enhancement of IL-1β-induced NO production is associated with p38-mediated activation of NF-κB. Direct effects of HIV-gp120 on CM may provide a previously unrecognized mechanism contributing to HIV cardiomyopathy.

Interplay of VIP and nitric oxide in regulation of the descending relaxation phase of peristalsis

1993 ◽  
Vol 264 (2) ◽  
pp. G334-G340 ◽  
Author(s):  
J. R. Grider
Keyword(s):  
Nitric Oxide ◽  
Smooth Muscle ◽  
Protein Kinase ◽  
Protein Kinase G ◽  
Rat Colon ◽  
No Production ◽  
Field Stimulation ◽  
Relaxant Effect ◽  
Relaxation Phase ◽  
Kinase A

Involvement of vasoactive intestinal peptide (VIP) and nitric oxide (NO) in neurally induced relaxation was examined in smooth muscle from rat colon. Relaxation induced by field stimulation or radial stretch (i.e., descending relaxation phase of the peristaltic reflex) was accompanied by VIP release and NO production. NG-nitro-L-arginine (L-NNA) abolished NO production in both preparations but only partly inhibited VIP release (45 +/- 8% at 8 Hz and 59 +/- 10% at 10 g stretch) and relaxation (62 +/- 5% and 35 +/- 6%); the effect of L-NNA was reversed by L-arginine but not D-arginine. The pattern implied that NO production normally acts to enhance VIP release. In addition, VIP induced relaxation and stimulated NO production in muscle strips and isolated colonic muscle cells: L-NNA abolished NO production but only partly inhibited relaxation (58 +/- 6%); oxyhemoglobin had no effect. The effect of L-NNA on relaxation was reversed by L-arginine but not by D-arginine. The protein kinase A inhibitor (R)-p-adenosine 3',5'-cyclic phosphorothioate [(R)-p-cAMPS] and the protein kinase G inhibitor KT5823 inhibited VIP-induced relaxation by 76 +/- 5 and 35 +/- 4%, respectively; a combination of the two inhibitors abolished relaxation. (R)-p-cAMPS blocked the direct relaxant effect of VIP, whereas KT5823 blocked the indirect effect of VIP mediated by NO.(ABSTRACT TRUNCATED AT 250 WORDS)

Interleukin-1 alpha stimulates dopamine release by activating type II protein kinase A in PC-12 cells

1995 ◽  
Vol 269 (6) ◽  
pp. E1083-E1088
Author(s):  
A. Joseph ◽  
A. Kumar ◽  
N. A. O'Connell ◽  
R. K. Agarwal ◽  
A. R. Gwosdow
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Dopamine Release ◽  
Interleukin 1 ◽  
Intracellular Camp ◽  
Type I ◽  
Camp Accumulation ◽  
Type Ii ◽  
Pc 12 Cells ◽  
Kinase A

A recent study from this laboratory [A. R. Gwosdow, N. A. O'Connell, and A. B. Abou-Samra. Am. J. Physiol. 263 (Endocrinol. Metab. 26): E461-E466, 1992] showed that the inflammatory mediator interleukin-1 alpha (IL-1 alpha) stimulates catecholamine release from primary cultures of rat adrenal cells. The present studies were conducted to determine whether 1) IL-1 alpha stimulates catecholamine/dopamine release from the adrenal medullary cell line PC-12 and 2) the adenosine 3',5'-cyclic monophosphate (cAMP)-protein kinase A (PKA) pathway is involved in IL-1 alpha-induced dopamine release from PC-12 cells. The results indicate that IL-1 alpha significantly (P < 0.05) elevated dopamine release after a 24-h incubation period. IL-1 alpha did not stimulate cAMP accumulation at any time period between 5 min and 2 h. In contrast, forskolin-treated cells elevated (P < 0.05) intracellular cAMP levels and increased dopamine release. Because IL-1 alpha did not affect cAMP accumulation, the effect of IL-1 alpha on PKA activity was investigated. IL-1 alpha increased (P < 0.05) PKA activity at 15 and 30 min and returned to control levels by 1 h. Forskolin also increased (P < 0.05) PKA activity. The type of PKA activated (P < 0.05) by IL-1 alpha was type II PKA. In contrast, forskolin activated (P < 0.05) type I and type II PKA. Inhibition of PKA with the PKA inhibitor H-8 blocked PKA activity and dopamine secretion by both IL-1 alpha and forskolin in PC-12 cells. These observations demonstrate that 1) IL-1 alpha stimulated dopamine release from PC-12 cells by activating PKA, 2) the mechanism of IL-1 alpha activation of PKA does not involve detectable increases in intracellular cAMP accumulation, and 3) IL-1 alpha activates type II PKA, which is used by IL-1 alpha to stimulate dopamine secretion from PC-12 cells.

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