Role of cGMP-PKG signaling in the protection of neonatal rat cardiac myocytes subjected to simulated ischemia/reoxygenation

A. Görbe, Z. Giricz, Andrea Szunyog, T. Csont, Dwaine S. Burley, Gary F. Baxter, P. Ferdinándy

Research output: Contribution to journalArticle

57 Citations (Scopus)

Abstract

Nitric oxide (NO) and B-type natriuretic peptide (BNP) are protective against ischemia-reperfusion injury as they increase intracellular cGMP level via activation of soluble (sGC) or particulate guanylate cyclases (pGC), respectively. The aim of the present study was to examine if the cGMP-elevating mediators, NO and BNP, share a common downstream signaling pathway via cGMP-dependent protein kinase (PKG) in cardiac cytoprotection. Neonatal rat cardiac myocytes in vitro were subjected to 2.5 h simulated ischemia (SI) followed by 2 h reoxygenation. Cell viability was tested by trypan blue exclusion assay. PKG activity of cardiac myocytes was assessed by phospholamban (PLB) phosphorylation determined by western blot. Cell death was 34 ± 2% after SI/reoxygenation injury in the control group. cGMP-inducing agents significantly decreased irreversible cell injury: the cGMP analog 8-bromo-cGMP (8-Br-cGMP, 10 nM) decreased it to 13 ± 1% (p <0.001), the direct NO-donor S-nitroso-N-acetylpenicillamine (SNAP, 1 μM) to 18 ± 6% (p <0.05) and BNP (10 nM) to 12 ± 2% (p <0.001), respectively. This protective effect was abolished by the selective PKG inhibitor KT-5823 (600 nM) in each case. As PLB is not a unique reporter for PKG activity since it is also phosphorylated by protein kinase A (PKA), we examined PLB phosphorylation in the presence of the PKA inhibitor KT-5720 (1 μM). The ratio of pPLB/PLB significantly increased after administration of both BNP and 8-Br-cGMP under ischemic conditions, which was abolished by the PKG inhibitor. This is the first demonstration that elevated cGMP produced either by the sGC activator SNAP or the pGC activator BNP exerts cytoprotective effects via a common downstream signaling pathway involving PKG activation.

Original languageEnglish
Pages (from-to)643-650
Number of pages8
JournalBasic Research in Cardiology
Volume105
Issue number5
DOIs
Publication statusPublished - Sep 2010

Fingerprint

Brain Natriuretic Peptide
Cardiac Myocytes
Ischemia
KT 1
Cyclic AMP-Dependent Protein Kinases
Nitric Oxide
Phosphorylation
S-Nitroso-N-Acetylpenicillamine
Cyclic GMP-Dependent Protein Kinases
Cytoprotection
Trypan Blue
Nitric Oxide Donors
Guanylate Cyclase
Wounds and Injuries
Protein Kinase Inhibitors
Reperfusion Injury
Cell Survival
Cell Death
Western Blotting
Control Groups

Keywords

  • BNP
  • Cardiomyocyte
  • CGMP
  • NO
  • Protein kinase G
  • Simulated ischemia

ASJC Scopus subject areas

  • Cardiology and Cardiovascular Medicine
  • Physiology (medical)
  • Physiology

Cite this

Role of cGMP-PKG signaling in the protection of neonatal rat cardiac myocytes subjected to simulated ischemia/reoxygenation. / Görbe, A.; Giricz, Z.; Szunyog, Andrea; Csont, T.; Burley, Dwaine S.; Baxter, Gary F.; Ferdinándy, P.

In: Basic Research in Cardiology, Vol. 105, No. 5, 09.2010, p. 643-650.

Research output: Contribution to journalArticle

@article{bed7605b62b2461bb74a4c97e781956e,
title = "Role of cGMP-PKG signaling in the protection of neonatal rat cardiac myocytes subjected to simulated ischemia/reoxygenation",
abstract = "Nitric oxide (NO) and B-type natriuretic peptide (BNP) are protective against ischemia-reperfusion injury as they increase intracellular cGMP level via activation of soluble (sGC) or particulate guanylate cyclases (pGC), respectively. The aim of the present study was to examine if the cGMP-elevating mediators, NO and BNP, share a common downstream signaling pathway via cGMP-dependent protein kinase (PKG) in cardiac cytoprotection. Neonatal rat cardiac myocytes in vitro were subjected to 2.5 h simulated ischemia (SI) followed by 2 h reoxygenation. Cell viability was tested by trypan blue exclusion assay. PKG activity of cardiac myocytes was assessed by phospholamban (PLB) phosphorylation determined by western blot. Cell death was 34 ± 2{\%} after SI/reoxygenation injury in the control group. cGMP-inducing agents significantly decreased irreversible cell injury: the cGMP analog 8-bromo-cGMP (8-Br-cGMP, 10 nM) decreased it to 13 ± 1{\%} (p <0.001), the direct NO-donor S-nitroso-N-acetylpenicillamine (SNAP, 1 μM) to 18 ± 6{\%} (p <0.05) and BNP (10 nM) to 12 ± 2{\%} (p <0.001), respectively. This protective effect was abolished by the selective PKG inhibitor KT-5823 (600 nM) in each case. As PLB is not a unique reporter for PKG activity since it is also phosphorylated by protein kinase A (PKA), we examined PLB phosphorylation in the presence of the PKA inhibitor KT-5720 (1 μM). The ratio of pPLB/PLB significantly increased after administration of both BNP and 8-Br-cGMP under ischemic conditions, which was abolished by the PKG inhibitor. This is the first demonstration that elevated cGMP produced either by the sGC activator SNAP or the pGC activator BNP exerts cytoprotective effects via a common downstream signaling pathway involving PKG activation.",
keywords = "BNP, Cardiomyocyte, CGMP, NO, Protein kinase G, Simulated ischemia",
author = "A. G{\"o}rbe and Z. Giricz and Andrea Szunyog and T. Csont and Burley, {Dwaine S.} and Baxter, {Gary F.} and P. Ferdin{\'a}ndy",
year = "2010",
month = "9",
doi = "10.1007/s00395-010-0097-0",
language = "English",
volume = "105",
pages = "643--650",
journal = "Basic Research in Cardiology",
issn = "0300-8428",
publisher = "D. Steinkopff-Verlag",
number = "5",

}

TY - JOUR

T1 - Role of cGMP-PKG signaling in the protection of neonatal rat cardiac myocytes subjected to simulated ischemia/reoxygenation

AU - Görbe, A.

AU - Giricz, Z.

AU - Szunyog, Andrea

AU - Csont, T.

AU - Burley, Dwaine S.

AU - Baxter, Gary F.

AU - Ferdinándy, P.

PY - 2010/9

Y1 - 2010/9

N2 - Nitric oxide (NO) and B-type natriuretic peptide (BNP) are protective against ischemia-reperfusion injury as they increase intracellular cGMP level via activation of soluble (sGC) or particulate guanylate cyclases (pGC), respectively. The aim of the present study was to examine if the cGMP-elevating mediators, NO and BNP, share a common downstream signaling pathway via cGMP-dependent protein kinase (PKG) in cardiac cytoprotection. Neonatal rat cardiac myocytes in vitro were subjected to 2.5 h simulated ischemia (SI) followed by 2 h reoxygenation. Cell viability was tested by trypan blue exclusion assay. PKG activity of cardiac myocytes was assessed by phospholamban (PLB) phosphorylation determined by western blot. Cell death was 34 ± 2% after SI/reoxygenation injury in the control group. cGMP-inducing agents significantly decreased irreversible cell injury: the cGMP analog 8-bromo-cGMP (8-Br-cGMP, 10 nM) decreased it to 13 ± 1% (p <0.001), the direct NO-donor S-nitroso-N-acetylpenicillamine (SNAP, 1 μM) to 18 ± 6% (p <0.05) and BNP (10 nM) to 12 ± 2% (p <0.001), respectively. This protective effect was abolished by the selective PKG inhibitor KT-5823 (600 nM) in each case. As PLB is not a unique reporter for PKG activity since it is also phosphorylated by protein kinase A (PKA), we examined PLB phosphorylation in the presence of the PKA inhibitor KT-5720 (1 μM). The ratio of pPLB/PLB significantly increased after administration of both BNP and 8-Br-cGMP under ischemic conditions, which was abolished by the PKG inhibitor. This is the first demonstration that elevated cGMP produced either by the sGC activator SNAP or the pGC activator BNP exerts cytoprotective effects via a common downstream signaling pathway involving PKG activation.

AB - Nitric oxide (NO) and B-type natriuretic peptide (BNP) are protective against ischemia-reperfusion injury as they increase intracellular cGMP level via activation of soluble (sGC) or particulate guanylate cyclases (pGC), respectively. The aim of the present study was to examine if the cGMP-elevating mediators, NO and BNP, share a common downstream signaling pathway via cGMP-dependent protein kinase (PKG) in cardiac cytoprotection. Neonatal rat cardiac myocytes in vitro were subjected to 2.5 h simulated ischemia (SI) followed by 2 h reoxygenation. Cell viability was tested by trypan blue exclusion assay. PKG activity of cardiac myocytes was assessed by phospholamban (PLB) phosphorylation determined by western blot. Cell death was 34 ± 2% after SI/reoxygenation injury in the control group. cGMP-inducing agents significantly decreased irreversible cell injury: the cGMP analog 8-bromo-cGMP (8-Br-cGMP, 10 nM) decreased it to 13 ± 1% (p <0.001), the direct NO-donor S-nitroso-N-acetylpenicillamine (SNAP, 1 μM) to 18 ± 6% (p <0.05) and BNP (10 nM) to 12 ± 2% (p <0.001), respectively. This protective effect was abolished by the selective PKG inhibitor KT-5823 (600 nM) in each case. As PLB is not a unique reporter for PKG activity since it is also phosphorylated by protein kinase A (PKA), we examined PLB phosphorylation in the presence of the PKA inhibitor KT-5720 (1 μM). The ratio of pPLB/PLB significantly increased after administration of both BNP and 8-Br-cGMP under ischemic conditions, which was abolished by the PKG inhibitor. This is the first demonstration that elevated cGMP produced either by the sGC activator SNAP or the pGC activator BNP exerts cytoprotective effects via a common downstream signaling pathway involving PKG activation.

KW - BNP

KW - Cardiomyocyte

KW - CGMP

KW - NO

KW - Protein kinase G

KW - Simulated ischemia

UR - http://www.scopus.com/inward/record.url?scp=77956190993&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=77956190993&partnerID=8YFLogxK

U2 - 10.1007/s00395-010-0097-0

DO - 10.1007/s00395-010-0097-0

M3 - Article

VL - 105

SP - 643

EP - 650

JO - Basic Research in Cardiology

JF - Basic Research in Cardiology

SN - 0300-8428

IS - 5

ER -