MAPK signaling pathways are needed for survival of H9c2 cardiac myoblasts under extracellular alkalosis.
pH is one of the most important physiological parameters, with its changes affecting the function of vital organs like the heart. However, the effects of alkalosis on the regulation of cardiac myocyte function have not been extensively investigated. Therefore, we decided to study whether the mitogen-activated protein kinase (MAPK) signaling pathways [c-Jun NH2-terminal kinases (JNKs), extracellular signal-regulated kinases (ERKs), and p38 MAPK] are activated by alkalosis induced with Tris-Tyrode buffer at two pH values, 8.5 and 9.5, in H9c2 rat cardiac myoblasts. These buffers also induced intracellular alkalinization comparable to that induced by 1 mM NH4Cl. The three MAPKs examined presented differential phosphorylation patterns that depended on the severity and the duration of the stimulus. Inhibition of Na+/H+ exchanger (NHE)1 by its inhibitor HOE-642 prevented alkalinization and partially attenuated the alkalosis (pH 8.5)-induced activation of these kinases. The same stimulus also promoted c-Jun phosphorylation and enhanced the binding at oligonucleotides bearing the activator protein-1 (AP-1) consensus sequence, all in a JNK-dependent manner. Additionally, mitogen- and stress-activated kinase 1 (MSK1) was transiently phosphorylated by alkalosis (pH 8.5), and this was abolished by the selective inhibitors of either p38 MAPK or ERK pathways. JNKs also mediated Bcl-2 phosphorylation in response to incubation with the alkaline medium (pH 8.5), while selective inhibitors of the three MAPKs diminished cell viability under these conditions. All these data suggest that alkalosis activates MAPKs in H9c2 cells and these kinases, in turn, modify proteins that regulate gene transcription and cell survival.
Stathopoulou
Konstantina
K
Department of Animal and Human Physiology, School of Biology, Faculty of Sciences, University of Athens, Panepistimioupolis, Athens 157 84 Greece.
Beis
Isidoros
I
Gaitanaki
Catherine
C
D000471:Alkalosis / Q000201:enzymology / Q000378:metabolism / Q000473:pathology*; D000818:Animals; D015153:Blotting, Western; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D024202:Electrophoretic Mobility Shift Assay; D004789:Enzyme Activation / Q000502:physiology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D066298:In Vitro Techniques; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D049449:Luminescence; D020928:Mitogen-Activated Protein Kinases / Q000502:physiology*; D032446:Myoblasts / Q000502:physiology*; D032383:Myocytes, Cardiac / Q000502:physiology*; D010766:Phosphorylation; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction / Q000502:physiology*; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D018808:Transcription Factor AP-1 / Q000096:biosynthesis / Q000235:genetics; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D016755:Proto-Oncogene Proteins c-jun; D017923:Sodium-Hydrogen Exchangers; D018808:Transcription Factor AP-1; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases