Antiapoptotic effect of calcitonin gene-related peptide on oxidative stress-induced injury in H9c2 cardiomyocytes via the RAMP1/CRLR complex.
Calcitonin gene-related peptide (CGRP) plays an important role in the mediation of protective effects observed in situations such as ischemic preconditioning in rat hearts. In this study, we investigated in H9c2 rat cardiomyoblasts if the protective effect of CGRP could be linked to an inhibitory effect on the apoptotic pathway. We also determined the specificity of observed effects by treatment with adrenomedullin (ADM) in stress conditions generated by 100 microM hydrogen peroxide. Using MTT assays, we demonstrate that a pretreatment with CGRP decreases by half the loss of cell viability induced by H(2)O(2). CGRP inhibits phosphatidylserine externalization, caspase 3 activation and DNA fragmentation due to oxidative stress. Using RT-PCR, we observed an increase in Bcl-2 mRNA expression induced by CGRP treatment. Dot blotting experiments showed that, in stress conditions, Bcl-2 protein level decreases while Bax is increased. CGRP administration prior to stress prevents these effects. The three-receptor activity modifying protein (RAMP) isotypes were detected by RT-PCR in H9c2 cells and in left ventricle rat tissue, RAMP1 and RAMP3 being the most abundant in both cases. RAMP1 expression was upregulated by CGRP while RAMP3 mRNA level was decreased. Cell viability assessed by MTT indicates that, contrary to CGRP, pretreatment of stressed cells with ADM, a RAMP2 agonist, fails to protect them while treatment with CGRP(8-37) (a RAMP1 and 2 inhibitor) abolished CGRP protective effect. Taken together, these data suggest that CGRP has antiapoptotic properties through the RAMP1/CRLR complex. CGRP could be used to prevent apoptosis in an ischemia-reperfusion context.
Sueur
Stéphanie
S
Laboratoire de Physiopathologie et Pharmacologie Cardiovasculaires Experimentales, Biologie Animale Cellulaire et Moléculaire, Faculté des Sciences Gabriel, Université de Bourgogne, IFR Santé 100, 6, boulevard Gabriel, 21000 Dijon, France.
Pesant
Matthieu
M
Rochette
Luc
L
Connat
Jean-Louis
JL
D000818:Animals; D015740:Calcitonin Gene-Related Peptide / Q000494:pharmacology*; D058286:Calcitonin Receptor-Like Protein; D002460:Cell Line; D053938:DNA Fragmentation / Q000187:drug effects*; D005786:Gene Expression Regulation / Q000187:drug effects; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D019157:Ischemic Preconditioning, Myocardial; D008565:Membrane Proteins / Q000378:metabolism*; D008916:Miotics / Q000494:pharmacology*; D046912:Multiprotein Complexes / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000378:metabolism*; D018384:Oxidative Stress / Q000187:drug effects*; D010446:Peptide Fragments / Q000494:pharmacology*; D051381:Rats; D058262:Receptor Activity-Modifying Protein 1; D058263:Receptor Activity-Modifying Protein 2; D058264:Receptor Activity-Modifying Protein 3; D058261:Receptor Activity-Modifying Proteins; D018003:Receptors, Calcitonin / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects
D058286:Calcitonin Receptor-Like Protein; C545580:Calcrl protein, rat; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D008916:Miotics; D046912:Multiprotein Complexes; D010446:Peptide Fragments; C545508:Ramp1 protein, rat; C545509:Ramp2 protein, rat; C545510:Ramp3 protein, rat; D058262:Receptor Activity-Modifying Protein 1; D058263:Receptor Activity-Modifying Protein 2; D058264:Receptor Activity-Modifying Protein 3; D058261:Receptor Activity-Modifying Proteins; D018003:Receptors, Calcitonin; C058636:calcitonin gene-related peptide (8-37); D015740:Calcitonin Gene-Related Peptide