Redox activation of Ref-1 potentiates cell survival following myocardial ischemia reperfusion injury.
A recent study showed that cardiac adaptation could potentiate translocation of thioredoxin-1 (Trx-1) into the nucleus, which then interacted with Ref-1, resulting in a survival signal. Here, we present evidence that such adaptation also causes nuclear translocation of Ref-1, which is almost completely inhibited when the hearts were pretreated with antisense Ref-1 that also abolished the cardioprotective adaptive response. Significant amounts of NFkappaB and Nrf2 were found to be associated with Ref-1 when the nuclear extract obtained from the left ventricle was immunoprecipitated with Ref-1. Such Ref-1-NFkappaB and Ref-1-Nrf2 interactions were significantly inhibited with antisense Ref-1. However, immunoprecipitation of nuclear extract with NFkappaB showed that the association of Trx-1 with NFkappaB is increased in the adapted heart, which was again significantly blocked by antisense Ref-1. Nrf2 was also associated with NFkappaB; however, such association appeared to be independent of Ref-1. In contrast, myocardial adaptation to ischemia inhibited the ischemia reperfusion-induced loss of Nrf2 from the nucleus, which was inhibited by antisense Ref-1. The nuclear translocation and activation of Ref-1 appeared to generate a survival signal as evidenced by the increased phosphorylation of Akt that was inhibited with antisense Ref-1. Finally, confocal microscopy confirmed the results of immunoblotting, clearly showing the nuclear translocation of Ref-1 and nuclear 3D colocalization of Ref-1 with NFkappaB in the adapted heart and its inhibition with antisense Ref-1. Our results show that PC potentiates a survival signal through the phosphorylation of Akt by causing nuclear translocation and activation of Ref-1, where significant interaction among NFkappaB and Ref-1, Trx-1, and Nrf2 appears to regulate Ref-1-induced survival signal.
Gurusamy
Narasimman
N
Cardiovascular Research Center, University of Connecticut School of Medicine, Farmington, CT 06030-1110, USA.
Malik
Gautam
G
Gorbunov
Nikolai V
NV
Das
Dipak K
DK
D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D002470:Cell Survival / Q000502:physiology; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase / Q000502:physiology*; D004789:Enzyme Activation; D008297:Male; D008315:Malondialdehyde / Q000378:metabolism; D015428:Myocardial Reperfusion Injury / Q000503:physiopathology*; D009206:Myocardium / Q000378:metabolism; D051267:NF-E2-Related Factor 2 / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D020319:Oligodeoxyribonucleotides, Antisense / Q000494:pharmacology; D010766:Phosphorylation; D021381:Protein Transport / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013879:Thioredoxins / Q000378:metabolism
D051267:NF-E2-Related Factor 2; D016328:NF-kappa B; C495637:Nfe2l2 protein, rat; D020319:Oligodeoxyribonucleotides, Antisense; C495697:Txn1 protein, rat; D008315:Malondialdehyde; D013879:Thioredoxins; D051057:Proto-Oncogene Proteins c-akt; C470319:Apex1 protein, rat; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase