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REDOX ACTIVATION OF REF‐1 POTENTIATES CELL SURVIVAL FOLLOWING MYOCARDIAL ISCHEMIA REPERFUSION INJURY
Author(s) -
Gurusamy Narasimman,
Malik Gautam,
Gorbunov Nikolai V,
Das Dipak K
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.5.a180-a
Subject(s) - chromosomal translocation , immunoprecipitation , microbiology and biotechnology , chemistry , protein kinase b , ischemia , phosphorylation , medicine , biology , biochemistry , gene
A recent study showed that cardiac adaptation could potentiate translocation of thioredoxin‐1 into the nucleus, which then interacted with Ref‐1 resulting a survival signal. Here, we present evidence that such adaptation also causes nuclear translocation of Ref‐1, which is inhibited when the hearts are treated with antisense Ref‐1 that also abolishes cardioprotective adaptive response. When the nuclear extract from the left ventricle was immunoprecipitated with Ref‐1 and the immunoprecipitate was probed for NFkB by Western immunoblotting, significant amount of NFkB was associated with Ref‐1. Such Ref‐1‐NFkB interaction was inhibited with antisense Ref‐1. When the nuclear extract was immunoprecipitated with NFkB and the immunoprecipitated pellet was probed for Trx‐1 and Nrf2, Trx‐1 showed increased association with NFkB in the adapted heart, which was again significantly blocked by antisense Ref‐1. Nrf2 was also associated with NFkB; however such association appeared to be independent of Ref‐1. In contrast, myocardial adaptation to ischemia inhibited the ischemia reperfusion‐induced loss of Nrf‐2 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, the confocal microscopy confirmed the results of immunoblotting clearly showing nuclear translocation of Ref‐1 in the adapted heart and its inhibition with antisense Ref‐1. Confocal immunofluorescence analysis further confirmed nuclear 3D co localization of Ref‐1 with NFkB in cardiomyocytes in the adapted hearts.