Abstract Postconditioning (PostC) can be obtained either with brief cycles of ischemia/reperfusion (I-PostC) or with a direct targeting of mitochondria with Diazoxide (phar- macological PostC, P-PostC). I-PostC may induce the activation of RISK and SAFE pathways and may favor nitric oxide production with S-Nitrosylation of proteins and redox signaling. It is not clear whether Diazoxide can lead to similar effects. We compared the effects of I-PostC and P-PostC on (a) kinases of RISK- and SAFE pathway, (b) S-Nitrosylation of mitochondrial proteins and (c) reduction of death signals (PKCd, cleaved caspase-3 and Beclin-1) in cytosolic and mitochondrial fractions. Isolated rat hearts underwent (1) perfusion without ische- mia (Sham), (2) ischemia/reperfusion (30-min ischemia plus 2-h reperfusion), (3) I-PostC (5 intermittent cycles of 10-s reperfusion and 10-s ischemia immediately after the 30-min ischemia), (4) P-PostC (Diazoxide 30 lM in the first of 3-min of reperfusion) or (5) I-PostC ? MPG or P-PostC ? MPG (MPG, 2-mercaptopropionylglycine 300 lM). Using Western blot and biotin switch assay, we found that P-PostC induced a redox sensible phosphor- ylation/translocation of Akt, ERK1/2 and GSK3b into the mitochondria, but not of phospho-STAT3, which was translocated into the mitochondria by I-PostC only. Either I-PostC or P-PostC increased mitochondrial S-Nitrosylated proteins (e.g., VDAC) and reduced the levels of phospho- PKCd, cleaved caspase-3 and Beclin-1. Therefore, direct targeting of mitochondria with Diazoxide (a) activates the RISK pathway via a redox signaling, (b) favors discrete mitochondrial protein S-Nitrosylation, including VDAC and (c) decreases signals of death. Intriguingly, phospho- STAT3 translocation is induced by I-PostC, but not by P-PostC, thus suggesting a redox-independent mechanism in the SAFE pathway

Diazoxide postconditioning induces mitochondrial protein S-Nitrosylation and a redox-sensitive mitochondrial phosphorylation/translocation of RISK elements:no role for SAFE

PENNA, Claudia;PERRELLI, MARIA-GIULIA;TULLIO, FRANCESCA;ANGOTTI, CARMELINA;CAMPOREALE, ANNALISA;POLI, Valeria;PAGLIARO, Pasquale
2013

Abstract

Abstract Postconditioning (PostC) can be obtained either with brief cycles of ischemia/reperfusion (I-PostC) or with a direct targeting of mitochondria with Diazoxide (phar- macological PostC, P-PostC). I-PostC may induce the activation of RISK and SAFE pathways and may favor nitric oxide production with S-Nitrosylation of proteins and redox signaling. It is not clear whether Diazoxide can lead to similar effects. We compared the effects of I-PostC and P-PostC on (a) kinases of RISK- and SAFE pathway, (b) S-Nitrosylation of mitochondrial proteins and (c) reduction of death signals (PKCd, cleaved caspase-3 and Beclin-1) in cytosolic and mitochondrial fractions. Isolated rat hearts underwent (1) perfusion without ische- mia (Sham), (2) ischemia/reperfusion (30-min ischemia plus 2-h reperfusion), (3) I-PostC (5 intermittent cycles of 10-s reperfusion and 10-s ischemia immediately after the 30-min ischemia), (4) P-PostC (Diazoxide 30 lM in the first of 3-min of reperfusion) or (5) I-PostC ? MPG or P-PostC ? MPG (MPG, 2-mercaptopropionylglycine 300 lM). Using Western blot and biotin switch assay, we found that P-PostC induced a redox sensible phosphor- ylation/translocation of Akt, ERK1/2 and GSK3b into the mitochondria, but not of phospho-STAT3, which was translocated into the mitochondria by I-PostC only. Either I-PostC or P-PostC increased mitochondrial S-Nitrosylated proteins (e.g., VDAC) and reduced the levels of phospho- PKCd, cleaved caspase-3 and Beclin-1. Therefore, direct targeting of mitochondria with Diazoxide (a) activates the RISK pathway via a redox signaling, (b) favors discrete mitochondrial protein S-Nitrosylation, including VDAC and (c) decreases signals of death. Intriguingly, phospho- STAT3 translocation is induced by I-PostC, but not by P-PostC, thus suggesting a redox-independent mechanism in the SAFE pathway
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371
383
cardioprotection; ischemia; nitric oxide; RISK; SAFE
C. Penna; M.-G. Perrelli; F. Tullio; C. Angotti; A. Camporeale; V. Poli; P. Pagliaro
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/2318/138647
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