Postconditioning leads to an increase in protein S-nitrosylation.
Previous studies have shown a role for nitric oxide and S-nitrosylation (SNO) in postconditioning (PostC), but specific SNO proteins and sites have not been identified in the myocardium after PostC. In this study, we examined SNO signaling in PostC using a Langendorff-perfused mouse heart model. After 20 min of equilibrium perfusion and 25 min of global ischemia, PostC was applied at the beginning of reperfusion with six cycles of 10 s of reperfusion and 10 s of ischemia. The total period of reperfusion was 90 min. Compared with the ischemia-reperfusion (I/R) control, PostC significantly reduced postischemic contractile dysfunction and infarct size. PostC-induced protection was blocked by treatment with N(G)-nitro-l-arginine methyl ester (l-NAME) (10 μmol/l; a constitutive NO synthase inhibitor), but not by either ODQ (10 μmol/l, a highly selective soluble guanylyl cyclase inhibitor) or KT5823 (1 μmol/l, a specific protein kinase G inhibitor). Two biotin switch based methods, two dimensional CyDye-maleimide difference gel electrophoresis (2D CyDye-maleimide DIGE) and SNO-resin-assisted capture (SNO-RAC), were utilized to identify SNO-modified proteins and sites. Using 2D CyDye-maleimide DIGE analysis, PostC was found to cause a 25% or greater increase in SNO of a number of proteins, which was blocked by treatment with l-NAME in parallel with the loss of protection. Using SNO-RAC, we identified 77 unique proteins with SNO sites after PostC. These results suggest that NO-mediated SNO signaling is involved in PostC-induced cardioprotection and these data provide the first set of candidate SNO proteins in PostC hearts.
Department of Cardiovascular Surgery, Guangzhou General Hospital of Guangzhou Military Command, Guangzhou, Guangdong Province, China;
Guang
G
Tong
Angel M
AM
Aponte
Mark J
MJ
Kohr
Charles
C
Steenbergen
Elizabeth
E
Murphy
Junhui
J
Sun
D000818:Animals; D002227:Carbazoles / Q000494:pharmacology; D057775:Ischemic Postconditioning; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D023421:Models, Animal; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism*; D017293:Protein S / Q000378:metabolism*; D015427:Reperfusion Injury / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects
D002227:Carbazoles; D017293:Protein S; C073601:KT 5823; D009569:Nitric Oxide; D019331:NG-Nitroarginine Methyl Ester
ischemic postconditioning; nitric oxide; protein S-nitrosylation; soluble guanylyl cyclase/cGMP