The novel
pro-drug of 3'4'-dihydroxyflavonol, NP202, potently reduces
myocardial infarct size resulting from
ischemia-reperfusion (I/R) through mechanisms that remain to be fully defined. In this study, we investigated whether cardioprotection induced by NP202 depended on activation of the
reperfusion injury survival
kinase (RISK) pathways. We therefore examined the effects of
PD98059 and
LY294002, specific inhibitors of the
MEK/ERK1/2 and PI3K/Akt pathways, respectively. In isolated cardiomyocytes, H2O2induced oxidative stress activated ERK1/2 and this was further enhanced by DiOHF, the active parent compound of NP202. Although oxidative stress did not stimulate Akt in cardiomyocytes, co-treatment with DiOHF substantially increased Akt phosphorylation. This suggests that DiOHF is a potent modulator of RISK pathways specifically in the context of stress stimulation. In anesthetised sheep, following 1h
ischemia and 3h reperfusion, the contribution of the RISK pathways to NP202-mediated cardioprotection was determined by treating the animals with
PD98059,
LY294002 or vehicle prior to NP202 administration and reperfusion.
Infarct size, as a percentage of the area-at-risk, was substantially reduced by NP202 (from 78±6 to 46±4%, P<0.05). Inhibition of
MEK/ERK1/2 abolished the cardioprotective effects of NP202 (
infarct size 81±4%), whereas inhibition of PI3K/Akt had no effect (
infarct size 53±4%). Our combined cellular and animal studies indicate that NP202 potently protects against myocardial I/R injury through complex mechanisms that involved augmentation of
MEK/ERK1/2 signaling, but not PI3K/Akt signaling.