Grape seed proanthocyanidins protect cardiomyocytes from ischemia and reperfusion injury via Akt‐NOS signaling

Abstract

Ischemia/reperfusion (I/R) injury in cardiomyocytes is related to excess reactive oxygen species (ROS) generation and can be modulated by nitric oxide (NO). We have previously shown that grape seed proanthocyanidin extract (GSPE), a naturally occurring antioxidant, decreased ROS and may potentially stimulate NO production. In this study, we investigated whether GSPE administration at reperfusion was associated with cardioprotection and enhanced NO production in a cardiomyocyte I/R model. GSPE attenuated I/R-induced cell death [18.0 +/- 1.8% (GSPE, 50 microg/ml) vs. 42.3 +/- 3.0% (I/R control), P < 0.001], restored contractility (6/6 vs. 0/6, respectively), and increased NO release. The NO synthase (NOS) inhibitor Nomega-nitro-L-arginine methyl ester (L-NAME, 200 microM) significantly reduced GSPE-induced NO release and its associated cardioprotection [32.7 +/- 2.7% (GSPE + L-NAME) vs. 18.0 +/- 1.8% (GSPE alone), P < 0.01]. To determine whether GSPE induced NO production was mediated by the Akt-eNOS pathway, we utilized the Akt inhibitor API-2. API-2 (10 microM) abrogated GSPE-induced protection [44.3% +/- 2.2% (GSPE + API-2) vs. 27.0% +/- 4.3% (GSPE alone), P < 0.01], attenuated the enhanced phosphorylation of Akt at Ser473 in GSPE-treated cells and attenuated GSPE-induced NO increases. Simultaneously blocking NOS activation (L-NAME) and Akt (API-2) resulted in decreased NO levels similar to using each inhibitor independently. These data suggest that in the context of GSPE stimulation, Akt may help activate eNOS, leading to protective levels of NO. GSPE offers an alternative approach to therapeutic cardioprotection against I/R injury and may offer unique opportunities to improve cardiovascular health by enhancing NO production and increasing Akt-eNOS signaling.

The protective effect of picroside II against hypoxia/reoxygenation injury in neonatal rat cardiomyocytes

“…These findings further suggest that overproduction of ROS plays a critical role in the cardiomyocytes injury induced by ischemia/reperfusion or H/R. It has also been reported that administration of antioxidants or free radical scavengers should give potential benefits by alleviating ischemia/ reperfusion injury (Shao et al, 2009).…”

The Molecular Mechanism of Protective Effects of Grape Seed Proanthocyanidin Extract on Reperfusion Arrhythmias in Rats in Vivo

“…Study had shown that GSPE stimulated nitric oxide production in untreated cardiomyocytes in a dose-dependent manner. 14) However the molecular mechanisms of the cardioprotection of GSPE are not fully elucidated.…”

“…A series of studies were conducted using GSPE to demonstrate its cardioprotective ability in animals and humans. [10][11][12][13][14] Our previous work had shown GSPE reduced reperfusioninduced arrhythmias in rabbits. 15) In isolated rat hearts GSPE had been proved to reduce the incidence of reperfusion-induced VF and VT. 11) However the mechanism of this protection against reperfusion-induced VT, VF by GSPE remained unclear.…”

Therapeutic hypothermia cardioprotection via Akt- and nitric oxide-mediated attenuation of mitochondrial oxidants

“…After the membranes were blocked in 5% fat-free milk in TBS-Tween 20, they were probed with antibodies against phosphorylated Akt (Thr308 and Ser473), phosphorylated heat shock protein 27 (HSP27) (Ser82), total Akt, tubulin, and/or total HSP-27. Signals were amplified and visualized with horseradish peroxidase-conjugated secondary antibody and enhanced chemiluminescence (30). Densitometry was performed using ImageJ software (version 1.38, NIH).…”

“…A chargedcoupled device camera was used to monitor contractions and membrane changes over time in the same field of cells (ϳ70 ϫ 90 m). Fluorescent images were acquired from a cooled Cool-SNAP-ES camera (Photometrics, Tuscon, AZ), and changes in fluorescent intensity over time were quantified with MetaMorph software (Molecular Devices, Downington, PA) (30).…”