Background
Epigallocatechin-3-gallate (EGCG) has been documented for its beneficial effects protecting oxidative stress to cardiac cells. Previously, we have shown the EGCG-mediated cardiac protection by attenuating reactive oxygen species and cytosolic Ca 2+ in cardiac cells during oxidative stress and myocardial ischemia. Here, we aimed to seek a deeper elucidation of the molecular anti-oxidative capabilities of EGCG in an H 2 O 2 -induced oxidative stress model of myocardial ischemia injury using H9c2 rat cardiomyoblasts.
Results
Proteomics analysis was used to determine the differential expression of proteins in H9c2 cells cultured in the conditions of control, 400 μM H 2 O 2 exposure for 30 min with and/or without 10 to 20 μM EGCG pre-treatment. In this model, eight proteins associated with energy metabolism, mitochondrial electron transfer, redox regulation, signal transduction, and RNA binding were identified to take part in EGCG-ameliorating H 2 O 2 -induced injury in H9c2 cells. H 2 O 2 exposure increased oxidative stress evidenced by increases in reactive oxygen species and cytosolic Ca 2+ overload, increases in glycolytic protein, α-enolase, decreases in antioxidant protein, peroxiredoxin-4 , as well as decreases in mitochondrial proteins, including aldehyde dehydrogenase-2 , o rnithine aminotransferase , and succinate dehydrogenase ubiquinone flavoprotein subunit . All of these effects were reversed by EGCG pre-treatment. In addition, EGCG attenuated the H 2 O 2 -induced increases of Type II inositol 3, 4-bisphosphate 4-phosphatase and relieved its subsequent inhibition of the downstream signalling for Akt and glycogen synthase kinase-3β (GSK-3β)/cyclin D1 in H9c2 cells. Pre-treatment with EGCG or GSK-3β inhibitor (SB 216763) significantly improved the H 2 O 2 -induced suppression on cell viability, phosphorylation of pAkt (S473) and pGSK-3β (S9), and level of cyclin D1 in cells.
Conclusions
Collectively, these findings suggest that EGCG blunts the H 2 O 2 -induced oxidative effect on the Akt activity through the modulation of PIP3 synthesis leading to the subsequent inactivation of GSK-3β mediated cardiac cell injury.