Aim:The aim of this study was to investigate whether Gs-Rbl relieves the CoCl 2 -induced apoptosis of hypoxic neonatal rat cardiomyocytes and in which the role of glucose transporter-4 (GLUT-4). Methods: 10, 50, 100, 200, 400, and 500 µmol/L), adenine 9-β-D-arabinofuranoside (ara A, 500 µmol/L; AMPK inhibitor) and wortmannin (0.5 µmol/L; PI3K inhibitor) only in combination with 200 µmol/L Gs-Rbl were administered in hypoxic cardiomyocytes, which were induced by 500 µmol/L CoCl 2 for 12 h. Then, the apoptotic rate (AR), 2-[ 3 H]-deoxy-D-glucose (2-[ 3 H]-DG) uptake, and the expression of GLUT-4 (including in plasma membrane, PM), phospho-AMPKα (Thr172), AMPKα and Akt in cells were assayed. Results: Compared with simple hypoxia (0 µmol/L Gs-Rbl), Gs-Rb1 greater than 10 µmol/L significantly decreased the apoptotic rate (P<0.01) and significantly increased 2-[ 3 H]-DG uptake (P<0.01), GLUT-4 content in cells and PM (P<0.01), AMPK activity (P<0.01) and Akt (P<0.01) levels in a dose-dependent manner. AMPK activity was completely suppressed by ara-A, just as Akt was suppressed by wortmannin. The AR, glucose uptake and GLUT-4 levels in cells and PM were partly down-regulated by ara-A or wortmannin. Conclusion: Gs-Rb1 may protect neonatal rat cardiomyocytes from apoptosis induced by CoCl 2 . The anti-apoptotic effect of Gs-Rb1 may occur by improving glucose uptake, in which GLUT-4 translocation and expression played a key role. Both the AMPK and the PI3K/Akt pathways may take part in the anti-hypoxic efficacy of Gs-Rb1.

Aim: To investigate whether mitochondria permeability transition pore (mPTP) opening was involved in ginsenoside Rb1 (Gs-Rb1) induced anti-hypoxia effects in neonatal rat cardiomyocytes ex vivo. Methods: Cardiomyocytes were randomly divided into 7 groups: control group, hypoxia group (500 µmol/L CoCl 2 ), Gs-Rb1 200 µmol/L group (CoCl 2 intervention+Gs-Rb1), wortmannin (PI3K inhibitor) 0.5 µmol/L group, wortmannin+Gs-Rb1 group, adenine 9-β-Darabinofuranoside (Ara A, AMPK inhibitor) 500 µmol/L group, and Ara A and Gs-Rb1 group. Apoptosis rate was determined by using flow cytometry. The opening of the transient mPTP was assessed by using co-loading with calcein AM and CoCl 2 in high conductance mode. Expression of GSK-3β, cytochrome c, caspase-3 and poly (ADP-ribose) polymerase (PARP) was measured by using Western blotting. ΔGSK-3β was defined as the ratio of p-Ser9-GSK-3β to total GSK-3β. Results: CoCl 2 significantly stimulated mPTP opening and up-regulated the level of ΔGSK-3β. There was a statistically significant positive correlation between apoptosis rate and mPTP opening, between apoptosis rate and ΔGSK-3β, and between mPTP opening and ΔGSK-3β. Gs-Rb1 significantly inhibited mPTP opening induced by hypoxia (41.3%±2.0%, P<0.001) . Gs-Rb1 caused a 77.3%±3.2% reduction in the expression of GSK-3β protein (P<0.001) and a significant increase of 1.182±0.007-fold (P=0.0001) in p-Ser9-GSK-3β compared with control group. Wortmannin and Ara A significantly inhibited the effect of Gs-Rb1 on mPTP opening and ΔGSK-3β. Gs-Rb1 significantly decreased expression of cytochrome c (66.1%±1.7%, P=0.001), caspase-3 (56.5%±2.7%, P=0.001) and cleaved poly ADP-ribose polymerase (PARP) (57.9%±1.4%, P=0.001). Conclusion: Gs-Rb1 exerted anti-hypoxia effect on neonatal rat cardiomyocytes by inhibiting GSK-3β-mediated mPTP opening.

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Gs-Rb1 may improve the viability of hypoxia cardiomyocytes by ameliorating cell autophagy via the upregulation of AMPK pathway.