Protective effects of ginsenoside Rb3 on oxygen and glucose deprivation-induced ischemic injury in PC12 cells

Aim:

To investigate the protective effects of ginsenoside Rb₃, a triterpenoid saponin isolated from the leaves of Panax notoginseng, on ischemic and reperfusion injury model of PC12 cells and elucidate the related mechanisms.

Methods:

PC12 cells exposed to oxygen and glucose deprivation (OGD) and restoration (OGD-Rep) were used as an in vitro model of ischemia and reperfusion. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and lactate dehydrogenase (LDH) leakage were used to evaluate the protective effects of ginsenoside Rb₃. Cellular apoptosis and mitochondrial membrane potential (MMP) were analyzed using flow cytometry. Intracellular calcium ion concentration ([Ca²⁺]_i) was detected using fluorophotometer system. Caspase-3, -8, and -9 activities were measured using assay kits with an ELISA reader. Western blotting assay was used to evaluate the release of cytochrome c and expression of caspase-3, Bcl-2 and Bax proteins.

Results:

It was shown that ginsenoside Rb₃ (0.1–10 μmol/L) significantly increased cell viability and inhibited LDH release in a dose-dependent manner on the ischemic model. In addition, ginsenoside Rb₃ also significantly inhibited ischemic injury-induced apoptosis, [Ca²⁺]_i elevation, and decrease of MMP. Meanwhile, pretreatment with ginsenoside Rb₃ significantly induced an increase of Bcl-2 protein expression and a decrease of cytosolic cytochrome c, cleaved-caspase 3 and Bax protein expression, the caspase-3, -8, and -9 activity were also inhibited.

Conclusion:

The results indicated that ginsenoside Rb₃ could markedly protected OGD-Rep induced ischemic injury and the mechanisms maybe related to its suppression of the intracellular Ca²⁺ elevation and inhibition of apoptosis and caspase activity. Ginsenoside Rb₃ could be a promising candidate in the development of a novel class of anti-ischemic agent.