Neuroprotective Effects And Mechanisms Of 20(R)-ginsenoside Rg₃ Against Rat Cerebral Ischemia-Reperfusion Injury

Objective To investigate the effects of 20(R)-ginsenoside Rg₃ on neuronalapoptosis and expression of calpain and caspase-3 mRNA in hippocampus CA1region after focal cerebral ischemia reperfusion in rat, and to elucidate theneuroprotective mechanisms of 20(R)-ginsenoside Rg₃ in the level of apoptoticregulation.Methods The model of focal ischemic reperfusion in Sprague-Dawley (SD) ratswas prepared by middle cerebral artery occlusion (MCAO) with a nylonmonofilament suture. Adult male SD rats were divided randomly into shamoperation group, model group, dissolvant group, 5, 10 and 20 mg/kg20(R)-ginsenoside Rg₃ groups, and 1 mg/kg nimodipine as positive control group,respectively. Based on different reperfusion time, each group was divided into 12 h-,24 h-and 72 h-reperfusion groups, respectively. The solvent (model group,dissolvant group and sham operation group) and all the above drugs wereintraperitoneally injected into rats at the volume of 1 ml/100 g body weight, twice aday. The behavioral disturbance was evaluated according to neurological deficitscores; the infarction volumes were evaluated by 2,3,5-Triphenyltetrazolium chloride(TTC) staining; the apoptosis in situ labeling was detected by terminaldeoxynucleotidyl transferase mediated d-UTP nick end labeling staining (TUNEL)and the expression of calpain and caspase-3 mRNA in hippocampus CA1 region wereassayed using in situ hybridization. Results Rg₃ (5, 10 and 20 mg/kg) could not improve the neurological deficit scoresof ischemia-reperfusion rats at 12 h; after 24 h and 72 h of ischemia-reperfusion,20(R)-ginsenoside Rg₃ significantly ameliorated the behavioral disturbance of rats ina dose-dependent manner and the curative effect of Rg₃ at high and middle doses wasnearly similar to that of nimodipine. It was obvious that high and middle doses ofRg₃ could reduce cerebral infarct volumes and degrade infarct rate of TTC-stainedcoronal brain sections from rats subjected to MCAO 2 h followed by 24 h ofreperfusion. TUNEL-positive cells were seldom expressed in sham operation groupand the number of apoptotic cells in model and dissolvent groups reached theirs peakvalue at 24 h in cerebral ischemia-reperfusion injury. Rg₃ markedly reduced theapoptotic cell number of neuron in a dose-dependent manner and the curative effect ofRg₃ at high and middle doses was equal to that of nimodipine. There were minorityexpressions of calpain I and caspase-3 mRNA at sham operation group, the mRNAexpressions of calpain I and caspase-3 of model and dissolvent groups, however,reached theirs peak values at 24 h in cerebral ischemia-reperfusion injury. Rg₃significantly suppressed the expression of calpain I and caspase-3 mRNA in adose-dependent manner, showing similar curative effect to that of nimodipine group.Conclusion These results indicated that 20(R)-ginsenoside Rg₃ attenuates theneuronal apoptosis due to cerebral ischemia-reperfusion injury. Inhibition of calpainand caspase-3 mRNA expression might be responsible for its neuroprotection.These findings suggested that the use of 20(R)-ginsenoside Rg₃ is of potentialtherapeutic application in human brain ischemial injuries.