Protective Effect And Mechanism Of Salidroside On Neuronal Mitochondrial Function Of Oxygen-Glucose-Deprived Neurons Via Optimizing Mitochondrial Quality Control

Background:Ischemic stroke(IS)is a life-threatening disease where sudden narrowing or occlusion of the blood supply arteries to the brain happens.Ischemic stroke is with the highest rate of death and disability worldwide.Related studies have confirmed that mitochondrial dysfunction is one of the most important mechanisms leading to neuronal damage in IS.Disruption of mitochondrial quality control(including mitochondrial dynamic imbalance,excessive mitophagy and mitochondrial calcium overload)eventually leads to neuronal death when neurons suffering from ischemia and hypoxia.Therefore,optimization of mitochondrial quality control is essential to alleviate neuronal damage in the IS state.Salidroside is the main active ingredient extracted from the traditional Chinese medicine Rhodiola rosea,which has antioxidant,neuronal protection and cognitive impairment improvement effects.Hence,it is interesting to ask whether salidroside can exert protective effects on neurons by optimizing mitochondrial quality control?Objective:In-depth investigation of the protective mechanism of salidroside in alleviating Oxygen-Glucose-Deprived(OGD)-induced neuronal injury.Methods:Cellular and animal models of ischemic stroke were constructed,intervened with salidroside,and the survival rate of neurons under the model conditions was examined by tissue section staining and CCK-8.We also used AMPK activator,inhibitor and siRNA to intervene in neurons under OGD conditions,and elucidated the protective effects of salidroside on mitochondria of oxygen-glucose deprived neurons in vitro and in vivo by Western blot,laser confocal microscopy,transmission electron microscopy and Real-time PCR.Results:In animal models,salidroside effectively reduced the infarct area of MCAO mice,preserved the number of neurons,restored neuronal mitochondrial ultrastructure.In the OGD cell model,salidroside significantly increased the survival rate of OGD neurons;restored the mitochondrial function of OGD neurons.Salidroside can bind directly to AMPK in vitro.By co-administrating Compound C or AICAR with salidroside,it was found that salidroside alleviated the imbalance of neuronal mitochondrial dynamics by inhibiting AMPK activity,decreasing the expression of mitochondrial fission protein in OGD neurons,increasing the expression of mitochondrial fusion proteins,and finally alleviating mitochondrial fragmentation in OGD neurons.In addition,salidroside alleviated excessive mitophagy in OGD neurons dependent on inhibition of AMPK activity,however,salidroside alleviated mitochondrial calcium overload was not dependent on AMPK signaling pathway.Salidroside reduced the area of mitochondria-associated membrane contact sites and decreased GRP75 expression in OGD neurons.Knockdown of HIF1α with siRNA significantly reduced GRP75 expression and alleviated mitochondrial calcium overload.Salidroside directly bound to HIF1α and promoted its degradation in OGD neurons,and simultaneously reduced HIF1α entry into the nucleus,inhibiting its transcriptional activity,decreasing GRP75 expression and alleviating mitochondrial calcium overload in OGD neurons.Conclusion:Salidroside alleviates mitochondrial damage in OGD neurons by optimizing mitochondrial quality control through AMPK-dependent and independent pathways.Salidroside directly binds to AMPK to alleviate the imbalance of mitochondrial dynamics in OGD neurons and inhibit excessive mitophagy.In addition,Salidroside increases HIF1αubiquitination and binds to it to reduce its entry into the nucleus,regulates its transcriptional activity,inhibits GRP75 expression,and alleviates mitochondrial calcium overload in OGD neurons...