The Role Of Endoplasmic Reticulum Chaperone GRP78in Ischemic Preconditioning-induced Autophagy Activation In Neural Cells

Aim: Our previous study showed that the brain ischemic preconditioning(IPC)mediates neuroprotection through endoplasmic reticulum (ER) stress induced autophagy.This study is aimed to explore the role of ER chaperone GRP78in IPC inducedautophagy activation in neural cellsMethods: IPC and oxygen glucose deprivation (OGD) model was established inrat pheochromocytoma(PC12) cells and primary cultured murine cortical neurons. Thecells were treated with BAPTA(dibromo-1,2-bis(aminophenoxy)ethane N,N,N9,N9-tetra acetic acid，0.125-2μM), small interfering RNA targeted GRP78and lentiviralvector mediated GRP78overexpression. Cell Counting Kit-8was used to determine cellviability. Immunoblotting and immunoflurescence was used to examine the proteinexpression of GRP78, LC3, Beclin1, p-AMPK, p-mTOR and p-p70S6K1.Transmissionelectron microscope was used to exmaine the formation of autophagosomes after IPC.Results: IPC exerts neuroprotection against subsequent OGD injury in both PC12cells and primary cultured cortical neurons. IPC upregulates GRP78and activatesautophagy simultaneously, as evidenced by upregulated LC3and Beclin1, and increasedformation of autophagosomes. BAPTA and GRP78siRNA abrogated IPC inducedneuroprotection, and decreased the expression of GRP78, LC3, Beclin1and p-AMPK,but increased p-mTOR expression. By contrast, GRP78overexpression protected PC12cells from OGD injury. GRP78overexpression also increased LC3, Beclin1andp-AMPK expression, but decreased p-mTOR, p-p70S6K expression. Importantly,GRP78knockdown abolished the LC3, Beclin1and p-AMPK upregulation induced byGRP78overexpression.Conclusion: IPC upregulates ER molecular chaperon GRP78and activatesautophagy in neural cells. Inhibition and interference of GRP78expression abolished the neuroprotection of IPC and inhibits autophagy. GRP78overexpression activatesautophagy through AMPK-mTOR pathway, and hence exerts protection againstischemic like injury in neural cells. These results suggest GRP78contributes toautophagy activation and neuroprotection of ischemic preconditioning（IPC) in neuralcells.