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Inhibition of Autophagy Attenuated Cell Damage after OGD/R in SH-SY5Y Cells by Down-Regulating AMPK-Mediated Autophagy Signaling Pathway

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Document pages: 12 pages

Abstract: Background: Activation of autophagy becomes a new therapeutic target for the treatment of stroke. The aim of this study was to observe the role of autophagy in the model of glucose-oxygen deprivation reoxygenation (OGD R) in SH-SY5Y cells by interfering with AMPK-mediated autophagy signaling pathway. Methods: The effects of autophagy on OGD R injury in SH-SY5Y cells were investigated by evaluating cell viability and morphologic change using Cell Counting Kit-8 (CCK-8) and inverted microscop, respectively. To investigate whether autophagy played a role via the AMPK-mTOR signaling pathway, the levels of phospho-AMPK, phos-pho-mTOR, ULK1, microtubule-associated protein 1 light chain 3 (LC3) and Beclin 1 were detected using Western blot. In order to detect changes in autophagic flow after OGD R, autophagic flux (number of autophagosomes and autophagosomes) was detected by tandem stably expressed a tandem GFP-mRFP-LC3 construct. Results: Autophagy was low in the control group. After OGD R, the expression of autophagy-related proteins LC3 and Beclin 1 and autophagic flux increased, phospho-AMPK and ULK1 expression increased and phospho-mTOR expression decreased, meanwhile abnormal cell morphological changes increased significantly and cell viability decreased. Inhibition of AMPK activity, the levels of phospho-mTOR, ULK1, LC3, Beclin1 and the expression of autophagic flux were opposite to those of the model group; cell abnormal morphology and structure improved, and cell viability increased. Conclusion: Inhibition of autophagy can attenuate cell damage after OGD R in SH-SY5Y cells by down-regulating AMPK-mediated autophagy signaling pathway.

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