Electroacupuncture pretreatment reduces cerebral ischemia/reperfusion injury through inhibiting the AMPK-Beclin1/Vps34 pathway-mediated autophagy

Abstract:

Objective：To explore the effect of electroacupuncture（EA） pretreatment on cerebral ischemia/reperfusion（I/R） injury in rats and its relevant mechanism. Methods：Seventy-five healthy adult male Sprague-Dawley rats were randomly divided into Sham group，I/R group，EA+I/R group，corpus striatum infection with adeno-associated virus-vector control（AAV-VC，an empty virus vector）+EA+I/R group，and corpus striatum infection with adeno-associated virus-Beclin1（AAV-Beclin1，a Beclin1-overexpressing virus）+EA+I/R group，with 15 rats in each group. The neurological function scores of the rats in each group were evaluated 1，2，and 3 days after a middle cerebral artery occlusion（MCAO）；the rats were sacrificed 7 days after the MCAO，and their cerebral infarct volumes were measured by TTC staining；cerebral cell apoptosis was evaluated by TUNEL staining；the protein expression levels of the autophagy-related molecules LC3-Ⅰ，LC3-Ⅱ，AMPK，and Beclin1 were determined by Western blot；the interaction between Beclin1 and Vps34 was evaluated by co-immunoprecipitation assay. Results：Compared with the Sham group，the I/R group showed significant increases in cerebral infarct volume and neurobehavioral score（both P<0.05）. The mechanism research showed a significantly increased LC3-Ⅱ/LC3-Ⅰ ratio and up-regulat－ed expression of the autophagy regulatory proteins AMPK and Beclin1（all P<0.05） with increased formation of Beclin1 and Vps34 complex after I/R treatment；meanwhile，the number of apoptotic cells increased significantly. The above indices were significantly improved after EA pretreatment（all P<0.05）. However，intracerebral overexpression of the key autophagy-related protein Beclin1 prior to EA pretreatment reversed the protective effect of EA pretreat－ment against I/R injury（P<0.05）. Conclusion：EA pretreatment may play a neuroprotective role through inhibiting the AMPK-Beclin1/Vps34 pathway to reduce autophagy.