Protective Effect of Buyang Huanwu Decoction on Cerebral Ischemia/Reperfusion Injury by Inhibiting Autophagy Based on AMPK/UIK1Signaling Pathway

摘要

The aim of the paper is to explore whether AMPK/U1K1 signaling pathway is involved in regulating autophagy and alleviating cerebral ischemia/reperfusion(I/R)injury.On this basis,study the regulatory effect of Buyang Huanwu Decoction(BHD)on AMPK/U1K1 signaling pathway,aiming to clarify the molecular mechanism of BHD in alleviating cerebral I/R injury and the protective effect of BHD containing serum(BHDS)on OGD/R neurons.The research group adopted the following methods:Neurons cultured for 7 days in vitro were randomly divided into normal control group,model group(OGD/R),AMPK activator(AICAR)group(3 mmol/L),Buyang Huanwu Decoction medicated serum(BHDS)group(3%),Buyang Huanwu Decoction medicated serum+AMPK activator(BHDS+AICAR)group.The protein expressions of p-AMPK,AMPK,p-UlKl,U1K1,Beclin-1 and LC3B in neurons were detected by Western blot,the fluorescence intensity and apoptosis rate of neuron autophagosome CYTO-ID staining were detected by flow cytometry,the fluorescence intensity and morphological changes of neuron LC3B were detected by immunofluorescence double labeling method,the morphological changes of neurons were observed by inverted microscope,and the survival rate of neurons was detected by CCK-8 method.Neuron OGD/R model and the optimal concentration of BHDS.The neurons in the model group were subjected to hypoxia and glucose deprivation for 1 h,2 h,3 h,4 h and reoxygenation and glucose deprivation for 24 h.The survival rate of neurons and the release rate of 1DH to determine whether the model was successfully established.And the results show that BHDS group,the survival rate of OGD/R neurons was significantly increased,and the apoptosis rate was significantly decreased.Among them,3%BHDS intervention at the same time of hypoxia glucose deprivation for 3 h and reoxygenation glucose for 24 h was the best condition.In BHDS group,the levels of p-AMPK/AMPK,p-UlKl/UlKl,beclin-1,CYTO-ID and LC3B fluorescence intensity,and the transformation from LC3B-I to LC3B-II of OGD/R neurons were significantly decreased.So we can draw a conclusion that BHD can inhibit autophagy and alleviate cerebral I/R injury through AMPK/U1K1 signaling pathway.