DHA Attenuates Aβ-Induced Toxic Effect Of Pericytes By Inhibiting Excessive Mitophagy

BackgroundAlzheimer’s disease（AD）is a progressive neurodegenerative disease of the brain,the main clinical manifestations are cognitive dysfunction and progressive memory loss.Among the many AD pathological hypotheses,the mainstream view supports the"Aβcascade hypothesis",and with the deepening of research,the "oxidative stress and free radical damage hypothesis" has also been supported by more and more evidence.Pericyte is a pericyte cell in the blood vessel wall that secretes Docosahexaenoic acid（DHA）in capillaries and the basal membrane of arteries and plays an important role in absorbing and degrading macromolecules such as Aβ,regulating inflammation,and regulating blood flow.Pericytes can phagocytic Aβ_1-42 and degrade it into Aβ_1-34,which is not easy to polymerize,so as to facilitate the removal of Aβ by drainage through the perivascular lymphatic pathway.DHA is an unsaturated fatty acid that has strong antioxidant and anti-inflammatory properties and can easily cross the blood-brain barrier.Studies have shown that DHA can relieve the symptoms of cognitive impairment in AD mice,but the specific mechanism of DHA in AD caused by pericellular dysfunction is unknown and no relevant reports have been reported.ObjectiveThe aim of this study was to investigate the protective effect and mechanism of DHA against Aβ-induced pericellular neurotoxicity.MethodsAPP/PS1 transgenic AD mice and primary pericellular cells were selected as study objects to construct in vivo and in vitro models.In vivo models,behavioral tests were performed to detect cognitive function in mice.The contents of Aβ_1-42,pericellular coverage and co-localization of Aβ_1-42 were detected by immunofluorescence and immunohistochemistry.The expressions of apoptosis,senescence and autophagy related proteins were detected by Western-blot.In the in vitro model,cell purity was determined and treated with Aβ_1-42 oligomer and DHA to establish cell model.Senescent β-galactosidase staining and Annexin-V-FITC/PI flow cytometry were performed to detect peri-cell senescence and apoptosis.The expression of SASP inflammatory cytokines was detected by qPCR.The expression of apoptosis,senescence and mitochondrial autophagy related proteins were detected by WB.ROS kit,ATP kit and JC-1 kit were used to detect the function of pericellular mitochondria.In order to further investigate the relationship between the inhibition of apoptosis,senescence and mitochondrial autophagy induced by DHA,autophagy agonists and autophagy inhibitors were used to treat cells,and the changes of apoptosis and senescence levels as well as mitochondrial autophagy status were detected.Results1.Compared with the control group,the cognitive function of mice in AD group was decreased,and the DHA treatment group could improve the cognitive impairment of AD mice.2.Compared with the control group,the formation of Aβ_1-42 plaques in the brain of mice in AD group was increased,the coverage of pericellular cells was decreased,ROS and senescence levels were increased,which were relieved in DHA group compared with AD group.3.In WB and qPCR experiments,brain senescence（p53,p21）,apoptosis（Bax,Bcl-2）and mitochondrial autophagy（LC3II/LC3I,p62,Pinkl,Parkin）related proteins were increased in AD group,while brain related indexes were decreased in DHA group.4.Compared with the control group,the therapeutic dose of DHA had no toxic effect on primary pericytes;Compared with the Aβ group,DHA treatment could significantly inhibit the decline of cell viability induced by Aβ,and the effect was most obvious on the third day of treatment.5.Compared with the control group,Aβ promoted the expression of primary pericellular senescence,apoptosis and mitochondrial autophagy related proteins,and DHA treatment significantly inhibited the occurrence of cell senescence,apoptosis and mitochondrial autophagy.6.Compared with the control group,Aβ caused primary pericellular mitochondrial dysfunction（ROS increased,JC-1 decreased,ATP decreased）,while DHA treatment improved the damage of Aβ to pericellular mitochondrial function.7.Compared with DHA+Aβ group,rapamycin alleviated the inhibition of Aβinduced mitochondrial autophagy and weakened the protective effect of DHA on Aβinduced apoptosis and senescence of pericytes;In contrast,3-MA enhanced the protective effect of DHA on pericellular cells.ConclusionIn vivo models,DHA improved cognitive dysfunction and decreased the rate of Aβ production in the brain of AD mice.In vitro models,DHA can inhibit the toxic effect of Aβ_1-42 oligomer on primary pericytes,improve the activity of pericytes,inhibit cell apoptosis and senescence,and alleviate mitochondrial dysfunction by inhibiting the overactivation of mitochondrial autophagy pathway.