Study On The Mechanism Of Nrf2 Regulating Oxidative Stress In Astrocytes

Objective:Oxidative stress is involved in the pathogenesis of Alzheimer ’s disease(AD)and is associated with reactive oxygen species(ROS)and lipid peroxidation in neurotoxicity during AD onset.The role of nuclear factor erythroid 2-related factor 2(Nrf2),a major antioxidant response component,in AD brain has been demonstrated,but the mechanism by which Nrf2 regulates astrocyte iron death in AD remains unclear.Therefore,in this study,AD patients,APP/PS1/tau triple transgenic AD model(3×Tg)mice and mouse astrocytes were taken as research objects to explore the regulatory mechanism of silting Nrf2 on the key protein of ferroptosis in the oxidative damage process of astrocytes,aiming to provide a basis for further understanding of the pathogenesis of Alzheimer’s disease.Methods:We selected the brain sections of AD patients and 3×Tg mice as the research objects,and detected the expression changes of Nrf2 and NADPH Oxidase 4(NOX4)in astrocytes through immunofluorescence co-localization staining.Mouse astrocytes were infected with lentivirus to achieve targeted silencing of Nrf2 expression in vitro.The expression of Heme Oxygenase-1(HO-1),Glutathione Peroxidase 4(GPX4)and cystine/glutamate antiporter system light chain subunit(x CT)was detected by immunoblotting assay.The ROS changes in mouse astrocytes after Nrf2 silencing were detected by flow cytometry.The mitochondrial structure,lipid peroxidation degree,DNA damage degree and tyrosine nitration degree of mouse astrocytes after Nrf2 silencing were detected by immunofluorescence staining.Results:The level of Nrf2 protein in brain astrocytes of AD patients and 3×Tg mice was significantly decreased,and the level of ROS marker NADPH Oxidase 4(NOX4)induced by oxidative stress was significantly increased.Lentivirus-targeted silencing of Nrf2 expression in mouse astrocytes decreased the levels of HO-1 and GPX4,while increasing the levels of x CT.Silencing Nrf2 also increased ROS production and mitochondrial fragmentation in mouse astrocytes,as well as lipid peroxidation,DNA oxidative damage and tyrosine nitration.Conclusions:Our study shows that Nrf2 levels are significantly reduced in damaged astrocytes in AD patients and in a 3×Tg AD mouse model.NOX4 levels were significantly increased in damaged astrocytes from AD patients and 3×Tg AD mice.We found that inhibition of Nrf2 expression promoted oxidative stress-induced ROS production,mitochondrial damage and lipid peroxidation in astrocytes.In addition,inhibiting the expression of Nrf2 in mouse astrocytes can inhibit the expression of GPX4,a key protein that regulate ferroptosis.Our results support that Nrf2 plays a key role in the oxidative stress of astrocytes.Since oxidative stress plays an important role in the occurrence and development of AD,targeting Nrf2 activation in astrocytes may provide a new idea for the treatment of AD.