Exploring The Mitochondrial Mechanisms Underlying The Neurotoxicity And The Pathological Damage Of Alzheimer’s Disease Caused By Copper Exposure

BACKGROUND AND AIMSAlzheimer’s disease(AD),was one of the most common neurodegenerative disease,which was characterized by learning and cognitive dysfunction,personality and behavior changes in clinically.With the advent of the aging society,the incidence of AD was rising sharply.Excessive copper intake could lead to neurotoxicity and accelerate the deterioration of neurodegenerative disease.However,the neurotoxicity and potential mechanism of copper was still unclear.In this study,we investigate the effects of copper exposure on neurotoxicity and mitochondrial function in AD model by using proteomics technology in vivo and vitro,and to screen out the key molecules and biomarkers and analyze the potential links.METHODS1.N2a-WT and N2a-APPswe cells were treated with increased doses of CuCl2(0,15,30,60μmol/L)for 24h.The protein levels of SDHB and COX5A were quantified by western blot analysis.2.N2a-WT and N2a-APPswe cells were pretreated with Melatonin(10μmol/L)for 2h,and then treated with CuCl2(60μmol/L)for 24h to extract proteins.The differential proteins were analyzed by Q Exactive mass spectrometry.Western blotting was used to verify the expression of these proteins and to explore the relationship of them.3.Five-month-old WT and APP/PS1-AD mice were fed with drinking water of 0.13 ppm copper chloride for 2 months,and which without copper exposure were used as the control group.Meantime,melatonin(10mg/kg)were administrated orally once every day for 2 months.Y maze test and Morris water maze test were used to evaluate the learning and memory function at the end of the experiment.4.T-AOC,MDA,GSH and CuZn-SOD/T-SOD levels were used to verify the damage of copper exposure on the oxidative stress function of cortex tissues of WT mice and APP/PS1-AD mice.5.Western blotting was used to detect the expression of apoptosis,antioxidant and electron transport chain(ECT)complex related proteins,and to evaluate the effects of copper exposure and melatonin treatment on mitochondrial biogenesis signaling pathway.RESULTS1.Compared to the control group,with the increase dose of copper exposure,the function of mitochondrial electron transport chain were decreased and the oxidative respiration was blocked in N2a-WT and N2a-APPswe mice.2.Proteomics analysis showed that the oxidative phosphorylation pathway and electron transport chain pathway were changed abnormally after copper exposure.3.The Y maze and Morris water maze behavioral results demonstrated that the learning and memory function were impaired after copper exposure while melatonin treatment improved behavioral dysfunction.4.The levels of oxidative stress signaling molecules were altered greatly after copper exposure,meanwhile,melatonin treatment blocked the overactivation of oxidative stress.5.Western blotting confirmed that the expression of apoptotic,antioxidant and ETC marker proteins were impaired in the cerebral cortex of WT mice and APP/S1AD mice and melatonin treatment reversed the abnormal things.CONCLUSIONSShort-term copper exposure could lead to learning and memory dysfunction in mice,and its mechanism of which may be related to the homeostasis of copper-induced oxidative stress and the inhibition of mitochondrial oxidative respiration.Meanwhile,proteomics studies have found that short-term copper exposure can induce redox and Mitochondrial signaling pathway dysfunction.