| Alzheimer’s disease(AD),commonly known as senile dementia,is a chronic neurodegenerative disease.The pathogenesis of AD is extremely complex,which has not yet been fully elucidated and has no effective therapeutic drugs either,making it a thorny social and medical problem.AD is characterized by the extracellular senile plaques consisting of amyloid-β(A β)peptides,and intracellular neurofibrillary tangles(NFTs)formed by abnormally hyperphosphorylated tau(p-tau).For a long time,the Aβ amyloid cascade hypothesis has been predominant both in clinical drug development and basic research.However,the pathological changes of tau protein are more relevant to clinical symptoms than Aβ.Besides,with the successive failures of drugs development targeting Aβ,people’s attention has begun to turn to tau protein.There are two major domains in tau protein:the overhanging domain at the N-terminus and the microtubule binding domain at the C-terminus.The latter one determines the physiological and pathological roles of tau.The dynamic balance of metal ions including copper,iron,zinc,aluminum,etc.is closely related to the occurrence and development of AD,especially the process of Aβ aggregation to form senile plaques and the phosphorylated tau protein aggregation to form neurofibrillary tangles.In addition,metal ions are also related to oxidative stress,synaptic plasticity,neurotoxicity,autophagy and apoptosis.However,the effect of Zn2+on the third repeat unit of the microtubule-binding domain of tau(tau-R3)has not been reported before.Therefore,the first part of this thesis is to systematically study the influence of Zn2+ on tau aggregation and neurotoxicity at molecular,cellular and animal levels by a series of advanced experimental methods.The result pointed out that 1)tau-R3 bound 0.426 Zn2+per monomer with dissociation constants of 14.7 nM;2)Zn2+ accelerated tau-R3 aggregation and promoted the formation of shorter and smaller oligomers;3)More Zn2++R3 entered the cells compared with tau-R3,which maybe one of the reasons that Zn2++R3 was more toxic to neuronal cells;4)Zn2++R3 increased p-tau levels,damaged synaptophysin and dendritic spines;5)Zn2++R3 decreased mitochondrial membrane potential,damaged mitochondrial respiratory ability,and reduced the expression of mitochondrial energy metabolism-related proteins;6)Zn2++R3 triggered autophagy initiation of N2A cell line,but the downstream of the autophagy flow was blocked,causing an autophagy disorder.According to the above results,a sporadic AD mouse model was established by injecting Zn2++R3 into the brains of mice.Typical pathological features of AD and significant cognitive deficits were appeared 1.5 months after injection according to a variety of behavioral experiments.Interestingly,we found that Zn2++R3 had no effect on serum biochemical indicators except total cholesterol(TC/CHO)and blood glucose levels,which are two risk factors of AD.Therefore,it is expected to develop into a widely used sporadic AD mouse model.The second part of this thesis is to study the Chinese medicine formulas that can interfere with early AD.For a long time,there are only two kinds of drugs for AD in clinical,i.e.acetylcholinesterase inhibitors and glutamate receptor antagonists,but without satisfactory anti-AD effects.For a long time,famous pharmaceutical companies have invested heavily in researching new AD drugs,but almost all of them have failed after entering phase Ⅱ and Ⅲ clinical trials.Single targeting and the lack of appropriate animal models may be the main reasons for the failure of AD drug development.Although many published papers showed the potential of traditional Chinese medicines on AD treatment,still none of them that can clearly prevent and treat AD has been approved for clinical use.In our study,we selected the effective ingredients of traditional Chinese medicines that have been reported to have clear preventive and treatment effects on AD,and composed Chinese medicine formulas according to the principles of traditional Chinese medicine(Jun,Chen,Zuo and Shi),which can play a synergistic role.In order to reduce the toxic side effects of the formulas,the components used are mainly from the national list of "Medicine and Food Homologous".The four components were:Ginsenosides,Icariin,Curcumin(Piperine is added to increase the bioavailability of Curcumin),and Fisetin.We composed these single medicines into two groups of formulas:curcumin,piperine,icariin,and ginsenosides;and fisetin,piperine,icariin,and ginsenosides.There are two doses in each formula.APP/PS1 mice were fed with the formulas-containing diet from 4-mo-old to 8-mo-old,before their cognitive abilities and AD related pathological characteristics were studied.It was found that the two formulas decreased the content of Aβ oligomers,p-tau protein and neurofibrillary tangles,increased synapse-associated proteins levels and reduced neuronal death in the brain tissues,and thus improved the cognitive abilities of AD mice.Overall,the formulas containing Fisetin performed better than those containing Curcumin and higher-dose formulas were superior than low-dose ones.Interesting,all the formulas showed no liver and kidney toxicity but significantly decreased the blood lipid levels.The results indicated that these formulas provided a novel therapeutic idea for the prevention of AD. |
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