Tau Protein Of Alzheimer's Disease Model Rats. Phlegm Law Abnormal Phosphorylation Regulation Research

ObjectiveTo establish the AD rat model induced by injection of basal nucleusof Meynert (BNM) with ibotenic acid(IBO) and 25-35β-amyloidpeptide(Aβ_25-35). And to explore the effect and potential mechanism ofreplenishing kidney-essence,removing phlegm therapy on tau proteinhyperphos-phorylation for AD.Methods15-month wistar rats(n=70) were randomly divided into sevengroups:normal group,blank group,AD model group, Huperzine A-ZhulinAntun treated group,high-dose,middle-dose and low-dose of BCHTgroups.There were ten rats in every group.To induce AD,five groups ratsreceived injection of IBO and Aβ_25-35 with basal nucleus of Meynert(BNM) through stereospeetific instrument. After treating for two weeks,BuShen HuaTan decoction were administered by oral gavage for the threedoses groups. Huperzine A-Zhulin Antun tablets served as control drug.Normal group, model group and blank group were administered byisovolumetric saline water.After treating for four weeks,Morris water maze was used to observethe ability of space learning and memory. Observe neurofibrillary tangles(NFT) of brain tissue through silver staining, senile plaque throughmethanol Congo red, ultramicro-pathological changes in the area of hippocampus were observed by transmission electron microscope(TEM)and radioimmunoassay was used to assay the activity of acetylcholine, acetylcholinesterase and choline acetyl transferase.Western blot wasused to detect volume dose of tau protein phosphorylation in hippo-campus homogenate, RT-PCR was used to detect the activity of cyclin-dependent kinase 5(cdk5), hybridization in situ was used to observe thechange of calciumlcalmodulin-dependent protein kinase 11 (CaMKⅡ) inhippocamp. enzyme linked immunosorbent assay was used to glycogensynthase kinase 3β(GSK-3β), to observe their effects on the abovemarks, we analyze the therapeutic mechanism of BSHT decoction inexperimental AD rats.Results1. In the behavior experiment, average incubation period of HuperzineA-Zhulin Antun treating group and high-dose,middle-dose andlow-dose groups have obvious shorten compared to model group(p＜0.01); Percen- tage of quadrant of high-dose,middle-dose andlow-dose groups have obvious increased compared to model group(p＜0.01), 20％and 40％area have obvious decreased compared bymodel group(p＜0.01), in space research of model group ,the frequencyin span of platform localization and percentage of area have obviouslydecerased compared to normal and blank group(p＜0.01). However,high-dose,middle-dose and low-dose groups have obviously increased compared to model groups (p＜0.01), and have a little dose-effectrelationship.2. The result of electron and light microscope have approved that thevolume of neuron of hippocampus decreased and denaturated in modelgroup and replenishing kidney-essence,removing phlegm therapyapproved those pathological change3. The activity of Ach and ChAT in hippocampus cortical area was lowerin AD model rats than that of other group's rats obviously(p＜0.01), but the activity of AchE climbed sharply. There is nosignificant difference between the model and normal group (p＞0.05);the significant differences were observed between the model groupand the high-, middle-, low-treated group of BSHT (p＜0.01). and nodifference with high-, middle- treated group(p＞0.05).There is nodifference between high-treated group and middle-treatedgroup(p＞0.05).4. The level of tau phosphorylation has no obvious difference betweennormal group and blank group (P＞0.05), the model group was prior toother groups(p＜0.01). Meanwhile,the expression of cdk5 mRNA,CaMKII-α, GSK-3βin model group have marked increased thannormal group(p＜0.01). After administrated by BSHT decoction, notnoly the total level of phosphorylation tau protein, but also theexpression of cdk5 mRNA,CaMKII-α, GSK-3βhave remarkedly increased, especially the high-treated group .Moreover,the high-treated,middle- treated and low-treated groups have dose-effect relationship(p＜0.01).Conclusion1. Injection of IBO and Aβ_25-35 with basal nucleus of Meynert (BNM)establish the AD model rat better simulate AD characteristic behaviourand pathological change,so they can be regarded as AD model rats.2. Replenishing kidney-essence and removing phlegm therapy canimprove remarkably disturbance learning and momory of AD ratsmodel.3. Replenishing kidney-essence and removing phlegm therapy canincrease the activity of Ach and ChAT, decrease the activity of AchE inorder to enhance the role of cholinergic neuron4. Replenishing kidney-essence and removing phlegm therapy canimprove pathomorphism change of hippocampus brain tissue in ADmodel rat.5. Replenishing kidney-essence and removing phlegm therapy caninhibit the activity ofcdk5 mRNA, CaMKII-α, GSK-3βand decreasethe level oftau protein hyper-phosphorylation.6. Kidney-essence weak and phlegm stasis is the basic pathogenesis forAD, and replenishing kidney-essence and removing phlegm therapy isoperative on treating AD.