OBJECTIVE：To study the effects and mechanism of panaxadiol （PD） on Tau protein phosphorylation in the SH-SY5Y cells transfected with APP gene（APP-SH-SY5Y）. METHODS ：The target of PD and non-receptor tyrosine kinases Fyn was verified by molecular docking. SH-SY 5Y cells were cultured in vitro ，and the APP-SH-SY 5Y cell models and green fluorescent （GFP）-SH-SY5Y cell model （control cell ）was constructed. The expression of Aβ1-42 was detected so as to verify the success of APP-SH-SY5Y cell model. Taking GFP-SH-SY 5Y cells as control ，the effects of 5，10，20，30，40 μmol/L PD and 125，250， 500，1 000，2 000 nmol/L PP 2（Fyn inhibitor ，positive control ）on the survival rate of APP-SH-SY 5Y cells were detected by CCK-8 assay after treated for 24 h，so as to confirm the optimal concentration. The concentration of Ca 2 + ，the ratio fophosphorylated Tau protein （p-Tau）/Tau，phosphorylatedn Src（p-Src）/Fyn and phosphorylated glutamate receptor 2B（p-GluN2B）/ GluN2B were detected in APP-SH-SY 5Y cells after trated with the optimal concentration of PD and PP 2 for 24 h. RESULTS ：The results of molecular simulation docking showed that PD could target Fyn protein. Compared with GFP-SH-SY 5Y cells ，the protein expression of Aβ1-42 in APP-SH-SY 5Y cell were increased significantly （P＜0.01）. The optimal concentration of PD and PP 2 were 20 μmol/L and 500 nmol/L. The 20 μmol/L PD and 500 nmol/L PP 2 could increase the survival rate of the cells and reduced the concentration of Ca 2+，the ratio of p-Tau/Tau ，p-Src/Fyn，and p-GluN 2B/GluN2B. CONCLUSIONS：PD can reduce the the phosphorylation of Tau protein through inhibiting Fyn/GluN 2B signaling pathway.

OBJECTIVE To investigate the improvement effect of Dihuang yinzi on neurological function of APP/PS1 double transgenic positive mice and its possible mechanism. METHODS APP/PS1 double transgenic positive mice were randomly divided into APP/PS1 positive model group， Dihuang yinzi low-dose， medium-dose and high-dose groups （12， 24， 48 g/kg，by the amount of crude drug）， western medicine group （donepezil hydrochloride 0.8 mg/kg）； APP/PS1 transgenic negative mice were included in as APP/PS1 negative normal group， with 6 mice in each group. Administration groups were given relevant medicine intragastrically； APP/PS1 positive model group and APP/PS1 negative normal group were given constant volume of normal saline intragastrically， once a day， for consecutive 28 d. After the last medication， the ability of learning and memory， intestinal flora diversity （only medium-dose group of Dihuang yinzi）， pathological changes of neural cells in hippocampus， and the expressions of Bcl-2， Bax and brain-derived neurotrophic factor （BDNF） in mice of each group were investigated. RESULTS Compared with APP/PS1 positive model group， escape latency of mice was shortened significantly in Dihuang yinzi medium-dose and high-dose groups， while the times of crossing the platform significantly increased （P＜0.05）. Compared with APP/PS1 positive model group， Chao1 index of Dihuang yinzi medium-dose group was lower， while Shannon index was higher （P＜0.05）； OTU abundance of microbial flora was decreased to some extent； dominant flora was Clostridia o_ Clostridia_ vadinBB60_ Group， verruca G_ UCG_ 005. Compared with the APP/PS1 positive model group， the pathological changes of the nerve cells in the hippocampal CA1 area of mice were improved in the medium-dose and high-dose groups of Dihuang yinzi； the protein expressions of Bcl-2 and BDNF in the hippocampal tissue were significantly increased， while the protein expression of Bax was significantly decreased （P＜0.05）. CONCLUSIONS Dihuang yinzi can enhance the diversity of flora， change the type of dominant flora， and can inhibit the apoptosis of brain neurons， and increase the level of BDNF； its mechanism may be to protect nerve cells by means of gut-brain axis， thereby improving the learning and memory ability of APP/PS1 positive model mice.

OBJECTIVE: To investigate the effect of osthole on the expression of amyloid precursor protein (APP) in Alzheimer's disease (AD) cell model and its mechanism. METHODS: The SH-SY5Y cell with over expression of APP was established by transfection by liposome 2000. The cells were treated with different concentrations of osthole, and the cell viability was determined by MTT and lactate dehydrogenase (LDH) assay. The differentially expressed miRNAs with and without osthole treatment were detected by miRNA array, and the target genes binding to the differentially expressed miRNAs were identified and verified by databases and Cytoscape. After the inhibitor of the differentially expressed miRNA was transduced into cells, the changes of APP and amyloid β (Aβ) protein were determined by immunofluorescence cytochemistry, and the mRNA expression of APP was determined by RT-PCR. RESULTS: The AD cell model with over expression of APP was established successfully. The results of MTT and LDH assay showed that osthole had a protective effect on cells and alleviated cell damage. miR-101a-3p was identified as the differentially expressed miRNA, which was binding to the 3'-UTR of APP. Compared with APP group, the expression of APP and Aβ protein and APP mRNA increased in the miR-101a-3p inhibitor group (all <0.01), while the expression of APP and Aβ protein and APP mRNA decreased in the cells with osthole treatment (all <0.01). CONCLUSIONS Osthole inhibits the expression of APP by up-regulating miR-101a-3p in AD cell model.
Alzheimer Disease ; Amyloid beta-Peptides ; Amyloid beta-Protein Precursor ; genetics ; Cell Line ; Coumarins ; pharmacology ; Gene Expression Regulation ; drug effects ; genetics ; Humans ; MicroRNAs ; genetics ; metabolism