This study aims to investigate the therapeutic effect and mechanism of Daotan Xixin Decoction on APP/PS1 mice. Twelve APP/PS1 male mice were randomized into four groups: APP/PS1 and low-, medium-, and high-dose Daotan Xixin Decoction. Three C57BL/6 wild-type mice were used as the control group. The learning and memory abilities of mice in each group were examined by the Morris water maze test. The pathological changes of hippocampal nerve cells were observed by hematoxylin-eosin staining and Nissl staining. Immunohistochemistry was employed to detect the expression of β-amyloid(Aβ)_(1-42) in the hippocampal tissue. The high-dose Daotan Xixin Decoction group with significant therapeutic effects and the model group were selected for high-throughput sequencing. The differentially expressed gene(DEG) analysis, Gene Ontology(GO) analysis, Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment analysis, and Gene Set Variation Analysis(GSVA) were performed on the sequencing results. RT-qPCR and Western blot were conducted to determine the mRNA and protein levels, respectively, of some DEGs. Compared with the APP/PS1 group, Daotan Xixin Decoction at different doses significantly improved the learning and memory abilities of APP/PS1 mice, ameliorated the neuropathological damage in the CA1 region of the hippocampus, increased the number of neurons, and decreased the deposition of Aβ_(1-42) in the brain. A total of 1 240 DEGs were screened out, including 634 genes with up-regulated expression and 606 genes with down-regulated expression. The GO analysis predicted the biological processes including RNA splicing and protein folding, the cellular components including spliceosome complexes and nuclear spots, and the molecular functions including unfolded protein binding and heat shock protein binding. The KEGG pathway enrichment analysis revealed the involvement of neurodegenerative disease pathways, amyotrophic lateral sclerosis, and splicing complexes. Further GSVA pathway enrichment analysis showed that the down-regulated pathways involved nuclear factor-κB(NF-κB)-mediated tumor necrosis factor-α(TNF-α) signaling pathway, UV response, and unfolded protein response, while the up-regulated pathways involved the Wnt/β-catenin signaling pathway. The results of RT-qPCR and Western blot showed that compared with the APP/PS1 group, Daotan Xixin Decoction at different doses down-regulated the mRNA and protein levels of signal transducer and activator of transcription 3(STAT3), NF-κB, and interleukin-6(IL-6) in the hippocampus. In conclusion, Daotan Xixin Decoction can improve the learning and memory abilities of APP/PS1 mice by regulating the STAT3/NF-κB/IL-6 signaling pathway.
Animals ; Drugs, Chinese Herbal/administration & dosage* ; Mice ; Male ; Alzheimer Disease/metabolism* ; Computational Biology ; Mice, Inbred C57BL ; High-Throughput Nucleotide Sequencing ; Amyloid beta-Protein Precursor/metabolism* ; Hippocampus/metabolism* ; Mice, Transgenic ; Presenilin-1/metabolism* ; Humans ; Memory/drug effects* ; Maze Learning/drug effects* ; Amyloid beta-Peptides/genetics* ; Disease Models, Animal

The aim of this study is to investigate the regulation of Kaixin San-medicated serum(KXS-MS) on autophagy induced by Aβ_(25-35) in SH-SY5Y cells. The SH-SY5Y cell model of Aβ_(25-35)(25 μmol·L~(-1))-induced injury was established, and different concentrations of KXS-MS were added into the culture media of cells, which were then incubated for 24 h. Cell viability was measured by the methyl thiazolyl tetrazolium(MTT) assay. The protein levels of microtubule-associated protein 1 light chain 3(LC3)Ⅰ, LC3Ⅱ, protein kinase B(Akt), p-Akt, mammalian target of rapamycin(mTOR), and p-mTOR were assessed by Western blot. Furthermore, the combination of rapamycin(Rapa)/3-methyladenine(3-MA) and low concentration of KXS-MS was added to the culture medium of SH-SY5Y cells injured by Aβ_(25-35), and the cell viability and the expression levels of the above proteins were determined. The results showed that Aβ_(25-35) decreased the cell viability, up-regulated the expression levels of LC3Ⅱ and LC3Ⅱ/LC3Ⅰ, and down-regulated the expression levels of p-Akt, p-mTOR, p-Akt/Akt, and p-mTOR/mTOR. Compared with the Aβ_(25-35) model group, KXS-MS treatment attenuated Aβ_(25-35)-induced injury and enhanced the survival of SH-SY5Y cells. Meanwhile, KXS-MS down-regulated the LC3Ⅱ/LC3Ⅰ level and up-regulated the p-Akt/Akt and p-mTOR/mTOR levels. Compared with the low-concentration KXS-MS group, Rapa did not affect the cell survival and the levels of p-Akt and p-Akt/Akt, while it up-regulated the levels of LC3Ⅱ and LC3Ⅱ/LC3Ⅰ and down-regulated the levels of p-mTOR and p-mTOR/mTOR. 3-MA significantly reduced the cell survival rate and p-Akt, p-Akt/Akt level in the KXS-MS group, while it had no significant effect on the levels of LC3Ⅱ, LC3Ⅱ/LC3Ⅰ, p-mTOR, and p-mTOR/mTOR. The above results indicate that KXS-MS exhibits protective effects against Aβ_(25-35)-induced damage in SH-SY5Y cells by up-regulating Akt/mTOR activity to inhibit autophagy.
Humans ; Autophagy/drug effects* ; TOR Serine-Threonine Kinases/genetics* ; Amyloid beta-Peptides/toxicity* ; Proto-Oncogene Proteins c-akt/genetics* ; Drugs, Chinese Herbal/pharmacology* ; Cell Line, Tumor ; Cell Survival/drug effects* ; Peptide Fragments/toxicity* ; Microtubule-Associated Proteins/genetics*

Alzheimer's disease (AD) poses one of the most urgent medical challenges in the 21st century as it affects millions of people. Unfortunately, the etiopathogenesis of AD is not yet fully understood and the current pharmacotherapy options are somewhat limited. Here, we report a novel inhibitor, Compound 44, for targeting cholinesterases, amyloid-β (Aβ) aggregation, and glycogen synthase kinase 3β (GSK-3β) simultaneously with the aim of achieving symptomatic relief and disease modification in AD therapy. We found that Compound 44 had good inhibitory effects on all intended targets with IC₅₀s of submicromolar or better, significant neuroprotective effects in cell models, and beneficial improvement of cognitive deficits in the triple transgenic AD (3 × Tg AD) mouse model. Moreover, we showed that Compound 44 acts as an autophagy regulator by inducing nuclear translocation of transcription factor EB through GSK-3β inhibition, enhancing the biogenesis of lysosomes and elevating autophagic flux, thus ameliorating the amyloid burden and tauopathy, as well as mitigating the disease phenotype. Our results suggest that triple-target inhibition via Compound 44 could be a promising strategy that may lead to the development of effective therapeutic approaches for AD.
Animals ; Alzheimer Disease/genetics* ; Mice, Transgenic ; Glycogen Synthase Kinase 3 beta/metabolism* ; Disease Models, Animal ; Mice ; Amyloid beta-Peptides/metabolism* ; Cholinesterase Inhibitors/therapeutic use* ; Humans ; Autophagy/drug effects* ; Cognitive Dysfunction/pathology* ; Neuroprotective Agents/pharmacology*

Alzheimer's disease (AD) is a neurodegenerative disorder with an insidious onset, primarily characterized by a progressive decline in cognitive function. MCP-1 is a cytokine with chemotactic effects on monocytes, which can regulate their migration and infiltration and participate in disease progression. Increasing evidence suggests that MCP-1 plays a key role in the progression of Alzheimer's disease and has the potential to act as an early diagnostic marker and intervention target. This paper reviews the regulatory role of MCP-1 in neuroinflammation, beta-amyloid (Aβ) deposition and Tau pathology, and explores the potential of MCP-1 as a biomarker and intervention target for the early diagnosis of Alzheimer's disease.
Alzheimer Disease/metabolism* ; Humans ; Chemokine CCL2/genetics* ; Amyloid beta-Peptides/metabolism* ; Animals ; tau Proteins/metabolism* ; Biomarkers/metabolism*

Alzheimer's disease (AD) is a multifactorial and heterogenic disorder. MiRNA is a class of non-coding RNAs with 19-22 nucleotides in length that can regulate the expression of target genes in the post-transcriptional level. It has been found that the miRNAome in AD patients is significantly altered in brain tissues, cerebrospinal fluid and blood circulation, as compared to healthy subjects. Experimental studies have suggested that expression changes in miRNA could drive AD onset and development via different mechanisms. Therefore, targeting miRNA expression to regulate the key genes involved in AD progression is anticipated to be a promising approach for AD prevention and treatment. Rodent AD models have demonstrated that targeting miRNAs could block biogenesis and toxicity of amyloid β, inhibit the production and hyper-phosphorylation of τ protein, prevent neuronal apoptosis and promote neurogenesis, maintain neural synaptic and calcium homeostasis, as well as mitigate neuroinflammation mediated by microglia. In addition, animal and human studies support the view that miRNAs are critical players contributing to the beneficial effects of cell therapy and lifestyle intervention to AD. This article reviews the most recent advances in the roles, mechanisms and applications of targeting miRNA in AD prevention and treatment based on rodent AD models and human intervention studies. The potential opportunities and challenges in clinical application of targeting miRNA for AD patients are also discussed.
Animals ; Humans ; MicroRNAs/genetics* ; Alzheimer Disease/prevention & control* ; Amyloid beta-Peptides ; Apoptosis ; Microglia

This study aimed to investigate the mechanism of Psoraleae Fructus in improving the learning and memory ability of APP/PS1 mice by serum metabolomics, screen the differential metabolites of Psoraleae Fructus on APP/PS1 mice, and reveal its influence on the metabolic pathway of APP/PS1 mice. Thirty 3-month-old APP/PS1 mice were randomly divided into a model group and a Psoraleae Fructus extract group, and another 15 C57BL/6 mice of the same age were assigned to the blank group. The learning and memory ability of mice was evaluated by the Morris water maze and novel object recognition tests, and metabolomics was used to analyze the metabolites in mouse serum. The results of the Morris water maze test showed that Psoraleae Fructus shortened the escape latency of APP/PS1 mice(P<0.01), and increased the number of platform crossing and residence time in the target quadrant(P<0.01). The results of the novel object recognition test showed that Psoraleae Fructus could improve the novel object recognition index of APP/PS1 mice(P<0.01). Eighteen differential metabolites in serum were screened out by metabolomics, among which the levels of arachidonic acid, tryptophan, and glycerophospholipid decreased after drug administration, while the levels of glutamyltyrosine increased after drug administration. The metabolic pathways involved included arachidonic acid metabolism, glycerophospholipid metabolism, tryptophan metabolism, linoleic acid metabolism, α-linolenic acid metabolism, and glycerolipid metabolism. Therefore, Psoraleae Fructus can improve the learning and memory ability of APP/PS1 mice, and its mechanism may be related to the effects in promoting energy metabolism, reducing oxidative damage, protecting central nervous system, reducing neuroinflammation, and reducing Aβ deposition. This study is expected to provide references for Psoraleae Fructus in the treatment of Alzheimer's disease(AD) and further explain the mechanism of Psoraleae Fructus in the treatment of AD.
Mice ; Animals ; Amyloid beta-Protein Precursor/genetics* ; Mice, Transgenic ; Arachidonic Acid ; Tryptophan ; Mice, Inbred C57BL ; Alzheimer Disease/genetics* ; Maze Learning ; Glycerophospholipids ; Disease Models, Animal ; Amyloid beta-Peptides/metabolism*

Objective To explore the effect of knocking down Rho-associated coiled-coil kinase (ROCK2) gene on the cognitive function of amyloid precursor protein/presenilin-1 (APP/PS1) double transgenic mice and its mechanism. Methods APP/PS1 double transgenic mice were randomly divided into AD model group (AD group), ROCK2 gene knock-down group (shROCK2 group), ROCK2 gene knock-down control group (shNCgroup), and wild-type C57BL/6 mice of the same age served as the wild-type control (WT group). Morris water maze and Y maze were employed to test the cognitive function of mice. Neuron morphology was detected by Nissl staining. Immunofluorescence histochemical staining was used to detect the expression of phosphorylated dynamin-related protein 1 (p-Drp1) and mitochondrial fusion 1 (Mfn1). Western blot analysis was used to detect the expression ROCK2, cleaved-caspase-3 (c-caspase-3), B-cell lymphoma 2 (Bcl2), Bcl2-related protein X (BAX), p-Drp1, mitochondrial fission 1 (Fis1), optic atrophy 1 (OPA1), Mfn1 and Mfn2. Results Compared with AD group mice, the expression of ROCK2 in shROCK2 group mice was significantly reduced; the cognitive function was significantly improved with the number of neurons in the hippocampal CA3 and DG areas increasing, and nissl bodies were deeply stained; the expression of c-caspase-3 and BAX was decreased, while the expression of Bcl2 was increased; the expression of mitochondrial division related proteins p-Drp1 and Fis1 were decreased, while the expression of mitochondrial fusion-related proteins OPA1, Mfn1 and Mfn2 were increased. Conclusion Knock-down of ROCK2 gene can significantly improve the cognitive function and inhibit the apoptosis of nerve cells of APP/PS1 mice. The mechanism may be related to promoting mitochondrial fusion and inhibiting its division.
Animals ; Mice ; Alzheimer Disease/pathology* ; Amyloid beta-Peptides/metabolism* ; Amyloid beta-Protein Precursor ; Apoptosis/genetics* ; bcl-2-Associated X Protein ; Caspase 3 ; Cognition ; Disease Models, Animal ; Mice, Inbred C57BL ; Mice, Transgenic ; Mitochondrial Dynamics/genetics*

Humans ; Alzheimer Disease/genetics* ; Apolipoprotein E4/genetics* ; Amyloid beta-Peptides

OBJECTIVES: The liver X receptors (LXRs) are members of the nuclear hormone receptor superfamily, and LXR-β is an important receptor for cholesterol content in brain cells. LXR-β/retinoic X receptor (RXR-α)/ATP binding cassette transporter A1 (ABCA1) cholesterol transmembrane transport system is closely related to the occurrence and development of Alzheimer's disease (AD). LXR agonist TO901317 can affect the accumulation of β- amyloid protein in the brain tissue of APP/PS1 double transgenic AD mice. However, the molecular mechanism is not clarified in detail. This study aims to evaluate the effects of LXR agonist TO901317 on the cognitive function of AD mice fed with high cholesterol diet, and to explore its possible mechanism from the perspective of cholesterol metabolism. METHODS: Twenty four male 6-month-old APP/PS1 double transgenic AD mice were randomly divided into 4 groups, 6 mice in each group: a control group (fed with normal diet), a cholesterol rich diet (CRD) group, a TO901317 group (fed with CRD combined with TO901317), and a GSK2033 group (fed with CRD combined with TO901317 and LXR antagonist GSK2033). The mice were fed with pellet feed made of high cholesterol feed, mixed with lard, egg yolk powder, and cod liver oil twice a day. TO901317 and GSK2033 were dissolved and diluted to a final concentration at 0.03%. The drugs were given to the mice daily through gastric tube according to their body weight. Meanwhile, the mice in the drug group were fed with high cholesterol diet . After feeding for 3 months, Morris water maze was used to observe the changes of spatial exploration and memory ability of AD mice in each group. The contents of TC, LDL, and HDL in serum of mice in each group were detected by cholesterol enzyme colorimetry, and the differences among the groups were compared. The expression of Aβ₄₂ in the brain of AD mice was detected by ELISA. Western blotting was used to detect the protein levels of LXR-β, RXR-α, ABCA1, and Caveolin-1 in the brain of each group. RESULTS: Morris water maze results showed that the times, distance and the duration of mice crossing the platform in the CRD group were significantly decreased compared with the control group (all P<0.05), while these three figures in TO901317 group were significantly increased compared with the CRD group (all P<0.05). Compared with the TO901317 group, there was a decrease of these figures in the GSK2033 group (all P<0.05). The serum TC and LDL levels in the CRD group were significantly higher than those in the control group, while HDL levels were significantly lower (all P<0.001). The figures of the TC and LDL contents level in the TO901317 group were lower than those in the CRD group, while HDL levels were higher (all P<0.001). Compared with TO901317 group, the contents of the TC and LDL in GSK2033 group were significantly increased, while HDL content was significantly decreased (all P<0.001). ELISA results showed that the production of Aβ₄₂ peptides in the brain of CRD group was the highest while the content in the TO901317 group was significantly decreased (P<0.001), which was the lowest among the groups. The figure in the control group was close to the GSK2033 group. Western blotting results showed that the protein levels of LXR-β, RXR-α, and ABCA1 in the CRD group were significantly decreased compared with the control group, but the protein level of Caveolin-1 was increased (all P<0.01). After TO901317 treatment, the protein levels of LXR-β, RXR-α and ABCA1 were significantly increased, while the protein level of Caveolin-1 was decreased partially (all P<0.001). In the GSK2033 group, the effect of TO901317 on AD mice was partially reversed by GSK2033. Compared to TO901317 group, the protein levels of LXR-β, RXR-α, and ABCA1 showed a decrease trend, while the protein level of Caveolin-1 showed an increase state (all P<0.05). CONCLUSIONS High cholesterol diet leads to severer spatial exploration, learning and memory impairment in transgenic AD mice, while the LXR agonist TO901317 attenuates this effect. The mechanism may be that TO901317 promotes cholesterol efflux by activating LXR-β/RXR-α/ABCA1 transmembrane transport system, reduces the expression of Caveolin-1, improves the composition of lipid raft, and ultimately reduces the production of Aβ₄₂ in the brain.
Male ; Animals ; Mice ; Liver X Receptors/metabolism* ; Mice, Transgenic ; Alzheimer Disease/genetics* ; Caveolin 1/metabolism* ; Hydrocarbons, Fluorinated/pharmacology* ; Cognition ; Amyloid beta-Peptides/metabolism* ; Cholesterol

Objective To study the effects of TYRO protein kinase-binding protein (TYROBP) deficiency on learning behavior, glia activation and pro-inflammatory cycokines, and Tau phosphorylation of a new Alzheimer's disease (AD) mouse model carrying a PSEN1 p.G378E mutation.Methods A new AD mouse model carrying PSEN1 p.G378E mutation was built based on our previously found AD family which might be ascribed to the PSEN1 mutation, and then crossed with TYROBP deficient mice to produce the heterozygous hybrid mice (PSEN1^G378E/WT; Tyrobp^+/-) and the homozygous hybrid mice (PSEN1^G378E/G378E; Tyrobp^-/-). Water maze test was used to detect spatial learning and memory ability of mice. After the mice were sacrificed, the hippocampus was excised for further analysis. Immunofluorescence was used to identify the cell that expresses TYROBP and the number of microglia and astrocyte. Western blot was used to detect the expression levels of Tau and phosphorylated Tau (p-Tau), and ELISA to measure the levels of pro-inflammatory cytokines. Results Our results showed that TYROBP specifically expressed in the microglia of mouse hippocampus. Absence of TYROBP in PSEN1^G378E mutation mouse model prevented the deterioration of learning behavior, decreased the numbers of microglia and astrocytes, and the levels of interleukin-6, interleukin-1β and tumor necrosis factor-α in the hippocampus (all P < 0.05). The ratios of AT8/Tau5, PHF1/Tau5, pT181/Tau5, pT231/Tau5 and p-ERK/ERK were all higher in homozygous hybrid mice (PSEN1^G378E/G378E; Tyrobp^-/- mice) compared with PSEN1^G378E/G378E mice (all P < 0.05). Conclusions TYROBP deficiency might play a protective role in the modulation of neuroinflammation of AD. However, the relationship between neuroinflammation processes involving microglia and astrocyte activation, and release of pro-inflammatory cytokines, and p-Tau pathology needs further study.
Mice ; Animals ; Alzheimer Disease/genetics* ; Neuroinflammatory Diseases ; Hippocampus/pathology* ; Mutation ; Cytokines/pharmacology* ; Disease Models, Animal ; tau Proteins/pharmacology* ; Amyloid beta-Peptides/metabolism* ; Adaptor Proteins, Signal Transducing/pharmacology*