ABSTRACT
The multiple-step cleavage of amyloid precursor protein (APP) generates amyloid-β peptides (Aβ), highly toxic molecules causing Alzheimer's disease (AD). The nonspecific cleavage between the transmembrane region of APP (APPTM) and γ-secretase is the key step of Aβ generation. Reconstituting APPTM under physiologically-relevant conditions is crucial to investigate how it interacts with γ-secretase and for future AD drug discovery. Although producing recombinant APPTM was reported before, the large scale purification was hindered by the use of biological protease in the presence of membrane protein. Here, we expressed recombinant APPTM in Escherichia coli using the pMM-LR6 vector and recovered the fusion protein from inclusion bodies. By combining Ni-NTA chromatography, cyanogen bromide cleavage, and reverse phase high performance liquid chromatography (RP-HPLC), isotopically-labeled APPTM was obtained in high yield and high purity. The reconstitution of APPTM into dodecylphosphocholine (DPC) micelle generated mono dispersed 2D 15N-1H HSQC spectra in high quality. We successfully established an efficient and reliable method for the expression, purification and reconstruction of APPTM, which may facilitate future investigation of APPTM and its complex in more native like membrane mimetics such as bicelle and nanodiscs.
Subject(s)
Humans , Amyloid beta-Protein Precursor/chemistry , Micelles , Amyloid Precursor Protein Secretases/metabolism , Magnetic Resonance Spectroscopy , Recombinant ProteinsABSTRACT
Silent information regulator 1 (SIRT1) is one of the seven mammalian proteins of the sirtuin family of NAD+-dependent deacetylases. SIRT1 plays a pivotal role in neuroprotection and ongoing research has uncovered a mechanism by which SIRT1 may exert a neuroprotective effect on Alzheimer's disease (AD). Growing evidence demonstrates that SIRT1 regulates many pathological processes including amyloid-β precursor protein (APP) processing, neuroinflammation, neurodegeneration, and mitochondrial dysfunction. SIRT1 has recently received enormous attention, and pharmacological or transgenic approaches to activate the sirtuin pathway have shown promising results in the experimental models of AD. In the present review, we delineate the role of SIRT1 in AD from a disease-centered perspective and provides an up-to-date overview of the SIRT1 modulators and their potential as effective therapeutics in AD.
Subject(s)
Animals , Humans , Alzheimer Disease , Amyloid beta-Protein Precursor , Animals, Genetically Modified , Sirtuin 1 , SirtuinsABSTRACT
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.
Subject(s)
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/geneticsABSTRACT
OBJECTIVE@#To explore whether the protein Deglycase protein 1 (DJ1) can ameliorate Alzheimer's disease (AD)-like pathology in Amyloid Precursor Protein/Presenilin 1 (APP/PS1) double transgenic mice and its possible mechanism to provide a theoretical basis for exploring the pathogenesis of AD.@*METHODS@#Adeno-associated viral vectors (AAV) of DJ1-overexpression or DJ1-knockdown were injected into the hippocampus of 7-month-old APP/PS1 mice to construct models of overexpression or knockdown. Mice were divided into the AD model control group (MC), AAV vector control group (NC), DJ1-overexpression group (DJ1 +), and DJ1-knockdown group (DJ1 -). After 21 days, the Morris water maze test, immunohistochemistry, immunofluorescence, and western blotting were used to evaluate the effects of DJ1 on mice.@*RESULTS@#DJ1 + overexpression decreased the latency and increased the number of platform traversals in the water maze test. DJ1 - cells were cured and atrophied, and the intercellular structure was relaxed; the number of age spots and the expression of AD-related proteins were significantly increased. DJ1 + increased the protein expression of Nuclear factor erythroid 2-related factor 2 (NRF2), heme oxygenase-1 (HO-1), light chain 3 (LC3), phosphorylated AMPK (p-AMPK), and B cell lymphoma-2 (BCL-2), as well as the antioxidant levels of total superoxide dismutase (T-SOD), total antioxidant capacity (T-AOC), and Glutathione peroxidase (GSH-PX), while decreasing the levels of Kelch-like hydrates-associated protein 1 (Keap1), mammalian target of rapamycin (mTOR), p62/sequestosome1 (p62/SQSTM1), Caspase3, and malondialdehyde (MDA).@*CONCLUSION@#DJ1-overexpression can ameliorate learning, memory, and AD-like pathology in APP/PS1 mice, which may be related to the activation of the NRF2/HO-1 and AMPK/mTOR pathways by DJ1.
Subject(s)
Animals , Mice , Alzheimer Disease/therapy , AMP-Activated Protein Kinases/metabolism , Amyloid beta-Protein Precursor/metabolism , Antioxidants/metabolism , Disease Models, Animal , Hippocampus/metabolism , Kelch-Like ECH-Associated Protein 1/metabolism , Mammals/metabolism , Mice, Inbred C57BL , Mice, Transgenic , NF-E2-Related Factor 2/metabolism , Presenilin-1/metabolism , TOR Serine-Threonine Kinases/metabolismABSTRACT
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.
Subject(s)
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/metabolismABSTRACT
Mutations in genes encoding amyloid precursor protein (APP) and presenilins (PSs) cause familial forms of Alzheimer's disease (AD), a neurodegenerative disorder strongly associated with aging. It is currently unknown whether and how AD risks affect early brain development, and to what extent subtle synaptic pathology may occur prior to overt hallmark AD pathology. Transgenic mutant APP/PS1 over-expression mouse lines are key tools for studying the molecular mechanisms of AD pathogenesis. Among these lines, the 5XFAD mice rapidly develop key features of AD pathology and have proven utility in studying amyloid plaque formation and amyloid β (Aβ)-induced neurodegeneration. We reasoned that transgenic mutant APP/PS1 over-expression in 5XFAD mice may lead to neurodevelopmental defects in early cortical neurons, and performed detailed synaptic physiological characterization of layer 5 (L5) neurons from the prefrontal cortex (PFC) of 5XFAD and wild-type littermate controls. L5 PFC neurons from 5XFAD mice show early APP/Aβ immunolabeling. Whole-cell patch-clamp recording at an early post-weaning age (P22-30) revealed functional impairments; although 5XFAD PFC-L5 neurons exhibited similar membrane properties, they were intrinsically less excitable. In addition, these neurons received smaller amplitude and frequency of miniature excitatory synaptic inputs. These functional disturbances were further corroborated by decreased dendritic spine density and spine head volumes that indicated impaired synapse maturation. Slice biotinylation followed by Western blot analysis of PFC-L5 tissue revealed that 5XFAD mice showed reduced synaptic AMPA receptor subunit GluA1 and decreased synaptic NMDA receptor subunit GluN2A. Consistent with this, patch-clamp recording of the evoked L23>L5 synaptic responses revealed a reduced AMPA/NMDA receptor current ratio, and an increased level of AMPAR-lacking silent synapses. These results suggest that transgenic mutant forms of APP/PS1 overexpression in 5XFAD mice leads to early developmental defects of cortical circuits, which could contribute to the age-dependent synaptic pathology and neurodegeneration later in life.
Subject(s)
Mice , Animals , Alzheimer Disease/pathology , Amyloid beta-Peptides/metabolism , Receptors, N-Methyl-D-Aspartate/metabolism , Amyloid beta-Protein Precursor/metabolism , Mice, Transgenic , Neurons/metabolism , Receptors, AMPA/metabolism , Disease Models, AnimalABSTRACT
Resveratrol (RES), a natural polyphenolic phytochemical, has been suggested as a putative anti-aging molecule for the prevention and treatment of Alzheimer's disease (AD) by the activation of sirtuin 1 (Sirt1/Sir2). In this study, we tested the effects of RES and Sirt1/Sir2 on sleep and courtship memory in a Drosophila model by overexpression of amyloid precursor protein (APP), whose duplications and mutations cause familial AD. We found a mild but significant transcriptional increase of Drosophila Sir2 (dSir2) by RES supplementation for up to 17 days in APP flies, but not for 7 days. RES and dSir2 almost completely reversed the sleep and memory deficits in APP flies. We further demonstrated that dSir2 acts as a sleep promotor in Drosophila neurons. Interestingly, RES increased sleep in the absence of dSir2 in dSir2-null mutants, and RES further enhanced sleep when dSir2 was either overexpressed or knocked down in APP flies. Finally, we showed that Aβ aggregates in APP flies were reduced by RES and dSir2, probably via inhibiting Drosophila β-secretase (dBACE). Our data suggest that RES rescues the APP-induced behavioral deficits and Aβ burden largely, but not exclusively, via dSir2.
Subject(s)
Animals , Alzheimer Disease/metabolism , Amyloid beta-Peptides , Amyloid beta-Protein Precursor/metabolism , Drosophila/physiology , Drosophila Proteins/metabolism , Resveratrol/pharmacology , Sirtuin 1 , SleepABSTRACT
Deficits in the clearance of amyloid β protein (Aβ) by the peripheral system play a critical role in the pathogenesis of sporadic Alzheimer's disease (AD). Impaired uptake of Aβ by dysfunctional monocytes is deemed to be one of the major mechanisms underlying deficient peripheral Aβ clearance in AD. In the current study, flow cytometry and biochemical and behavioral techniques were applied to investigate the effects of polysaccharide krestin (PSK) on AD-related pathology in vitro and in vivo. We found that PSK, widely used in therapy for various cancers, has the potential to enhance Aβ uptake and intracellular processing by human monocytes in vitro. After administration of PSK by intraperitoneal injection, APP/PS1 mice performed better in behavioral tests, along with reduced Aβ deposition, neuroinflammation, neuronal loss, and tau hyperphosphorylation. These results suggest that PSK holds promise as a preventive agent for AD by strengthening the Aβ clearance by blood monocytes and alleviating AD-like pathology.
Subject(s)
Animals , Mice , Alzheimer Disease/pathology , Amyloid beta-Peptides/metabolism , Amyloid beta-Protein Precursor/metabolism , Cognition , Disease Models, Animal , Mice, Transgenic , Monocytes/pathology , Polysaccharides/therapeutic use , ProteoglycansABSTRACT
BACKGROUND@#Circular RNAs (circRNAs) are considered to be important regulators in cancer biology. In this study, we focused on the effect of circRNA baculoviral inhibitor of apoptosis protein (IAP) repeat containing 6 (circBIRC6) on non-small cell lung cancer (NSCLC) progression.@*METHODS@#The NSCLC and adjacent non-tumor tissues were collected at Shanghai Ninth People's Hospital. Quantitative real-time polymerase chain reaction was conducted for assessing the levels of circBIRC6, amyloid beta precursor protein binding protein 2 (APPBP2) messenger RNA (mRNA), baculoviral IAP repeat containing 6 mRNA (BIRC6), and microRNA-217 (miR-217). Western blot assay was adopted for measuring the protein levels of APPBP2, E-cadherin, N-cadherin, and vimentin. Colony formation assay, transwell assay, and flow cytometry analysis were utilized for evaluating cell colony formation, metastasis, and apoptosis. Dualluciferase reporter assay and RNA immunoprecipitation assay were carried out to determine the interaction between miR-217 and circBIRC6 and APPBP2 in NSCLC tissues. The murine xenograft model assay was used to investigate the function of circBIRC6 in tumor formation in vivo. Differences were analyzed via Student's t test or one-way analysis of variance. Pearson's correlation coefficient analysis was used to analyze linear correlation.@*RESULTS@#CircBIRC6 was overexpressed in NSCLC tissues and cells. Knockdown of circBIRC6 repressed the colony formation and metastasis and facilitated apoptosis of NSCLC cells in vitro and restrained tumorigenesis in vivo. Mechanically, circBIRC6 functioned as miR-217 sponge to promote APPBP2 expression in NSCLC cells. MiR-217 inhibition rescued circBIRC6 knockdown-mediated effects on NSCLC cell colony formation, metastasis, and apoptosis. Overexpression of miR-217 inhibited the malignant phenotypes of NSCLC cells, while the effects were abrogated by elevating APPBP2.@*CONCLUSIONS@#CircBIRC6 aggravated NSCLC cell progression by elevating APPBP2 via sponging miR-217, which might provide a fresh perspective on NSCLC therapy.
Subject(s)
Animals , Humans , Mice , Amyloid beta-Peptides/metabolism , Amyloid beta-Protein Precursor/metabolism , Carcinoma, Non-Small-Cell Lung/pathology , Cell Movement/genetics , Cell Proliferation/genetics , China , Gene Expression Regulation, Neoplastic/genetics , Lung Neoplasms/pathology , MicroRNAs/metabolism , RNA, Circular/genetics , RNA, MessengerABSTRACT
Objective: To uncover the time-dependent expression pattern of ptk2b gene and ptk2b-encoded protein, protein tyrosine kinase 2 beta(PTK2B), in the brain tissues of transgenic animal models of Alzheimer's disease (AD) and its relationship with the levels of Aβ1-42, phosphorylation of Tau (p-Tau) and low density lipoprotein receptor-related protein-1(LRP-1) in blood and brain tissues. Methods: In this study, 5-, 10- and 15-month-old APPswe/PS1dE9 double-transgenic mice harboring the genotype of AD confirmed by the gene test were divided into the 5-, 10- and 15-month-old experiment groups, and simultaneously, age-matched C57BL/6J mice were placed into the corresponding control groups, with 8 mice in each group. All mice were subjected to the Morris Water Maze for test of cognitive and behavioral ability. Expression profiles of PTK2B, Aβ1-42, p-Tau/Tau and LRP-1 in the hippocampus or blood of mice were quantified by using the immunohistochemistry staining, Western blot or enzyme-linked immunosorbent assay (ELISA), while the mRNA expression of ptk2b in the hippocampus was quantified by using the real-time quantitative polymerase chain reaction (qRT-PCR). Results: Results of experiment groups demonstrated that as mice aged, the expression levels of PTK2B, ptk2b mRNA, Aβ1-42 and p-Tau/Tau in the hippocampus were increased, and the expression of LRP-1 was decreased gradually. While in the blood, the level of Aβ1-42 was decreased, and the cognitive and behavioral ability was decreased in an age-dependent manner (all P< 0.05). However, comparisons among the control groups, only the age-dependent downregulation of LRP-1 were observed in hippocampus(P<0.05), but other indicators had no significant differences (P>0.05). Conclusion: In the hippocampus of APP/PS1 double-transgenic mice, the expressions of PTK2B, Aβ1-42 and p-Tau/Tau are upregulated, LRP-1 is downregulated, while cognitive and behavioral ability is decreased, and such changes are presented in a time-dependent manner.
Subject(s)
Animals , Mice , Alzheimer Disease/metabolism , Amyloid beta-Peptides , Amyloid beta-Protein Precursor/genetics , Focal Adhesion Kinase 2/metabolism , Hippocampus/metabolism , Low Density Lipoprotein Receptor-Related Protein-1 , Maze Learning , Mice, Inbred C57BL , Mice, Transgenic , RNA, MessengerABSTRACT
OBJECTIVE@#To compare the clinical efficacy of abdominal acupoint thread embedding therapy based on "brain-intestinal connection" combined with donepezil hydrochloride tablets and oral donepezil hydrochloride tablets alone for mild-to-moderate Alzheimer's disease (AD) and observe its effects on amyloid precursor protein (APP) and β-amyloid protein@*METHODS@#Sixty patients with AD were randomly divided into an observation group (30 cases, 3 cases dropped off) and a control group (30 cases, 3 cases dropped off). The patients in the control group were treated with donepezil hydrochloride tablets (5 mg per day); based on the treatment in the control group, the patients in the observation group were treated with abdominal acupoint thread embedding therapy at Zhongwan (CV 12), Xiawan (CV 10), Huaroumen (ST 24), Wailing (ST 26), Daheng (SP 15), etc., once every 10 days. Both groups were treated for 2 months. The mini-mental state examination (MMSE), Alzheimer's disease assessment scale-cognitive subscale (ADAS-Cog), activity of daily living scale (ADL), neuropsychiatric inventory questionnaire (NPI) as well as the serum levels of APP and Aβ@*RESULTS@#After treatment, the MMSE scores in the two groups were higher than those before treatment (@*CONCLUSION@#The abdominal acupoint thread embedding therapy based on the theory of "brain-intestinal connection" combined with donepezil hydrochloride tablets can improve cognitive function, self-care ability of daily life and mental behavior, and reduce the serum levels of APP and Aβ
Subject(s)
Humans , Acupuncture Points , Alzheimer Disease/drug therapy , Amyloid beta-Peptides , Amyloid beta-Protein Precursor , Brain , Donepezil , Peptide FragmentsABSTRACT
To investigate the effects of on behavior and blood brain barrier (BBB) in Alzheimer's disease mice. Thirty-eight 4-month-old APP/PS1 double transgenic mice were randomly divided into three groups: model group, low-dose group and high-dose group. Saline, and 12 g·kg·d were given to each group by continuous gavage once a day for respectively. The changes in activities of daily live and fear conditioning memory behavior of mice were examined by nesting behavior test and fear conditioning test, respectively. The β-amyloid protein (Aβ) depositions in cortex and hippocampal CA1 area of mice were detected by thioflavin T staining. The CD34 and activities fibrinogen (Fib) immunofluorescence double staining were used to determine the vascular endothelial integrity and BBB exudation. Compared with model mice, activities of daily live were significantly improved in low-dose and high-dose groups (both <0.01), the fear memory ability was significantly increased in high-dose group (<0.01). The amount of Aβ deposition in cortex and hippocampal CA1 decreased significantly in high-dose group, the area ratio decreased significantly; the area ratio of Aβ deposition in hippocampal CA1 region in low-dose group also decreased (all <0.05). The proportions of CD34 positive area of cortex in low and high dose groups increased, the percentage of fibrinogen positive area decreased (all <0.05). The proportion of CD34 positive area in hippocampal CA1 region in high-dose group was significantly increased, the percentage of fibrinogen positive area decreased significantly (both <0.05). especially high-dose can improve the activities of daily live and fear conditioning memory function of APP/PS1 mice, reduce the deposition of Aβ in brain. The mechanism may be related to the reduction of BBB permeability and the protection of the integrity of BBB.
Subject(s)
Animals , Mice , Alzheimer Disease , Amyloid beta-Protein Precursor , Blood-Brain Barrier/metabolism , Disease Models, Animal , Hippocampus/metabolism , Mice, Inbred C57BL , Mice, TransgenicABSTRACT
Sleep exerts important functions in the regulation of cognition and emotion. Recent studies have found that sleep disorder is one of the important risk factors for Alzheimer's disease (AD), but the effects of chronic sleep deprivation on the cognitive functions of AD model mice and its possible mechanism are still unclear. In the present study, 8-month-old male APP/PS1/tau triple transgenic AD model (3xTg-AD) mice and wild type (WT) mice (n = 8 for each group) were subjected to chronic sleep deprivation by using the modified multiple platform method, with 20 h of sleep deprivation each day for 21 days. Then, open field test, elevated plus maze test, sugar water preference test, object recognition test, Y maze test and conditioned fear memory test were performed to evaluate anxiety- and depression-like behaviors, and multiple cognitive functions. In addition, the immunohistochemistry technique was used to observe pathological characteristics in the hippocampus of mice. The results showed that: (1) Chronic sleep deprivation did not affect anxiety- (P = 0.539) and depression-like behaviors (P = 0.874) in 3xTg-AD mice; (2) Chronic sleep deprivation exacerbated the impairments of object recognition memory (P < 0.001), working memory (P = 0.002) and the conditioned fear memory (P = 0.039) in 3xTg-AD mice; (3) Chronic sleep deprivation increased amyloid β (Aβ) deposition (P < 0.001) and microglial activation (P < 0.001) in the hippocampus of 3xTg-AD mice, without inducing abnormal tau phosphorylation and neurofibrillary tangles. These results indicate that chronic sleep deprivation exacerbates the impairments of recognition memory, working memory and conditioned fear memory in 3xTg-AD mice by aggravating Aβ deposition and the excessive activation of microglia in the hippocampus.
Subject(s)
Animals , Male , Mice , Alzheimer Disease , Amyloid beta-Peptides , Amyloid beta-Protein Precursor/genetics , Cognition , Disease Models, Animal , Mice, Inbred C57BL , Mice, Transgenic , Presenilin-1 , Sleep Deprivation , tau ProteinsABSTRACT
OBJECTIVE@#To compare the therapeutic effect of electroacupuncture (EA) combined with donepezil hydrochloride and donepezil hydrochloride alone on improving learning-memory ability in patients with Alzheimer's disease (AD), and to explore its action mechanism.@*METHODS@#Sixty patients of AD were randomly divided into an observation group and a control group, 30 cases in each group. The patients in the observation group were treated with EA at governor vessel (GV) combined with donepezil hydrochloride. EA was applied at Baihui (GV 20) and Fengfu (GV 16) with dilatational wave (10 Hz/50 Hz of frequency, 0.5 to 5.0 mA of intensity), and the needles were kept for 40 min, EA was given once a day; the donepezil hydrochloride tablet was taken orally, 5 mg, once a day, and after 4 weeks the dosage might be increased to 10 mg per day according to the specific situation. All the treatment was given for 8 weeks. The patients in the control group were only treated with donepezil hydrochloride with the identical procedure as the observation group. The Montreal cognitive assessment (MoCA) and Alzheimer's disease assessment scale cognitive part (ADAS-Cog) were evaluated before and after treatment; P300 (latency and amplitude of N2 and P3) was detected by EEG/ERP system brain event related potential instrument, and amyloid precursor protein (APP) and β-amyloid protein 1-42 (Aβ) were detected by ELISA.@*RESULTS@#Compared before treatment, the MoCA scores were increased after treatment in the two groups (<0.05), and the MoCA score in the observation group was higher than that in the control group (<0.05). Compared before treatment, the ADAS-Cog scores were decreased after treatment in the two groups (<0.05), and the ADAS-Cog score in the observation group was lower than that in the control group (<0.05). Compared before treatment, the latency of N2 and P3 was shortened and the amplitude was increased after treatment in the two groups (<0.05); after treatment, the latency of N2 and P3 in the observation group was shorter than that in the control group and the amplitude was higher than that in the control group (<0.05). Compared before treatment, the serum levels of APP and Aβ were lower after treatment in the two groups (<0.05), and the serum levels of APP and Aβ in the observation group were lower than those in the control group (<0.05).@*CONCLUSION@#EA at Baihui (GV 20) and Fengfu (GV 6) combined with donepezil hydrochloride can effectively reduce the serum levels of APP and Aβ and improve the scores of MoCA and ADAS-Cog and the levels of N2 and P3 of P300 in AD patients, which has superior effect to donepezil hydrochloride alone in improving the learning-memory of AD patients.
Subject(s)
Humans , Alzheimer Disease , Blood , Therapeutics , Amyloid beta-Peptides , Blood , Amyloid beta-Protein Precursor , Blood , Cognition , Donepezil , Therapeutic Uses , Electroacupuncture , Learning , Memory , Peptide Fragments , BloodABSTRACT
Hepatocellular carcinoma (HCC) is a malignant tumor with high morbidity and mortality globally. It accounts for the majority of primary liver cancer cases. Amyloid precursor protein (APP), a cell membrane protein, plays a vital role in the pathogenesis of Alzheimer's disease, and has been found to be implicated in tumor growth and metastasis. Therefore, to understand the relationship between APP and 5-fluorouracil (5-FU) resistance in liver cancer, Cell Counting Kit-8, apoptosis and cell cycle assays, western blotting, and reverse transcription-quantitative polymerase chain reaction (qPCR) analysis were performed. The results demonstrated that APP expression in Bel7402-5-FU cells was significantly up-regulated, as compared with that in Bel7402 cells. Through successful construction of APP-silenced (siAPP) and overexpressed (OE) Bel7402 cell lines, data revealed that the Bel7402-APP751-OE cell line was insensitive, while the Bel7402-siAPP cell line was sensitive to 5-FU in comparison to the matched control group. Furthermore, APP overexpression decreased, while APP silencing increased 5-FU-induced apoptosis in Bel7402 cells. Mechanistically, APP overexpression and silencing can regulate the mitochondrial apoptotic pathway and the expression of apoptotic suppressor genes (B-cell lymphoma-2 (Bcl-2) and B-cell lymphoma-extra large (Bcl-xl)). Taken together, these results preliminarily revealed that APP overexpression contributes to the resistance of liver cancer cells to 5-FU, providing a new perspective for drug resistance.
Subject(s)
Humans , Amyloid beta-Protein Precursor/physiology , Apoptosis/drug effects , Carcinoma, Hepatocellular/drug therapy , Cell Line, Tumor , Drug Resistance, Neoplasm , Fluorouracil/pharmacology , Liver Neoplasms/drug therapy , Mitochondria/physiology , Proto-Oncogene Proteins c-bcl-2/genetics , bcl-X Protein/geneticsABSTRACT
OBJECTIVE@#To study the effect of β-sheet blocking peptide H102 on the expression of AMPK-mTOR autophagy pathway-related protein in APP/PS1 double transgenic AD mice.@*METHODS@#Thirty male APP/PS1 transgenic male AD mice of 6 months old were randomly divided into AD group and H102 intervention group, and C57BL/6J male mice of the same age were used as control group (n=15). The mice in the HF group were administered with 5 μl (5.8 mg/kg) of H102 polypeptide solution through the nasal cavity at the same time period, and the mice in the control group and the AD group were given the same amount of blank adjuvant solution daily. The memory recognition ability was tested by a new object recognition experiment 30 days after continuous administration. Immunohistochemistry and Western blot were used to detect the expressions of phosphorylated AMP-activated protein kinase(P-AMPK),phosphorylated mammalian target of rapamycin (P-mTOR) and ratio of LC32to LC31(LC3II/I )in brain tissue.@*RESULTS@#Compared with the control group, the new object recognition index (RI) of the AD group was significantly lower (P<0.05), and the P-AMPK and LC3II/I ratios in the brain of the mice were significantly lower (P<0.05). The expression of P-mTOR protein was increased significantly (P<0.05). Compared with the AD group, the RI of the H102 intervention group was increased significantly (P<0.05), and the P-AMPK and LC3II/I ratios in the brain tissue of the mice were increased significantly (P<0.05). The expression of P-mTOR protein was decreased significantly (P<0.05).@*CONCLUSION@#H102 can improve the recognition and memory ability of AD mice by activating the AMPK-mTOR autophagy-related pathway.
Subject(s)
Animals , Male , Mice , AMP-Activated Protein Kinases , Alzheimer Disease , Amyloid beta-Protein Precursor , Autophagy , Disease Models, Animal , Memory , Mice, Inbred C57BL , Mice, Transgenic , Peptides , Pharmacology , Random Allocation , TOR Serine-Threonine KinasesABSTRACT
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.
Subject(s)
Humans , Alzheimer Disease , Amyloid beta-Peptides , Amyloid beta-Protein Precursor , Genetics , Cell Line , Coumarins , Pharmacology , Gene Expression Regulation , Genetics , MicroRNAs , Genetics , MetabolismABSTRACT
Alzheimer's disease (AD) is the most common form of dementia among the elderly, characterized by amyloid plaques, neurofibrillary tangles, and neuroinflammation in the brain, as well as impaired cognitive behaviors. A sex difference in the prevalence of AD has been noted, while sex differences in the cerebral pathology and relevant molecular mechanisms are not well clarified. In the present study, we systematically investigated the sex differences in pathological characteristics and cognitive behavior in 12-month-old male and female APP/PS1/tau triple-transgenic AD mice (3×Tg-AD mice) and examined the molecular mechanisms. We found that female 3×Tg-AD mice displayed more prominent amyloid plaques, neurofibrillary tangles, neuroinflammation, and spatial cognitive deficits than male 3×Tg-AD mice. Furthermore, the expression levels of hippocampal protein kinase A-cAMP response element-binding protein (PKA-CREB) and p38-mitogen-activated protein kinases (MAPK) also showed sex difference in the AD mice, with a significant increase in the levels of p-PKA/p-CREB and a decrease in the p-p38 in female, but not male, 3×Tg-AD mice. We suggest that an estrogen deficiency-induced PKA-CREB-MAPK signaling disorder in 12-month-old female 3×Tg-AD mice might be involved in the serious pathological and cognitive damage in these mice. Therefore, sex differences should be taken into account in investigating AD biomarkers and related target molecules, and estrogen supplementation or PKA-CREB-MAPK stabilization could be beneficial in relieving the pathological damage in AD and improving the cognitive behavior of reproductively-senescent females.
Subject(s)
Animals , Female , Humans , Male , Alzheimer Disease , Metabolism , Pathology , Psychology , Amyloid beta-Protein Precursor , Genetics , Metabolism , Cyclic AMP Response Element-Binding Protein , Metabolism , Cyclic AMP-Dependent Protein Kinases , Metabolism , Disease Models, Animal , Hippocampus , Metabolism , Pathology , Inflammation , Metabolism , Pathology , Psychology , Maze Learning , Physiology , Mice, Inbred C57BL , Mice, Transgenic , Neurofibrillary Tangles , Metabolism , Pathology , Plaque, Amyloid , Metabolism , Pathology , Psychology , Presenilin-1 , Genetics , Metabolism , Sex Characteristics , Spatial Memory , Physiology , p38 Mitogen-Activated Protein Kinases , Metabolism , tau Proteins , Genetics , MetabolismABSTRACT
Astrocytes are closely associated with Alzheimer's disease (AD). However, their precise roles in AD pathogenesis remain controversial. One of the reasons behind the different results reported by different groups might be that astrocytes were targeted at different stages of disease progression. In this study, by crossing hAPP (human amyloid precursor protein)-J20 mice with a line of GFAP-TK mice, we found that astrocytes were activated specifically at an early stage of AD before the occurrence of amyloid plaques, while microglia were not affected by this crossing. Activation of astrocytes at the age of 3-5 months did not affect the proteolytic processing of hAPP and amyloid plaque loads in the brains of hAPP-J20 mice. Our data suggest that early activation of astrocytes does not affect the deposition of amyloid β in an animal model of AD.
Subject(s)
Animals , Humans , Mice , Aldehyde Dehydrogenase , Metabolism , Alzheimer Disease , Genetics , Metabolism , Pathology , Amyloid beta-Peptides , Metabolism , Amyloid beta-Protein Precursor , Genetics , Metabolism , Astrocytes , Metabolism , Brain , Pathology , Calcium-Binding Proteins , Metabolism , Cell Proliferation , Disease Models, Animal , Gene Expression Regulation , Genetics , Glial Fibrillary Acidic Protein , Glutamine , Metabolism , Green Fluorescent Proteins , Genetics , Metabolism , Ki-67 Antigen , Metabolism , Mice, Transgenic , Microfilament Proteins , Metabolism , Mutation , Genetics , Nerve Tissue Proteins , MetabolismABSTRACT
OBJECTIVE@#To observe the gait changes of Alzheimer's disease PS1M146V/APPswe/tauP301L triple-transgenic (3xTg-AD) mice and to investigate the improvement effect of single chain variable domain antibody fragment 17 (scFv17) on the gait.@*METHODS@#In the present study, a selection of 6-month-old 3xTg-AD mice (=18) and C57BL/6 wild-type mice (=24) was performed. First, we observed their gait changes and found that the gait of 12-month-old 3xTg-AD mice was severely damaged. Then, the two groups of mice were randomly divided into four groups:WT+PBS(=12), WT+scFv17(=12), 3xTg-AD+PBS(=9) and 3xTg-AD+scFv17(=9). The gait behavior test and pathological test were performed after 12 weeks'continuous administration of scFv17 (1.5 mg/kg) or an equal volume of PBS (0.01 mol/L) by nasal gavage twice a week.@*RESULTS@#Compared with the same month old wild type mice, the rear track width of 12 month old 3xTg-AD mice was increased(<0.01), swing time percent was decreased (<0.01), stance time percent was increased(<0.01), so the ability of movement coordination and balance was seriously damaged. ScFv17 could improve the coordination and balance ability of 12 month old 3xTg-AD mice(<0.01). The morphological structure of 3xTg-AD mice cerebellar Purkinje cells was improved. The treatment of scFv17 increased the Nissl body number of the cerebellar Purkinje cells of 3xTg-AD mice (<0.01). scFv17 reduced the amyloid β protein (Aβ) plaques in the cerebellar cortex of 3xTg-AD mice (<0.01), and scFv17 reduced the intracellular neurofibrillary tangles (NFT) of the cerebellar Purkinje cells of the 3xTg-AD mice (<0.01).@*CONCLUSIONS@#The coordination and balance ability of 3xTg-AD mice was significantly impaired. ScFv17 can improve gait behaviour in the 3xTg-AD significantly.The mechanism may be related to the improvement of the structure and protein function of cerebellar Purkinje cells, and the eliminating of the Aβ plaques and the neurofibrillary tangles.