Spatial-Temporal Mapping Reveals the Golgi as the Major Processing Site for the Pathogenic Swedish APP Mutation: Familial APP Mutant Shifts the Major APP Processing Site.

Alzheimer's disease is associated with increased levels of amyloid beta (Aß) generated by sequential intracellular cleavage of amyloid precursor protein (APP) by membrane-bound secretases. However, the spatial and temporal APP cleavage events along the trafficking pathways are poorly defined. Here, we use the Retention Using Selective Hooks (RUSH) to compare in real time the anterograde trafficking and temporal cleavage events of wild-type APP (APPwt) with the pathogenic Swedish APP (APPswe) and the disease-protective Icelandic APP (APPice). The analyses revealed differences in the trafficking profiles and processing between APPwt and the APP familial mutations. While APPwt was predominantly processed by the ß-secretase, BACE1, following Golgi transport to the early endosomes, the transit of APPswe through the Golgi was prolonged and associated with enhanced amyloidogenic APP processing and Aß secretion. A 20°C block in cargo exit from the Golgi confirmed ß- and γ-secretase processing of APPswe in the Golgi. Inhibition of the ß-secretase, BACE1, restored APPswe anterograde trafficking profile to that of APPwt. APPice was transported rapidly through the Golgi to the early endosomes with low levels of Aß production. This study has revealed different intracellular locations for the preferential cleavage of APPwt and APPswe and Aß production, and the Golgi as the major processing site for APPswe, findings relevant to understand the molecular basis of Alzheimer's disease.

RACK1 and IRE1 participate in the translational quality control of amyloid precursor protein in Drosophila models of Alzheimer's disease.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by dysregulation of the expression and processing of the amyloid precursor protein (APP). Protein quality control systems are dedicated to remove faulty and deleterious proteins to maintain cellular protein homeostasis (proteostasis). Identidying mechanisms underlying APP protein regulation is crucial for understanding AD pathogenesis. However, the factors and associated molecular mechanisms regulating APP protein quality control remain poorly defined. In this study, we show that mutant APP with its mitochondrial-targeting sequence ablated exhibited predominant endoplasmic reticulum (ER) distribution and led to aberrant ER morphology, deficits in locomotor activity, and shortened lifespan. We searched for regulators that could counteract the toxicity caused by the ectopic expression of this mutant APP. Genetic removal of the ribosome-associated quality control (RQC) factor RACK1 resulted in reduced levels of ectopically expressed mutant APP. By contrast, gain of RACK1 function increased mutant APP level. Additionally, overexpression of the ER stress regulator (IRE1) resulted in reduced levels of ectopically expressed mutant APP. Mechanistically, the RQC related ATPase VCP/p97 and the E3 ubiquitin ligase Hrd1 were required for the reduction of mutant APP level by IRE1. These factors also regulated the expression and toxicity of ectopically expressed wild type APP, supporting their relevance to APP biology. Our results reveal functions of RACK1 and IRE1 in regulating the quality control of APP homeostasis and mitigating its pathogenic effects, with implications for the understanding and treatment of AD.

Eta-secretase-like processing of the amyloid precursor protein (APP) by the rhomboid protease RHBDL4.

The amyloid precursor protein (APP) is a key protein in Alzheimer's disease synthesized in the endoplasmic reticulum (ER) and translocated to the plasma membrane where it undergoes proteolytic cleavages by several proteases. Conversely, to other known proteases, we previously elucidated rhomboid protease RHBDL4 as a novel APP processing enzyme where several cleavages likely occur already in the ER. Interestingly, the pattern of RHBDL4-derived large APP C-terminal fragments resembles those generated by the η-secretase or MT5-MMP, which was described to generate so-called Aη fragments. The similarity in large APP C-terminal fragments between both proteases raised the question of whether RHBDL4 may contribute to η-secretase activity and Aη-like fragments. Here, we identified two cleavage sites of RHBDL4 in APP by mass spectrometry, which, intriguingly, lie in close proximity to the MT5-MMP cleavage sites. Indeed, we observed that RHBDL4 generates Aη-like fragments in vitro without contributions of α-, ß-, or γ-secretases. Such Aη-like fragments are likely generated in the ER since RHBDL4-derived APP-C-terminal fragments do not reach the cell surface. Inherited, familial APP mutations appear to not affect this processing pathway. In RHBDL4 knockout mice, we observed increased cerebral full-length APP in comparison to wild type (WT) in support of RHBDL4 being a physiologically relevant protease for APP. Furthermore, we found secreted Aη fragments in dissociated mixed cortical cultures from WT mice, however significantly fewer Aη fragments in RHBDL4 knockout cultures. Our data underscores that RHBDL4 contributes to the η-secretease-like processing of APP and that RHBDL4 is a physiologically relevant protease for APP.

Single-cell transcriptomic analysis in clear cell renal cell carcinoma: Deciphering the role of APP within the tumour microenvironment.

Clear cell renal cell carcinoma (ccRCC) represents a significant challenge in oncology, primarily due to its resistance to conventional therapies. Understanding the tumour microenvironment (TME) is crucial for developing new treatment strategies. This study focuses on the role of amyloid precursor protein (APP) in tumour-associated macrophages (TAMs) within the ccRCC TME, exploring its potential as a prognostic biomarker. Basing TAM-related genes, the prognostic model was important to constructed. Employing advanced single-cell transcriptomic analysis, this research dissects the TME of ccRCC at an unprecedented cellular resolution. By isolating and examining the gene expression profiles of individual cells, particularly focusing on TAMs, the study investigates the expression levels of APP and their association with the clinical outcomes of ccRCC patients. The analysis reveals a significant correlation between the expression of APP in TAMs and patient prognosis in ccRCC. Patients with higher APP expression in TAMs showed differing clinical outcomes compared to those with lower expression. This finding suggests that APP could serve as a novel prognostic biomarker for ccRCC, providing insights into the disease progression and potential therapeutic targets. This study underscores the importance of single-cell transcriptomics in understanding the complex dynamics of the TME in ccRCC. The correlation between APP expression in TAMs and patient prognosis highlights APP as a potential prognostic biomarker. However, further research is needed to validate these findings and explore the regulatory mechanisms and therapeutic implications of APP in ccRCC.

Early Animal Origin of BACE1 APP/Aß Proteolytic Function.

Alzheimer's disease is characterized, in part, by the accumulation of ß-amyloid (Aß) in the brain. Aß is produced via the proteolysis of APP by BACE1 and γ-secretase. Since BACE1 is the rate-limiting enzyme in the production of Aß, and a target for therapeutics, it is of interest to know when its proteolytic function evolved and for what purpose. Here, we take a functional evolutionary approach to show that BACE1 likely evolved from a gene duplication event near the base of the animal clade and that BACE1 APP/Aß proteolytic function evolved during early animal diversification, hundreds of millions of years before the evolution of the APP/Aß substrate. Our examination of BACE1 APP/Aß proteolytic function includes cnidarians, ctenophores, and choanoflagellates. The most basal BACE1 ortholog is found in cnidarians, while ctenophores, placozoa, and choanoflagellates have genes equally orthologous to BACE1 and BACE2. BACE1 from a cnidarian (Hydra) can cleave APP to release Aß, pushing back the date of the origin of its function to near the origin of animals. We tested more divergent BACE1/2 genes from a ctenophore (Mnemiopsis) and a choanoflagellate (Monosiga), and neither has this activity. These findings indicate that the specific proteolytic function of BACE1 evolved during the very earliest diversification of animals, most likely after a gene-duplication event.

ADAM10 isoforms: optimizing usage of antibodies based on protein regulation, structural features, biological activity and clinical relevance to Alzheimer's disease.

A Disintegrin and Metalloproteinase 10 (ADAM10) is a crucial transmembrane protein involved in diverse cellular processes, including cell adhesion, migration, and proteolysis. ADAM10's ability to cleave over 100 substrates underscores its significance in physiological and pathological contexts, particularly in Alzheimer's disease (AD). This review comprehensively examines ADAM10's multifaceted roles, highlighting its critical function in the non-amyloidogenic processing of the amyloid precursor protein (APP), which mitigates amyloid beta (Aß) production, a critical factor in AD development. We summarize the regulation of ADAM10 at multiple levels: transcriptional, translational, and post-translational, revealing the complexity and responsiveness of its expression to various cellular signals. A standardized nomenclature for ADAM10 isoforms is proposed to improve clarity and consistency in research, facilitating better comparison and replication of findings across studies. We address the challenges in detecting ADAM10 isoforms using antibodies, advocating for standardized detection protocols to resolve discrepancies in results from different biological matrices. This review underscores the potential of ADAM10 as a biomarker for early diagnosis and a therapeutic target in AD. By consolidating current knowledge on ADAM10's regulation and function, we aim to provide insights that will guide future research and therapeutic strategies in the AD context.

Altered amyloid-ß structure markedly reduces gliosis in the brain of mice harboring the Uppsala APP deletion.

Deposition of amyloid beta (Aß) into plaques is a major hallmark of Alzheimer's disease (AD). Different amyloid precursor protein (APP) mutations cause early-onset AD by altering the production or aggregation properties of Aß. We recently identified the Uppsala APP mutation (APPUpp), which causes Aß pathology by a triple mechanism: increased ß-secretase and altered α-secretase APP cleavage, leading to increased formation of a unique Aß conformer that rapidly aggregates and deposits in the brain. The aim of this study was to further explore the effects of APPUpp in a transgenic mouse model (tg-UppSwe), expressing human APP with the APPUpp mutation together with the APPSwe mutation. Aß pathology was studied in tg-UppSwe brains at different ages, using ELISA and immunohistochemistry. In vivo PET imaging with three different PET radioligands was conducted in aged tg-UppSwe mice and two other mouse models; tg-ArcSwe and tg-Swe. Finally, glial responses to Aß pathology were studied in cell culture models and mouse brain tissue, using ELISA and immunohistochemistry. Tg-UppSwe mice displayed increased ß-secretase cleavage and suppressed α-secretase cleavage, resulting in AßUpp42 dominated diffuse plaque pathology appearing from the age of 5-6 months. The γ-secretase cleavage was not affected. Contrary to tg-ArcSwe and tg-Swe mice, tg-UppSwe mice were [11C]PiB-PET negative. Antibody-based PET with the 3D6 ligand visualized Aß pathology in all models, whereas the Aß protofibril selective mAb158 ligand did not give any signals in tg-UppSwe mice. Moreover, unlike the other two models, tg-UppSwe mice displayed a very faint glial response to the Aß pathology. The tg-UppSwe mouse model thus recapitulates several pathological features of the Uppsala APP mutation carriers. The presumed unique structural features of AßUpp42 aggregates were found to affect their interaction with anti-Aß antibodies and profoundly modify the Aß-mediated glial response, which may be important aspects to consider for further development of AD therapies.

GDI2 deletion alleviates neurodegeneration and memory loss in the 5xFAD mice model of Alzheimer's disease.

Accumulation of insoluble deposits of amyloid ß-peptide (Aß), derived from amyloid precursor protein (APP) processing, represents one of the major pathological hallmarks of Alzheimer's disease (AD). Perturbations in APP transport and hydrolysis could lead to increased Aß production. However, the precise mechanisms underlying APP transport remain elusive. The GDP dissociation inhibitor2 (GDI2), a crucial regulator of Rab GTPase activity and intracellular vesicle and membrane trafficking, was investigated for its impact on AD pathogenesis through neuron-specific knockout of GDI2 in 5xFAD mice. Notably, deficiency of GDI2 significantly ameliorated cognitive impairment, prevented neuronal loss in the subiculum and cortical layer V, reduced senile plaques as well as astrocyte activation in 5xFAD mice. Conversely, increased activated microglia and phagocytosis were observed in GDI2 ko mice. Further investigation revealed that GDI2 knockout led to more APP co-localized with the ER rather than the Golgi apparatus and endosomes in SH-SY5Y cells, resulting in decreased Aß production. Collectively, these findings suggest that GDI2 may regulate Aß production by modulating APP intracellular transport and localization dynamics. In summary, our study identifies GDI2 as a pivotal regulator governing APP transport and process implicated in AD pathology; thus highlighting its potential as an attractive pharmacological target for future drug development against AD.

Recent Insights into the Neurobiology of Alzheimer's Disease and Advanced Treatment Strategies.

In recent years, significant advancements have been made in understanding Alzheimer's disease from both neurobiological and clinical perspectives. Exploring the complex systems underlying AD has unveiled insights that could potentially revolutionize therapeutic approaches. Recent investigations have highlighted intricate interactions among genetic, molecular, and environmental factors in AD. Optimism arises from neurobiological advancements and diverse treatment options, potentially slowing or halting disease progression. Amyloid-beta plaques and tau protein tangles crucially influence AD onset and progression. Emerging treatments involve diverse strategies, such as approaches targeting multiple pathways involved in AD pathogenesis, such as inflammation, oxidative stress, and synaptic dysfunction pathways. Clinical trials using humanized monoclonal antibodies, focusing on immunotherapies eliminating amyloid-beta, have shown promise. Nonpharmacological interventions such as light therapy, electrical stimulation, cognitive training, physical activity, and dietary changes have drawn attention for their potential to slow cognitive aging and enhance brain health. Precision medicine, which involves tailoring therapies to individual genetic and molecular profiles, has gained traction. Ongoing research and interdisciplinary collaboration are expected to yield more effective treatments.

Neuroprotective Strategies and Cell-Based Biomarkers for Manganese-Induced Toxicity in Human Neuroblastoma (SH-SY5Y) Cells.

Manganese (Mn) is an essential heavy metal in the human body, while excess Mn leads to neurotoxicity, as observed in this study, where 100 µM of Mn was administered to the human neuroblastoma (SH-SY5Y) cell model of dopaminergic neurons in neurodegenerative diseases. We quantitated pathway and gene changes in homeostatic cell-based adaptations to Mn exposure. Utilizing the Gene Expression Omnibus, we accessed the GSE70845 dataset as a microarray of SH-SY5Y cells published by Gandhi et al. (2018) and applied statistical significance cutoffs at p < 0.05. We report 74 pathway and 10 gene changes with statistical significance. ReactomeGSA analyses demonstrated upregulation of histones (5 out of 10 induced genes) and histone deacetylases as a neuroprotective response to remodel/mitigate Mn-induced DNA/chromatin damage. Neurodegenerative-associated pathway changes occurred. NF-κB signaled protective responses via Sirtuin-1 to reduce neuroinflammation. Critically, Mn activated three pathways implicating deficits in purine metabolism. Therefore, we validated that urate, a purine and antioxidant, mitigated Mn-losses of viability in SH-SY5Y cells. We discuss Mn as a hypoxia mimetic and trans-activator of HIF-1α, the central trans-activator of vascular hypoxic mitochondrial dysfunction. Mn induced a 3-fold increase in mRNA levels for antioxidant metallothionein-III, which was induced 100-fold by hypoxia mimetics deferoxamine and zinc.

Investigation of the Iron Oxide Nanoparticle Effects on Amyloid Precursor Protein Processing in Hippocampal Cells.

Introduction: Iron oxide nanoparticles (Fe2O3-NPs) are small magnetic particles that are widely used in different aspects of biology and medicine in modern life. Fe2O3-NP accumulated in the living cells due to the absence of an active system to excrete the iron ions and damages cellular organelles by high reactivity. Methods: Herein cytotoxic effects of Fe2O3-NP with a size of 50 nm on the primary culture of neonatal rat hippocampus were investigated using 2,5-diphenyltetrazolium bromide (MTT) assay. Pathophysiological signs of Alzheimer's disease such as amyloid precursor protein (APP) expression, Aß aggregation, soluble APPα, and APPß secretion were also investigated in hippocampal cells treated with various concentrations of nanoparticle (NP) for different exposure times. Results: Our results revealed that Fe2O3-NP treatment causes oxidative stress in cells which is accompanied by upregulation of the APP and Aß in a concentration-dependent manner. NP exposure also leads to more secretion of sAPPß rather than sAPPα, leading to increased activation of ß-secretase in NP-received cells. All the harmful effects accumulate in neurons that cannot be renovated, leading to neurodegeneration in Alzheimer's disease. Conclusion: This study approved iron-based NPs could help to develop Alzheimer's and related neurological disorders and explained why some of the iron chelators have therapeutic potential in Alzheimer's disease. Highlights: Fe2O3-NP induced oxidative stress in hippocampal cells in a concentration dependent manner.Fe2O3-NP imposed up-regulation of APP in hippocampal cells.Fe2O3-NP activated ß-secretase and elevated sAPPß/sAPPα ratio.Cumulative effects of Fe2O3-NP damages increased cell death in neurons. Plain Language Summary: The most common type of dementia is Alzheimer's disease (AD), which is characterized by chronic neurodegeneration, impairment of memory, and disturbed planning, language, and thinking ability. In recent years, the use of nanoparticles has been increased in all aspects of life. Among these nanoparticles, iron oxide nanoparticles (Fe2O3-NP) are vital in biological sciences, medicine, magnetic resonance imaging, ultrasound, and optical imaging. Considering the general application and high reactivity of iron, growing concerns exist about the Fe2O3-NP application harms, especially in the central nervous system. Hippocampus tissue is one of the affected tissues in AD, which is widely investigated in recent years. This study aimed to investigate the cytotoxic effects of Fe2O3-NP on the primary culture of the hippocampus as one of the main tissues damaged in patients with AD. Our results revealed that treatment with different concentrations of Fe2O3-NP caused cellular damage in hippocampal cells. Exposure to Fe2O3-NP also caused oxidative stress. Our results showed a close association between oxidative stress and AD's pathological symptoms. The Fe2O3-NP application in medicine and biology should be limited.

Huanglian Jiedutang Regulates HIF-1α/VEGF Signaling Pathway to Improve Learning and Memory Abilities of APP/PS1 Transgenic Mice / 中国实验方剂学杂志

ObjectiveTo reveal the effects of Huanglian Jiedutang （HLJDT） on the learning and memory abilities of APP/PS1 transgenic mice via hypoxia-inducible factor-1α （HIF-1α）/vascular endothelial growth factor （VEGF） signaling pathway. MethodForty 5-month-old β-amyloid precursor protein （APP）/presenilin 1（PS1） mice were randomized into the model， donepezil （0.001 g·kg-1·d-1）， and low-， medium-， and high-dose （1.5， 3， 6 g·kg-1·d-1， respectively） HLJDT groups， and 8 C57BL/6 mice were taken as the normal group. After 45 days of continuous administration， Morris water maze test was conducted， and the organ indexes were calculated. The morphological structure of cerebral vascular endothelial cells in mice was observed under a transmission electron microscope. Western blot was employed to measure the protein levels of APP， HIF-1α， VEGF，VEGFA， and brain-derived neurotrophic factor （BDNF） in the hippocampus. The mRNA levels of APP， HIF-1α， and VEGF were determined by real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）. ResultCompared with the normal group， the model group showed prolonged escape latency （P<0.05）， reduced distance and time around the target platform （P<0.05）， decrease brain and spleen indexes （P<0.05）， vascular endothelial cells with karyopyknosis and not abundant cytoplasm， up-regulated protein levels of APP， HIF-1α， VEGF， and VEGFA （P<0.05）， down-regulated protein level of BDNF （P<0.05）， and up-regulated mRNA levels of APP， HIF-1α， and VEGF （P<0.05） in the hippocampus. Compared with the model group， high-dose HLJDT shortened the escape latency （P<0.05）， increased the distance and time around the target platform （P<0.05）， raised the brain and spleen indexes （P<0.05）， repaired the organelles of vascular endothelial cells， down-regulated the protein levels of APP， HIF-1α， VEGF， and VEGFA （P<0.05）， up-regulated the protein level of BDNF （P<0.05）， and down-regulated the mRNA levels of APP， HIF-1α， and VEGF （P<0.05） in the hippocampus. ConclusionHLJDT can improve the learning and memory abilities of mice by reducing the expression of HIF-1α and VEGF， thus protecting the nerves.

Expression, purification and micelle reconstruction of the transmembrane domain of the human amyloid precursor protein for NMR studies / 生物工程学报

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.

Abdominal acupoint thread embedding therapy based on "brain-intestinal connection" for mild-to-moderate Alzheimer's disease and its effects on serum levels of APP and Aβ / 中国针灸

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

Effect of Huangjingwan on Antioxidant Functions and Aβ1-42 and APP Protein Expressions in Brain of Alzheimer' s Disease Rats / 中国实验方剂学杂志

Objective::To observe the effect of Huangjingwan (HW) on antioxidant functions and β-amyloid 1-42 (Aβ1-42) and amyloid precursor protein (APP) expressions in the brain of Alzheimer' s disease (AD) rats. Method::SD rats were randomly divided into normal control group, sham model control group, AD model group, and low, medium, high-dose (equivalent raw drug dose 1, 3, 9 g·kg-1·d-1) HW groups.The AD models were established through intraperitoneal injection with 1.25% D-galactose (120 mg·kg-1·d-1, 6 consecutive weeks) and then one-time right ventricular injection with Aβ1-42 (10 μg). Two weeks after modeling, the rats in each HW group received corresponding drugs through intragastric administration, once a day, while the rats in sham model control group, AD model group were given normal saline 1 mL through intragastric administration, once a day.Gastric perfusion lasted for 8 weeks.At the end of the experiment, learning and memory abilities of the rats were assessed by Platform Jumping Test.The changes of physical endurance in rats were tested by 10% weight swimming under load.The activities of superoxide dismutase (SOD), glutathione reductase (GR), glutathione peroxidase (GSH-Px) antioxidant enzymes and the contents of glutathione (GSH) and malondialdehyde (MDA) in rat brain tissue were detected by colorimetry.The changes of interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), Aβ1-42 and APP protein in rat brain tissues were determined by enzyme linked immunosorbent assay (ELISA). Western blot analysis was used to measure the expression of APP protein in rat brain. Result::Compared with the normal control group, rats in AD model group showed an obvious dementia state, that is more lying and less movement, longer learning response time, significant increase in the number of learning and memory errors, significant attenuation in physical fitness, significant decrease in the activities of antioxidant enzymes (SOD, GR, GSH-Px) and anti-inflammatory factors GSH in brain, significant rise in the levels of inflammatory factors MDA, IL-1β and TNF-α and the content of Aβ1-42 protein, and significant reduction in the content of APP protein in brain (P<0.01). Low, medium and high-dose HW could ameliorate dementia symptoms in AD rats, improve the achievement of learning and memory, antagonize body weakness and increase physical fitness, promote SOD, GR, GSH-Px activities and anti-inflammatory factor GSH level in the brain, reduce the levels of MDA, IL-1β and TNF-α in the brain, decrease the level of Aβ1-42 and increase the level of APP protein in the brains of AD rats compared with the AD model group (P<0.05, P<0.01), besides, within the dose range of 1-9 g·kg-1·d-1, HW has a more obvious effect with the increase of dose. Conclusion::HW has the effects in preventing and treating AD, which is related to the HW' s mechanisms in enhancing the function of antioxidant system in brain, reducing neuroinflammatory reaction and deposition of Aβ1-42 induced by oxidative stress, and maintaining the expression level of APP protein.

Neuroprotective role and mechanism of S14G-humanin in amyloid precursor protein/presenolin-1 double transgenic mice / 中华神经医学杂志

Objective:To investigate the effect of S14G-humanin (HNG) on spatial learning and memory abilities and hippocampal neuron autophagy in amyloid precursor protein (APP)/presenolin-1 (PS1) double-transgenic mice.Methods:The APP/PSl transgenic mice were randomly divided into model group and HNG treatment group ( n=8). Another 8 C57BL/6J mice were chosen as controls; 0.5 mL double-steamed water was intraperitoneally injected into mice in the control group and model group every day, and 0.5 mL of HNG (50 μg/kg) was intraperitoneally injected into mice in the HNG treatment group every day; after 4 weeks of continuous injection, Morris water maze test was used to test the spatial learning and memory abilities of mice in each group. Western blotting and reverse transcription-PCR were used to detect the protein and mRNA expressions of ubiquitin like kinase1 (ULK1), P62, anti-microtubule associated protein 1 light chain 3 (LC3) II/LC3 I and cathepsin D in the hippocampus. Immunohistochemical staining was used to detect the deposition of amyloid β protein (Aβ) in the hippocampus of mice in each group. Results:Morris water maze test showed that the escape latency of mice in the control group, HNG treatment group and model group was statistically prolonged in turn, and the number of times traversing through the quadrant of the original platform was significantly smaller in turn ( P<0.05). Western blotting and reverse transcription-PCR showed that the ULK1 protein and mRNA expressions in the hippocampus decreased successively, P62 protein and mRNA expressions increased successively, LC3 II protein/LC3 I protein and LC3 II mRNA/ LC3 I mRNA ratio increased successively, and cathepsin D protein and mRNA expressions decreased successively in the control group, HNG treatment group and model group, with significant differences ( P<0.05). Immunohistochemical staining showed that the Aβ positive expressions in the hippocampal neurons of the control group, HNG treatment group and model group increased successively, with statistical differences ( P<0.05). Conclusion:HNG treatment can improve the spatial learning and memory abilities, which may be attributed to ameliorate autophagic network dysfunction and reduce Aβ plaques in the hippocampi of APP/PSl transgenic mice.

Effect of Electro-acupuncture on Behavior Alterations in Mice with Isoflurane-induced Alzheimer's Disease and Its Mechanism / 广州中医药大学学报

Objective To investigate the effect of electro-acupuncture on behavior alterations,the expression of amyloid precursor protein(APP),amyloid β-protein(Aβ) proteins and neuroapoptosis in the cerebral cortex in the APPswe/PS1dE9 double transgenic mice with Alzheimer's disease (AD) induced by isoflurane.Methods Sixmonth-old AD mice and wild-type mice at the same age were randomly divided into wile-type control group,AD group,isoflurane group,electro-acupuncture group (N =8).The mice were given pretreatment with electro-acupuncture at Baihui(GV20) acupoint and Yongquan(KI 1) acupoint once a day for 3 successive days,15 min each time.And then the mice in electro-acupuncture group and isoflurane group were exposed to a box full of 1.2％ isoflurane for 4 hours.Morris water maze was used to test the learning and memory abilities of the mice,Western blotting method was used to detect the expression of APP-C83,APP-C99 and Aβ,and terminal deoxynucleotidyl transferase(TdT)-mediated dUTP nick end labeling(TUNEL) was used to detect neuroapoptosis in the cerebral cortex.Results The escape latency of AD group was longer than that of wild-type mice group(P＜0.05),and the latency of isoflurane group was longer than that of AD group (P ＜ 0.05),while the latency of electro-acupuncture group was shorter than that of isoflurane group(P ＜ 0.05).The percentage of retention time in the target quadrant and the times for crossing the target quadrant in isoflurane group were lower than those of AD group (P ＜ 0.05),but were higher in electro-acupuncture group than those in isoflurane group (P ＜ 0.05).APPC83 expression level in isoflurane group was significantly lower than that in AD group (P ＜ 0.05),while APP-C83 expression level in electro-acupuncture group was higher than that in isoflurane group (P ＜ 0.05).APP-C99 expression level in isoflurane group was significantly higher than that in AD group (P ＜ 0.05),and APP-C99 expression level in electro-acupuncture group was lower than that in isoflurane group (P ＜ 0.05).The cortical apoptosis index in isoflurane group was significantly higher than that in AD group (P ＜ 0.05),and the cortical apoptosis index in electro-acupuncture group was lower than that in isoflurane group (P ＜ 0.05).The expression level of Aβ in AD group,isoflurane group and electro-acupuncture group was significantly higher than wild-type control group (P ＜ 0.05).Conclusion Electro-acupuncture can relieve the AD-like neurotoxicity induced by isoflurane and inhibit the decline of learning and memory abilities of AD mice,and the mechanism is probably related with suppressing the overexpression of APP-C99 and reducing the production and accumulation of Aβ,thereby alleviating the neuroapoptosis.

Over-expressed human amyloid precursor protein depressed M cholinergic receptor binding in SH-SY5Y cells / 基础医学与临床

Objective To observe the effect of over-expressed amyloid procursor protein(APP) gene on the cholinergic receptor binding and the ChAT activity in human neuroblastoma cell line SH-SY5Y.Methods SH-SY5Y cells were transfected with pCMV695 plasmid containing wild type human APP695 gene by Lipofectamine 2000 method.The expression of APP was detected by Western blot.A? contents were test by ELISA assay in over-expression SH-SY5Y cell clones(SH-SY5Y-APP).The special binding of muscarinic and nicotinic cholinergic receptors in those A?-overproducing cell clones was determined by radio-ligand binding method.The cholinergic acetyl transferase(ChAT) activity was assayed by radiao-immunoassay.Results No evident morphologic changes of cytotoxicity were detected after transfection.When A? production was 2～2.6 times as much as that of normal cells,muscarinic receptor binding was decreased from 18.5 % to 21.9 %(P

Therapeutic Potential of ?-Secretase in Alzheimer's Disease / 生物化学与生物物理进展

Alzheimer!s disease (AD) is one of the commonest neurodegenerative diseases affected mainly theelderly. AD is characterized by the formation of neuritic plaque in brain, which is composed mainly of extracellular?amyloid deposion, the A?. A?is deprived from serial hydrolysis of amyloid precursor protein (APP) by twosecretases, the ?and ?-secretase respectively. Alternatively, APP can also be sequential processed by ?-secretaseand ?-secretase, which not only preclude the formation of A?, but also generate a large ectodomain (sAPP?) whohas several neuroprotective properties. Thus the secondary processing pathway has become the focus of ADresearch. Many results have indicated that members of the adamalysin family of proteins, mainly the ADAM 10,ADAM 17 and ADAM 9, fulfill some of the criteria required of ?-secretase. Here the biological characteristics of?-secretase, its activity regulation and its potential function as targets for the treatment of AD were summerized.