Cellular response toβ-amyloid neurotoxicity in Alzheimer's disease and implications in new therapeutics

β-Amyloid(Aβ)is a specific pathological hallmark of Alzheimer's disease(AD).Because of its neurotoxicity,AD patients exhibit multiple brain dysfunctions.Disease-modifying therapy(DMT)is the central concept in the development of AD thera-peutics today,and most DMT drugs that are currently in clinical trials are anti-Aβdrugs,such as aducanumab and lecanemab.Therefore,understanding Aβ's neurotoxic mechanism is crucial for Aβ-targeted drug development.Despite its total length of only a few dozen amino acids,Aβis incredibly diverse.In addition to the well-known Aβ_(1-42),N-terminally truncated,glutaminyl cyclase(QC)catalyzed,and pyroglutamate-modified Aβ(pEAβ)is also highly amyloidogenic and far more cytotoxic.The extracel-lular monomeric Aβ_(x-42)(x=1-11)initiates the aggregation to form fibrils and plaques and causes many abnormal cellular responses through cell membrane receptors and receptor-coupled signal pathways.These signal cascades further influence many cel-lular metabolism-related processes,such as gene expression,cell cycle,and cell fate,and ultimately cause severe neural cell damage.However,endogenous cellular anti-Aβdefense processes always accompany the Aβ-induced microenvironment alterations.Aβ-cleaving endopeptidases,Aβ-degrading ubiquitin-proteasome system(UPS),and Aβ-engulfing glial cell immune responses are all essential self-defense mechanisms that we can leverage to develop new drugs.This review discusses some of the most recent advances in understanding Aβ-centric AD mechanisms and suggests prospects for promising anti-Aβstrategies.

β-Amyloid对PC12细胞毒性损害的机制研究

目的利用体外PC1 2细胞培养来研究Aβ对神经元细胞毒性作用 ,以探索早老性痴呆 (AD)的发生机制。 方法通过DNA末端标记及电子显微镜超微结构观察研究了Aβ诱导培养的PC1 2细胞的形态学和分子生化改变。 结果Aβ可诱导PC1 2细胞核DNA发生降解 ,出现染色质浓缩成块状 ,胞浆浓缩 ,胞膜内陷 ,凋亡小体形成等。 结论Aβ在AD发生中可能是通过诱导神经元凋亡而引起神经元丢失的。

淫羊藿黄酮对APP转基因小鼠学习记忆及β-amyloid生成的影响

观察不同月龄APP(amyloid precursor protein)转基因模型小鼠学习记忆功能的改变,以及中药有效部位淫羊藿黄酮对10月龄转基因小鼠学习记忆功能和脑内APP、BACE的表达及β-淀粉样肽(β-amyloid,Aβ)生成及含量的影响.用药组小鼠自4月龄开始灌胃给予淫羊藿黄酮小(0.03hg·kg-1/d)、大剂量(0.1g·kg-1/d)6个月至10月龄,正常对照组、转基因阴性对照组及模型组以同样方式灌胃给予蒸馏水.应用Morris水迷宫和物体识别方法测试小鼠学习记忆能力,应用免疫组化学及WesternBlot方法分别检测海马CA1区及皮层中APP、BACE的表达,采用双抗体夹心ELISA试剂盒测定海马中不溶性Aβ1-42含量.研究结果表明,APP转基因小鼠在4月龄即出现学习记忆能力障碍,在水迷宫实验中,比转基因阴性对照组小鼠潜伏期延长28%(p<0.05).增龄至10月龄,APP转基因小鼠学习记忆能力明显下降,水迷宫潜伏期及游泳距离与转基因阴性对照组的差异分别加大为40%(p<0.01)和35%(p<0.05),物体识别实验中分辨指数的差异为61%(p<0.05).与正常对照组及转基因阴性对照组相比,10月龄转基因模型小鼠海马CA1区及皮层中APP和BACE的表达明显增加,海马中Aβ1-42的含量明显升高.淫羊藿黄酮大剂量可明显改善10月龄APP转基因小鼠Morris水迷宫作业成绩,提高模型鼠物体识别能力,明显减少APP转基因模型小鼠海马和皮层APP及BACE的表达,降低海马Aβ1-42的含量.提示淫羊藿黄酮能改善APP转基因模型小鼠学习记忆能力和减少Aβ经由淀粉源途径生成及含量,对防治AD等神经退行性疾病具有良好的应用前景.

未知病变类型脑血管中β-amyloid、α-actin、collagen Ⅳ的含量

目的研究未知病变类型脑血管病变的结构特征。方法通过刚果红染色、免疫组织化学染色、计算机图像分析技术对未知病变类型脑血管病变的β-amyloid、α-actin、collagenⅣ的含量进行研究。结果未知病变类型脑血管壁α-actin、collagenⅣ呈少量阳性染色,与正常脑血管存在显著差异(P<0.05);β-amyloid染色呈阴性,与正常脑血管无差异(P>0.05)。病变血管壁中上述三种蛋白的表达特点与脑血管淀粉样变(cerebralamyloidangiopathy,CAA)及小动脉硬化玻璃样变不同。结论未知病变类型脑血管病变具有不同于CAA的病变特征。

Relationship between β-amyloid protein 1-42, thyroid hormone levels and the risk of cognitive impairment after ischemic stroke

BACKGROUND Post-stroke cognitive impairment(PSCI)is not only a common consequence of stroke but also an important factor for adverse prognosis of patients.Biochemical indicators such as blood lipids and blood pressure are affected by many factors,and the ability of evaluating the progress of patients with PSCI is insufficient.Therefore,it is necessary to find sensitive markers for predicting the progress of patients and avoiding PSCI.Recent studies have shown thatβ-amyloid protein 1-42(Aβ1-42)and thyroid hormone levels are closely related to PSCI,which may be the influencing factors of PSCI,but there are few related studies.AIM To investigate the relationship between serum levels of Aβand thyroid hormones in acute stage and PSCI and its predicted value.METHODS A total of 195 patients with acute cerebral infarction confirmed from June 2016 to January 2018 were enrolled in this study.Baseline data and serological indicators were recorded to assess cognitive function of patients.All patients were followed up for 1 year.Their cognitive functions were evaluated within 1 wk,3 mo,6 mo and 1 yr after stroke.At the end of follow-up,the patients were divided into PSCI and non-PSCI according to Montreal cognitive assessment score,and the relationship between biochemical indexes and the progression of PSCI was explored.RESULTS Compared with patients with non-PSCI,the levels of Aβ1-42,triiodothyronine(T3)and free thyroxin were lower in the patients with PSCI.Repeated measures analysis of variance showed that the overall content of Aβ1-42 and T3 in PSCI was also lower than that of the non-PSCI patients.Further analysis revealed that Aβ1-42(r=0.348),T3(r=0.273)and free thyroxin(r=0.214)were positively correlated with disease progression(P<0.05),suggesting that these indicators have the potential to predict disease progression and outcome.Cox regression analysis showed that Aβ1-42 and T3 were important factors of PSCI.Then stratified analysis showed that the lower the Aβ1-42 and T3,the higher risk of PSCI in patients who were aged over 70,female and illiterate.CONCLUSION Aβ1-42 and T3 have the ability to predict the progression of PSCI,which is expected to be applied clinically to reduce the incidence of PSCI and improve the quality of life of patients.

血清β-Amyloid水平在膝骨关节炎发生评估中的应用价值

目的:探讨膝骨关节炎(KOA)患者血清中β-Amyloid水平与KOA发生及预后的相关性。方法:从温州医科大学附属第二医院育英儿童医院住院患者中选取56例经全膝关节置换或膝关节镜手术治疗的KOA患者为试验组和25例非KOA患者为对照组。试验组根据X线Kellgren-Lawrence(K-L)分级方法分级,同时收集手术患者术前术后血清标本,采用双抗体夹心酶联免疫吸附法(ELISA)测定血清β-Amyloid水平后,绘制受试者工作特征(ROC)曲线评价血清β-Amyloid对KOA的预测价值,采用Spearman秩相关分析患者血清β-Amyloid水平与KOA的相关性,应用Pearson相关性分析患者手术前后血清β-Amyloid变化值与患者住院时间的相关性。结果:试验组患者血清β-Amyloid水平高于对照组(P<0.001);Logistic回归分析显示β-Amyloid是KOA患病的危险因素(OR=15.122,P<0.05);ROC曲线分析显示,β-Amyloid cut off值等于0.770,曲线下面积(AUC)为0.752,95%CI=0.636~0.867,敏感度85.5%,特异度60.0%;K-L2组患者血清β-Amyloid低于K-L3/4组患者(P=0.041);β-Amyloid的表达与KOA严重程度分级呈显著正相关(r=0.332,P=0.013);术前KOA患者血清β-Amyloid水平高于术后(P=0.002),KOA患者手术前后血清β-Amyloid变化值与患者住院时间呈显著负相关(r=-0.949,P<0.001)。结论:β-Amyloid是KOA患病的危险因素,血清β-Amyloid水平可能成为一种潜在的评估KOA发生及预后的生物标志物。

THE PROTECTIVE EFFECTS OF THE TOTAL SAPONIN OF DIPSACUS ASPEROIDES ON THE APOPTOSIS OF HIPPOCAMPAL NEURONS INDUCED BY β-AMYLOID PROTEIN

Objective To investigate the effects of the total saponin of Dipsacus asperoides (tSDA) and ginsenoside Rb1 (GRb1) on the apoptosis of primary cultured hippocampal neurons induced by β-amyloid protein (Aβ). Methods Primary cultured hippocampal neurons, the cultures were pretreated with tSDA and GRb1 on 10d for 24 hours respectively. Then the cultures were treated with 35 μmol·L -1 Aβ25-35 for 24 hours, observed the changing of survival rate of neurons and the apoptosis of neurons with biochemical analysis combining immunofluorescent cytochemical double-staining technique. Results Hippocampal neurons were treated with 35 μmol·L -1 Aβ for 24 hours, and survival rate of neurons downed to 52.6%. When neurons were pretreated by tSDA and GRb1, survival rate of neurons increased 11% to 15%. The findings of immunofluorescent cytochemical double-staining indicated that apoptotic neurons were obviously more than that of the blank group, reaching 43.9%.When neurons were pretreated by tSDA and GRb1, apoptotic neurons were downed to 16.6%, 10.8% respectively. Conclusion tSDA had the same effects as GRb1, protecting the neurons, antagonizing neurotoxicity of Aβ, increasing survival rate of neurons, and reducing apoptotic neurons induced by Aβ.

Comparison ofβ-Amyloid Plaque Labeling Methods:Antibody Staining,Gallyas Silver Staining,and Thioflavin-S Staining

Objective To evaluate senile plaque formation and compare the sensitivity of three differentβ-amyloid(Aβ)labeling methods(antibody staining,Gallyas silver staining,and thioflavin-S staining)to detect Aβdeposition.Methods APPswe/PSEN1dE9 transgenic mice(APP/PS1)of different ages were used to examine spatiotemporal changes in Aβplaque deposition.Antibody staining,Gallyas silver staining,and thioflavin-S staining were used to detect Aβplaque deposition in the same brain region of adjacent slices from model mice,and the results were compared.Results With aging,Aβplaques first appeared in the cortex and then the deposition increased throughout the whole brain.Significantly greater plaque deposition was detected by 6E10 antibody than that analyzed with Gallyas silver staining or thioflavin-S staining(P<0.05).Plaque deposition did not show significant difference between the APP/PS1 mice brains assayed with Gallyas silver staining and ones with thioflavin-S staining(P=0.0033).Conclusions The APP/PS1 mouse model of Alzheimer’s disease could mimick the progress of Aβplaques occurred in patients with Alzheimer’s disease.Antibody detection of Aβdeposition may be more sensitive than chemical staining methods.

Mesenchymal stem cells:As a multi-target cell therapy for clearingβ-amyloid deposition in Alzheimer’s disease

Extracellularβ-amyloid(Aβ)plaques and neurofibrillary tangles(NFTs)are the pathological hallmarks of Alzheimer’s disease(AD).Studies have shown that aggregates of extracellular Aβcan induce neuroinflammation mediated neurotoxic signaling through microglial activation and release of pro-inflammatory factors.Thus,modulation of Aβmight be a potential therapeutic strategy for modifying disease progression.Recently,a large number of reports have confirmed the beneficial effects of mesenchymal stem cells(MSCs)on AD.It is believed to reduce neuroinflammation,reduce Aβamyloid deposits and NFTs,increase acetylcholine levels,promote neurogenesis,reduce neuronal damage,and improve working memory and cognition.In this review,we focus on the role of MSCs in clearing Aβdeposition.MSCs have the potential to modulate Aβ-related microenvironments via enhancement of autophagy,proteolysis of Aβaggregates,phagocytic clearance of Aβby microglial M2 polarization,decrease oxidative stress(OS),and correction of abnormal sphingolipid(SL)metabolism.With advantages in clinical applications,these data suggest that the use of MSCs as a multi-target modulator of Aβwould be an effective therapeutic approach in AD.

Multifaceted neuroprotective effects of(-)-epigallocatechin-3-gallate(EGCG)in Alzheimer's disease:an overview of pre-clinical studies focused onβ-amyloid peptide

Alzheimer’s disease(AD)is the most common neurodegenerative disease characterized by cognitive decline and memory impairment.Many lines of evidence indicate that excessiveβ-amyloid peptide(Aβ)generation and aggregation play pivotal roles in the initiation of AD,leading to various biochemical alteration including oxidative damage,mitochondrial dysfunction,neuroinflammation,signaling pathway and finally resulting in neuronal death.AD has a complex pathogenic mechanism,and a single-target approach for anti-AD strategy is thus full of challenges.To overcome these limitations,the present study focused to review on one of multiple target-compounds,(-)-epigallocatechin-3-gallate(EGCG)for the prevention and treatment of AD.EGCG is a main bioactive polyphenol in green tea and has been reported to exert potent neuroprotective properties in a wide array of both cellular and animal models in AD.This review demonstrated multiple neuroprotective efficacies of EGCG by focusing on the involvement of Aβ-evoked damage and its Aβregulation.Furthermore,to understand its mechanism of action on the brain,the permeability of the blood-brain barrier was also discussed.

Effects of Yizhi Capsule (益智胶囊) on Learning and Memory Disorder and β-amyloid Peptide Induced Neurotoxicity in Rats

To explore the effects of Yizhi Capsule （益智胶囊, YZC） on learning and memory disorder and β-amyloid peptide induced neurotoxicity in rats. Methods： Various doses of YZC were administered to Sprague-Dawley （SD） rats for 8 consecutive days, twice a day. On the 8th day of the experiment, scopolamine hydrobromide was intraperitoneally injected to every rat and Morris water maze test and shuttle dark avoidance test were carried out respectively to explore the changes of learning and memory capacities in the rats. Resides, after the cerebral cortical neurons of newborn SD rats aged within 3 days were cultured in vitro for 7 days, drug serum containing YZC was added to the cultured neurons before or after β amyloid peptide25-35 （Aβ25-35） intoxication to observe the protective effect of YZC on neurotoxicity by MTT assay and to determine the LDH content in the supernatant. Results： Compared with those untreated with YZC, the rats having received YZC treatment got superiority in shorter time of platform seeking in Morris water maze test, as well as elongated latent period and less times of error in shuttle dark avoidance test. On the cultured neurons, YZC drug serum could effectively increase the survival rate of Aβ25-35 intoxicated neurons and reduce the LDH contents in cultured supernatant. Conclusion： YZC has an action of improving learning and memory disorder, and good protective effect on Aβ25-35 induced neurotoxicity in SD rats. KEY WORDS

β-amyloid诱异神经细胞凋亡的研究

Aβ与AD的发生密切相关，然而其致神经元丢失的机制仍不清楚。该实验通过DNA末端标记及电子显微镜超微结构观察研究了Aβ透导培养的神经元细胞损害的形态学和分子生化改变．结昊显示：Aβ可诱导神经妊胞核DNA发生降解，出现染色质浓缩成块状，胞浆浓缩，胞膜内陷，调亡小体形成等。提示Aβ在AD中可能是通过诱导神经元凋亡而引起砷经元丢失的。

Phytochemical composition of Caesalpinia crista extract as potential source for inhibiting cholinesterase and β-amyloid aggregation: Significance to Alzheimer's disease

Objective: To screen plant extract fractions and elucidate the components present in Caesalpinia crista(C. crista) leaves for cholinergic and anti-amyloidogenic activities for the treatment of Alzheimer's diseases. Methods: This work has been carried out to study the action of C. crista extracts from nonpolar to polar solvents toward inhibition of oxidative stress, cholinergic and amyloidosis. The antioxidant activity was studied using DPPH total antioxidant assay; cholinergic assay by Ellman's method and anti-amyloidogenic assay by thioflavin-T fluorescence and transmission electron microscopy. Results: The quantification of polyphenols was carried out following C. crista methanolic extract(CCMeOH) HPLC fingerprinting, along with LC-MS and elucidated by MS LAMPS database. GC-MS of CCMeOH was screened for potential moieties. In vitro experimental results showed that the CCMe OH was potential extract that exhibited active inhibition of antioxidant property, cholinergic enzymes acetylcholinesterase and butyrylcholinesterase. For anti-amyloidogenic evaluations, among all the extracts, the CCMe OH was found to have the potential toward inhibiting the oligomers, fibrillation of Aβ42 with good defibrillation of amyloid cascading properties. Conclusions: These results are also supported by the presence of polyphenols as the active ingredients. Multi-potent target drug therapy is a promising option in treating the Alzheimer's diseases. Methanolic extract of C. crista shows potential activity against cholinergic enzymes, Aβ42 aggregation with antioxidant activity.

The β-amyloid protein induces S100β expression in rat hippocampus through a mechanism that involves IL-1

Objective To explore the effect of β-amyloid protein (Aβ) on S100β expression in rat hippocampus and its mechanisms. Methods At 7 days after bilateral stereotaxis injection of different dose of fibrillar Aβ 25-35 and interluekin-1 receptor antagonist (IL-1ra) into the rat CA1 region, the learning and memory abilities of rats were tested with passive avoidance task. Amyloid deposition was detected by using Congo red staining technique. Nissl staining and immunohistochemical techniques were used to analyze the number of neurons, and GFAP and the S100β expression in hippocampal CA1 region , respectively. Results After fibrillar Aβ injection, the step-through latency of rats was significantly shortened compared to that of the control group. The GFAP positive astrocytes were found surrounding amyloid deposition. Neuronal loss occurred in the pyramidal cell layer of CA1 region. The number of S100β positive cells in Aβ-treated group was significantly increased compared with that in the control group. After IL-1ra injection, the number of S100β positive cells was significantly decreased. Conclusion Intrahippocampal injection of Aβ 25-35 could cause similar pathologic changes of Alzheimer’s disease. Aβ 25-35 was capable of up-regulating S100β expression in a dose-dependent manner. The injection of IL-1ra could attenuate the effect of Aβ on S100β expression.

STUDY ON THE THERAPEUTIC EFFECTS OF GINSENOSIDE Rg-1 AND GASTRODINE ON AD MODEL RATS INDUCED BY β-AMYLOID PEPTIDE (25-35)

Objective To study the therapeutic effects of Ginsenoside Rg-1 and Gastrodine on rats model of Alzheimer's disease(AD). Methods Aggregated β-Amyloid peptide (25-35) was injected into the lateral ventricle of rats to establish AD models. Ginsenoside Rg-1, Gastrodine and Ginsenoside Rg-1+Gastrodine were intraperitoneally injected into rats of each test group(Ginsenoside Rg-1∶10mg/kg·day; Gastrodine 100mg/kg·day) for 4 weeks, the rats of control group received equal volume of saline. Passive avoidance task and Morris maze test were done to assess the ability of learning and memory. The content of superoxide dismutase (SOD), malondiadehyde (MDA), total-antioxidative capability (T-AOC), Choline acetyltransferase (ChAT) and acetylcholinesterase (AchE) in brain tissue were measured. Results Ginsenoside Rg-1 and Gastrodine significantly improved learning and memory deficits in the rats with AD induced by β-Amyloid peptide (25-35) (P<0.05). Ginsenoside Rg-1+Gastrodine group were better than Ginsenoside Rg-1 group and Gastrodine group (P<0.05). Ginsenoside Rg-1 reduced the increase of SOD, MDA, but inhibited the decrease of T-AOC, AchE and ChAT; Gastrodine reduced the increase of SOD, MDA, while inhibited the decrease of T-AOC. Gastrodine could also prevent the activity of ChAT and AchE decline in AD rats. Conclusion Both Ginsenoside Rg-1 and Gastrodine have therapeutic effects on rats with AD; Ginsenoside Rg-1 and Gastrodine injection at the same time were better than only using one of them. Their mechanisms might different. Ginsenoside Rg-1 can not only inhibit peroxidation but also increase the activity of AchE and ChAT in brain tissue, while Gastrodine can inhibit peroxidation only, but it can't prevent the decline of ChAT and AchE activity in AD rats.

Effect of Bak Foong Pills on the expression of β-amyloid in rat retina with optic nerve transection

AIM:To investigate the effect of Bak Foong Pills(BFP) on the expression of β-amyloid(Aβ) in rats retina with optic nerve transaction,and its roles and possible mechanisms in protecting optic nerve damage.· METHODS:Seventy-two healthy,Sprague-Dawley,adult rats were randomly assigned to three groups:negative control group(control group),optic nerve transection group(model group) and BFP treatment group(BFP group,100μg/mL) followed by establishing optic nerve transection model.The expression of Aβ was measured at 48 hours by Western-blotting.Moreover,the expressions of Bcl-2,Bax and Caspase-3 mRNA were evaluated at 48 hours by reverse transcriptase polymerase chain reaction(RT-PCR).RESULTS:There were significant differences among the control,model and BFP groups in the expression of Aβ(all P <0.01).Aβ expression was significantly higher in the model and BFP groups than that in the control group(P < 0.01),with a more significant reduction in the BFP group than that in the model group(P <0.01).Moreover,there were also significant differences among the three groups in the expressions of Bcl-2/Bax(Bcl-2:anti-apoptotic;Bax:proapoptotic) and Caspase-3 mRNA(proapoptotic)(all P<0.01).Bcl-2/Bax ratio was significantly lower and Caspase-3 mRNA expression was significantly higher in the model and BFP groups than those in the control group(P <0.01),with a significant growing of Bcl-2/Bax and reduction of Caspase-3 in the BFP group than those in the model group(P<0.01).· CONCLUSION:BFP can down-regulate Aβ expression in retina and may inhibit apoptosis and protect optic nerve by enhancing Bcl-2/Bax ratio and inhibiting Caspase-3 pathway.

Experimental Analysis of Interacting HT22 Plasma Membrane Cholesterol and β-Amyloid

The peptide β-Amyloid (β-A) is known to be one of the primary factors causing neurodegeneration in the Alzheimer disease. Hence, one would like to know the factors that would increase or decrease the toxicity of β-Amyloid in the brain. One of the factors that are debated in the literature is cholesterol, where it is not clear if modulating the levels of cholesterol would affect the degree of toxicity of β-Amyloid on neuron cells in the brain. In order to investigate this problem, data were collected and analyzed for three types of experiments: 1) Correspondence between cholesterol and methyl-β-cyclodextrin (MβCD) measurements;2) measurements of the relative fluorescence unit (RFU) with respect to MβCD concentration (with/without β-A);and 3) RFU measurements with respect to β-A concentration (with/without MβCD). HT22 hippocampal neurons immortalized with the simian virus SV-40 large T-antigen plasmid vector were used to conduct the experiments. Mito-ID Membrane potential cytotoxicity was used as a measure of mitochondrial potential change. The statistical analysis of the presented experimental results indicates that cholesterol has no statistically significant effect on the degree of toxicity of β-Amyloid.

NKX6.3 protects against gastric mucosal atrophy by downregulatingβ-amyloid production

BACKGROUND Atrophic gastritis is characterized by loss of appropriate glands and reduction in gastric secretory function due to chronic inflammatory processes in gastric mucosa. Moreover, atrophic gastritis is considered as a precancerous condition of gastric cancer. However, little is known about the molecular mechanism underlying gastric mucosal atrophy and its contribution to gastric carcinogenesis.Thus, we hypothesized that transcription factor NKX6.3 might be involved in maintaining gastric epithelial homeostasis by regulating amyloid β(Aβ)production.AIM To determine whether NKX6.3 might protect against gastric mucosal atrophy by regulating Aβ production.METHODS We identified NKX6.3 depletion induced cell death by cell count and Western blot assay. Production and mechanism of Aβ oligomer were analyzed by enzymelinked immunosorbent assay, Western blot, immunoprecipitation, real-timequantitative polymerase chain reaction and immunofluorescence analysis. We further validated the correlation between expression of NKX6.3, Helicobacter pylori CagA, Aβ oligomer, apolipoprotein E(ApoE), and β-secretase 1(Bace1) in 55 gastric mucosae.RESULTS NKX6.3 depletion increased both adherent and floating cell populations in HFE-145 cells. Expression levels of cleaved caspase-3,-9, and poly ADP ribose polymerase were elevated in floating HFE-145^(shNKX6.3) cells. NKX6.3 depletion produced Aβ peptide oligomers, and increased expression of ApoE, amyloid precursor protein, Aβ, Bace1, low-density lipoprotein receptor, nicastrin, high mobility group box1, and receptor for advanced glycosylation end product proteins. In immunoprecipitation assay, γ-secretase complex was stably formed only in HFE-145^(shNKX6.3) cells. In gastric mucosae with atrophy, expression of Aβpeptide oligomer, ApoE, and Bace1 was detected and inversely correlated with NKX6.3 expression. Treatment with recombinant Aβ 1-42 produced Aβoligomeric forms and decreased cell viability in HFE-145^(shNKX6.3) cells. Additionally,NKX6.3 depletion increased expression of inflammatory cytokines and cyclooxygenase-2.CONCLUSION NKX6.3 inhibits gastric mucosal atrophy by regulating Aβ accumulation and inflammatory reaction in gastric epithelial cells.

Key gene and protein changes in the beta-amyloid pathway following Longyanshen polysaccharides treatment in a mouse model of Alzheimer's disease

BACKGROUND： During onset and development of Alzheimer＇s disease, β-amyloid （Aβ） precursor protein （APP）, β-site amyloid precursor protein cleaving enzyme （BACE）, and β-amyloid are key genes and proteins in the Aβ pathway, and over-expression of these genes can lead to Aβ deposit/on in the brain. OBJECTIVE： To observe the influence of Longyanshen polysaccharides on expression of BACE, APP, and Aβ in the senescence-accelerated mouse prone/8 （SAMP8） brain, and to compare these effects with huperzine A treatment. DESIGN, TIME AND SETTING： A randomized, controlled, neurobiochemical experiment was performed at the Department of Pharmacology and Scientific Experimental Center of Guangxi Medical University from September 2005 to January 2008. MATERIALS： Longyanshen polysaccharfdes powder was extracted from the dried slices of the medicinal plant Longyanshen. The active component, Longyanshen polysaccharides, was provided by the Department of Pharmacology, Guangxi Medical University; huperzine A was purchased from Yuzhong Drug Manufactory, China. METHODS： Healthy SAMP8 mice were used to establish a model of Alzheimer＇s disease. A total of 50 SAMP8 mice were randomly assigned to 5 groups （n = 10）： SAMP8, huperzine A, low-, middle-, and high-dose polysaccharides. In addition, 10 senescence-accelerated mouse resistant 1 （SAMR1） mice were selected as normal controls. SAMP8 and SAMR1 mice were administered 30 mL/kg normal saline; the huperzine A group was administered 0.02 mg/kg huperzine A; the low-, middle-, and high-dose polysaccharides groups were respectively administered 45, 90, and 180 mg/kg Longyanshen polysaccharides. Each group was treated by intragastric administration, once per day, for 50 consecutive days. MAIN OUTCOME MEASURES： One hour after the final administration, immunohistochemical analysis was used to determine Aβ expression in the cortex and hippocampus of SAMP8 mice. Reverse-transcription polymerase chain reaction was used to determine mRNA levels of BACE and APP in SAMP8 brain tissue. RESULTS： Compared with the SAMR1 group, Aβ expression in the cerebral cortex and hippocampus, as well as expression of BACE, APP mRNA in the brain was significantly increased in the SAMP8 group （P 〈 0.05-0.01）. Compared with the SAMP8 group, Aβ expression, as well as BACE and APP mRNA expression, were significantly decreased in the cerebral cortex and hippocampus of huperzine A and low-, middle-, and high-dose polysaccharides groups （P 〈 0.05-0.01）. In particular, the effect of high-dose polysaccharides was the most significant （P 〈 0.05-0.01 ）. CONCLUSION： Longyanshen polysaccharides reduced or inhibited over-expression of BACE, APP, and Aβ in SAMP8 mice in a dose-dependent manner, and the effect was not worse than huperzine A.

Beta-amyloid precursor protein cleavage enzyme-1 expression in adult rat retinal neurons in the early period after lead exposure

Previous studies have reported that non-human primates and rodents exposed to lead during brain development may become dependent on the deposition of pre-determined β-amyloid protein （Aβ）,and exhibit upregulation of β-site amyloid precursor protein expression in old age.However,further evidence is required to elucidate the precise relationship and molecular mechanisms underlying the effects of early lead exposure on excessive Aβ production in adult mammals.The present study investigated the effects of lead exposure on expression of β-amyloid precursor protein cleavage enzyme-1 （BACE-1） in the rat retina and the production of Aβ in early development,using the retina as a window for studying Alzheimer＇s disease.Adult rats were intraocularly injected with different doses of lead acetate （10μmol/L,100μmol/L,1 mmol/L,10 mmol/L and 100 mmol/L）.The results revealed that retinal lead concentration,BACE-1 and its cleavage products β-C-terminal fragment and retina Aβ1-40 were all significantly increased in almost all of the lead exposure groups 48 hours later in a dose-dependent manner.The only exception was the 10μmol/L group.The distribution of BACE-1 in the retina did not exhibit obvious changes,and no distinctive increase in the activation of retinal microglia was apparent.Similarly,retinal synaptophysin expression did not exhibit any clear changes.These data suggest that lead exposure can result in the upregulation of retinal neuron BACE-1 expression in the early period of development and further increase the overproduction of Aβ1-40 in the retina.Our results provided novel insight into the molecular mechanisms underlying environmentally-induced Alzheimer＇s disease.