Neuroprotective effects of ZL006 in Aβ1–42-treated neuronal cells

Amyloid beta (Aβ)-induced neurotoxicity and oxidative stress plays an important role in the pathogenesis of Alzheimer’s disease (AD). ZL006 is shown to reduce over-produced nitric oxide and oxidative stress in ischemic stroke by interrupting the interaction of neuronal nitric oxide synthase and postsynaptic density protein 95. However, few studies are reported on the role of ZL006 in AD. To investigate whether ZL006 exerted neuroprotective effects in AD, we used Aβ_1–42 to treat primary cortical neurons and N2a neuroblastoma cells as an in vitro model of AD. Cortical neurons were incubated with ZL006 or dimethyl sulfoxide for 2 hours and treated with Aβ_1–42 or NH₃•H₂O for another 24 hours. The results of cell counting Kit-8 (CCK-8) assay and calcein-acetoxymethylester/propidium iodide staining showed that ZL006 pretreatment rescued the neuronal death induced by Aβ_1–42. Fluorescence and western blot assay were used to detect oxidative stress and apoptosis-related proteins in each group of cells. Results showed that ZL006 pretreatment decreased neuronal apoptosis and oxidative stress induced by Aβ_1–42. The results of CCK8 assay showed that inhibition of Akt or NF-E2-related factor 2 (Nrf2) in cortical neurons abolished the protective effects of ZL006. Moreover, similar results were also observed in N2a neuroblastoma cells. ZL006 inhibited N2a cell death and oxidative stress induced by Aβ_1–42, while inhibition of Akt or Nrf2 abolished the protective effect of ZL006. These results demonstrated that ZL006 reduced Aβ_1–42-induced neuronal damage and oxidative stress, and the mechanisms might be associated with the activation of Akt/Nrf2/heme oxygenase-1 signaling pathways.

Chinese Library Classification No. R459.9; R453; R364     

In pursuit of a sensitive EEG functional connectivity outcome measure for clinical trials in Alzheimer’s disease

ObjectiveIn clinical trials in Alzheimer’s Disease (AD), an improvement of impaired functional connectivity (FC) could provide biological support for the potential efficacy of the drug. Electroencephalography (EEG) analysis of the SAPHIR-trial showed a treatment induced improvement of global relative theta power but not of FC measured by the phase lag index (PLI). We compared the PLI with the amplitude envelope correlation with leakage correction (AEC-c), a presumably more sensitive FC measure.MethodsPatients with early AD underwent 12 weeks of placebo or treatment with PQ912, a glutaminylcyclase inhibitor. Eyes-closed task free EEG was measured at baseline and follow-up (PQ912 n = 47, placebo n = 56). AEC-c and PLI were measured in multiple frequency bands. Change in FC was compared between treatment groups by using two models of covariates.ResultsA significant increase in global AEC-c in the alpha frequency band was found with PQ912 treatment compared to placebo (p = 0.004, Cohen’s d = 0.58). The effect remained significant when corrected for sex, country, ApoE ε4 carriage, age, baseline value (model 1; p = 0.006) and change in relative alpha power (model 2; p = 0.004).ConclusionsFunctional connectivity in early AD, measured with AEC-c in the alpha frequency band, improved after PQ912 treatment.SignificanceAEC-c may be a robust and sensitive FC measure for detecting treatment effects.     

黄芩茎叶总黄酮对AD大鼠海马神经元的保护作用及机制研究

目的：探讨黄芩茎叶总黄酮(SSTF)对大鼠双侧海马注射Aβ25－35引起的大鼠学习记忆功能和海马神经元形态变化的影响及机制。方法：将30只雄性Wistar大鼠随机分为对照组、模型组、总黄酮组,对照组及模型组灌胃纯化水,qd,d 8对照组海马注射生理盐水,继续灌胃;模型组双侧海马注射Aβ25－35（5 μL）,其他同对照组;给药组总黄酮（50 mg?kg-1,ig,qd),d 8海马注射Aβ,继续灌胃,三组于d 15采用Morris水迷宫实验观察大鼠学习记忆能力,测5 d,处死,硫堇Nissl染色观察海马神经元的变化,检测血清中丙二醛（MDA）含量。结果：模型组大鼠逃避潜伏期较对照组明显延长（P<0.05）,给药组大鼠逃避潜伏期较模型组明显缩短（P<0.05）;模型组海马CA1区局部神经细胞带脱失,脱失处胶质细胞增多,给药组细胞损伤较轻;模型组大鼠血清中MDA显著高于对照组（P<0.05）,给药组血清中MDA明显低于模型组（P<0.05）。结论：黄芩茎叶总黄酮对海马注射Aβ25－35引起大鼠学习记忆能力降低及海马神经元形态变化具有保护作用,其机制可能是减少Aβ引起的脂质过氧化产物增多引起的氧化应激,并可对抗Aβ引起的胶质细胞增多。     

Amyloid-β and tau synergistically impair the oxidative phosphorylation system in triple transgenic Alzheimer's disease mice

Alzheimer''s disease (AD) is characterized by amyloid-beta (Aβ)-containing plaques, neurofibrillary tangles, and neuron and synapse loss. Tangle formation has been reproduced in P301L tau transgenic pR5 mice, whereas APP^swPS2^N141I double-transgenic APP152 mice develop Aβ plaques. Cross-breeding generates triple transgenic (^tripleAD) mice that combine both pathologies in one model. To determine functional consequences of the combined Aβ and tau pathologies, we performed a proteomic analysis followed by functional validation. Specifically, we obtained vesicular preparations from ^tripleAD mice, the parental strains, and nontransgenic mice, followed by the quantitative mass-tag labeling proteomic technique iTRAQ and mass spectrometry. Within 1,275 quantified proteins, we found a massive deregulation of 24 proteins, of which one-third were mitochondrial proteins mainly related to complexes I and IV of the oxidative phosphorylation system (OXPHOS). Notably, deregulation of complex I was tau dependent, whereas deregulation of complex IV was Aβ dependent, both at the protein and activity levels. Synergistic effects of Aβ and tau were evident in 8-month-old ^tripleAD mice as only they showed a reduction of the mitochondrial membrane potential at this early age. At the age of 12 months, the strongest defects on OXPHOS, synthesis of ATP, and reactive oxygen species were exhibited in the ^tripleAD mice, again emphasizing synergistic, age-associated effects of Aβ and tau in perishing mitochondria. Our study establishes a molecular link between Aβ and tau protein in AD pathology in vivo, illustrating the potential of quantitative proteomics.     

大鼠阿尔茨海默病模型中溶酶体组织蛋白酶D的表达及其意义

目的：观察阿尔茨海默病（AD）大鼠皮质神经元中溶酶体蛋白酶Cathepsin D的表达。方法采用β-淀粉样肽（Aβ）大鼠海马注射制作AD动物模型,用免疫组织化学染色和Western-blot印迹法检测大鼠颞底皮层Cathepsin D的表达。Y迷宫检测大鼠空间辨别性能和学习记忆能力。TUNEL法检测细胞凋亡情况。结果Aβ脑池内灌注造成Aβ沉积的AD模型在行为学和病理学改变上一定程度地模拟了AD。免疫组化和Werstern-bolt检测均显示实验组Cathepsin D阳性神经元数量较假手术组和正常组明显增加（P＜0.05）。TUNEL法检测显示AD大鼠颞底皮层神经元凋亡与正常组和假手术组相比显著升高（P＜0.05）。结论Cathepsin D在AD大鼠皮层脑组织中表达升高,可能参与了神经元凋亡等病理过程。     

Could Pin1 help us conquer essential hypertension at an earlier stage? A promising early-diagnostic biomarker and its therapeutic implications for the disease

Essential hypertension is a major risk factor for cardiovascular morbidity and mortality, and the early-diagnosis is very important for the prevention of essential hypertension. Previously, we found that Pin1, the only known enzyme isomerizing pSer/pThr-Pro motifs in proteins, may gradually become inactive under conditions of stress such as intracellular acidification and fever. Interestingly, essential hypertension and the dysfunction of Pin1 often synchronously occur with the increasing age. Recent evidence indicates that Pin1 primarily increases the activity of endothelial nitric oxide synthase (eNOS) and the production of nitric oxide (NO) in multiple ways, significantly promoting the relaxation response of blood vessels and preventing the elevation of blood pressure. Further, the inhibition of Pin1 results in significantly increased blood pressure in rats. So, we hypothesized and evaluated the potential of Pin1 to be a new early-diagnostic biomarker as well as a therapeutic drug for essential hypertension. The unique activity of Pin1 and some epidemiological and experimental data evidence that the decreased activity of Pin1 may be closely associated with the development of essential hypertension. The factors that may impact the activity of Pin1 and correlate with the risk of essential hypertension were also discussed. These findings indicate that Pin1 plays a key and permanent role in efficiently preventing the development of essential hypertension, and that Pin1 may be a promising early-diagnostic biomarker as well as an effective therapeutic drug for the early-diagnosis, prevention, and treatment of essential hypertension, potentially decreasing the risk of cardiovascular morbidity and mortality.     

黄芩茎叶总黄酮对AD模型大鼠学习、记忆能力及神经元损伤的影响

目的探讨黄芩茎叶总黄酮（SSTF）对AD的防治作用。方法将30只雄性Wistar大鼠随机分为三组各10只,观察组予SSTF100mg／（kg&#183;d）灌胃1次／d,第8天双侧海马注射凝胶态淀粉样蛋白（Aβ）25-35 5μl（10μg）,继续SSTF灌胃;对照组及模型组予蒸馏水灌胃1次／d,第8天双侧海马分别注射生理盐水及Aβ23-35,5μl（10μg）,继续灌胃。三组均于第15天采用Morris水迷宫试验观察大鼠学习记忆能力（逃避潜伏期时间）,连续观察5d后处死,硫堇Nissl染色观察海马神经元变化。结果模型组及观察组大鼠逃避潜伏期较对照组明显延长,观察组明显短于模型组;模型组海马CAI区神经细胞带脱失、不完整,脱失处胶质细胞增多,观察组细胞损伤较轻。结论SSTF可明显改善Aβ25-35所致AD大鼠学习、记忆能力,可能机制为减少Aβ所产生的氧自由基,减轻胶质细胞增殖所致损害。     

Screening for a human single chain Fv antibody against epitope on amyloid-beta 1-40 from a human phage display library

Amyloid-beta peptides （Aβ） are believed to be .responsible for the mental decline in patients with Alzheimer＇s reported that pathology in vaccination disease （AD）. In 1999, Schenk et all immunization with Aβ attenuated AD-like the PDAPP mouse, and developed a new approach to AD. Such vaccines were successfully tested in mouse models of AD for the reduction of Aβ plaque burden and the improvement of cognitive performance. However, 6% of AD patients developed symptoms of brain inflammation after vaccination that resembled enceohalitis or meningitis.     

GPR120 Signaling Controls Amyloid-β Degrading Activity of Matrix Metalloproteinases

Alzheimer''s disease (AD) is characterized by the extensive deposition of amyloid-β peptide (Aβ) in the brain. Brain Aβ level is regulated by a balance between Aβ production and clearance. The clearance rate of Aβ is decreased in the brains of sporadic AD patients, indicating that the dysregulation of Aβ clearance mechanisms affects the pathologic process of AD. Astrocytes are among the most abundant cells in the brain and are implicated in the clearance of brain Aβ via their regulation of the blood–brain barrier, glymphatic system, and proteolytic degradation. The cellular morphology and activity of astrocytes are modulated by several molecules, including ω3 polyunsaturated fatty acids, such as docosahexaenoic acid, which is one of the most abundant lipids in the brain, via the G protein-coupled receptor GPR120/FFAR4. In this study, we analyzed the role of GPR120 signaling in the Aβ-degrading activity of astrocytes. Treatment with the selective antagonist upregulated the matrix metalloproteinase (MMP) inhibitor-sensitive Aβ-degrading activity in primary astrocytes. Moreover, the inhibition of GPR120 signaling increased the levels of Mmp2 and Mmp14 mRNAs, and decreased the expression levels of tissue inhibitor of metalloproteinases 3 (Timp3) and Timp4, suggesting that GPR120 negatively regulates the astrocyte-derived MMP network. Finally, the intracerebral injection of GPR120-specific antagonist substantially decreased the levels of TBS-soluble Aβ in male AD model mice, and this effect was canceled by the coinjection of an MMP inhibitor. These data indicate that astrocytic GPR120 signaling negatively regulates the Aβ-degrading activity of MMPs.SIGNIFICANCE STATEMENT The level of amyloid β (Aβ) in the brain is a crucial determinant of the development of Alzheimer''s disease. Here we found that astrocytes, which are the most abundant cell type in the CNS, harbor degrading activity against Aβ, which is regulated by GPR120 signaling. GPR120 is involved in the inflammatory response and obesity in peripheral organs. However, the pathophysiological role of GPR120 in Alzheimer''s disease remains unknown. We found that selective inhibition of GPR120 signaling in astrocytes increased the Aβ-degrading activity of matrix metalloproteases. Our results suggest that GPR120 in astrocytes is a novel therapeutic target for the development of anti-Aβ therapeutics.     

Amyloid-Beta Mediates Homeostatic Synaptic Plasticity

The physiological role of the amyloid-precursor protein (APP) is insufficiently understood. Recent work has implicated APP in the regulation of synaptic plasticity. Substantial evidence exists for a role of APP and its secreted ectodomain APPsα in Hebbian plasticity. Here, we addressed the relevance of APP in homeostatic synaptic plasticity using organotypic tissue cultures prepared from APP^−/− mice of both sexes. In the absence of APP, dentate granule cells failed to strengthen their excitatory synapses homeostatically. Homeostatic plasticity is rescued by amyloid-β and not by APPsα, and it is neither observed in APP^+/+ tissue treated with β- or γ-secretase inhibitors nor in synaptopodin-deficient cultures lacking the Ca²⁺-dependent molecular machinery of the spine apparatus. Together, these results suggest a role of APP processing via the amyloidogenic pathway in homeostatic synaptic plasticity, representing a function of relevance for brain physiology as well as for brain states associated with increased amyloid-β levels.