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.

Justicidin A Reduces β-Amyloid via Inhibiting Endocytosis of β-Amyloid Precursor Protein

β-amyloid precursor protein (APP) can be cleaved by α-, and γ-secretase at plasma membrane producing soluble ectodomain fragment (sAPPα). Alternatively, following endocytosis, APP is cleaved by β-, and γ-secretase at early endosomes generating β-amyloid (Aβ), the main culprit in Alzheimer's disease (AD). Thus, APP endocytosis is critical for Aβ production. Recently, we reported that Monsonia angustifolia, the indigenous vegetables consumed in Tanzania, improved cognitive function and decreased Aβ production. In this study, we examined the underlying mechanism of justicidin A, the active compound of M. angustifolia, on Aβ production. We found that justicidin A reduced endocytosis of APP, increasing sAPPα level, while decreasing Aβ level in HeLa cells overexpressing human APP with the Swedish mutation. The effect of justicidin A on Aβ production was blocked by endocytosis inhibitors, indicating that the decreased APP endocytosis by justicidin A is the underlying mechanism. Thus, justicidin A, the active compound of M. angustifolia, may be a novel agent for AD treatment.

3D-QSAR Analysis and Molecular Docking Study on Biaryl Aminothiazine BACE1 Inhibitor / 中国药房

OBJECTIVE:To provide theoretic basis for the design and synthesis of novel high-activity biaryl aminothiazineβ-amyloid precursor protein cleaving enzyme 1 (BACE1) inhibitor,the research and development of new AD therapy drugs. METHODS:Totally 41 molecules of biaryl aminothiazine BACE1 inhibitors were selected. By SYBYL-X 2.0 software package, CoMFA and CoMSIA method were used to construct 3D-QSAR model of derivatized compounds. Surflex-dock molecular docking was applied to analyze binding mode of the compounds with BACE1. RESULTS:The q2 value of 3D-QSAR model established by CoMFA and CoMSIA method were all higher than 0.5,indicating good predictability. The established three dimensional contour plots could manifest the effect of substituents at different sites on activity of compounds. Surflex-dock analysis showed that biaryl aminothiazine and amino acid residues as ASP80, ASP276 and TYR246 in BACE1 had a key effect on hydrogen bonds. CONCLUSIONS:3D-QSAR model established on the basis of biaryl aminothiazine derivatized compounds show good predictability,which provides guidance for the structure optimization of the compound. TYR246 may be another potential active functional residue of biaryl aminothiazine inhibitor compound molecule combined with BACE1. Through 3D-QSAR analysis and molecular docking,new biaryl aminothiazine BACE1 inhibitor can be designed and synthesized so as to research and develop new drugs for AD.

The relationship between differentially expressed miR-19b-3p and cognitive function in patients with Alzheimer's disease / 中华行为医学与脑科学杂志

Objective To explore the relationship between differentially expressed miR-19b-3p and cognitive function in patients with Alzheimer's disease (AD).Methods The miRNA expression profiles of patients with AD(AD group,n=30) or healthy elderly people (NC group,n=30) were analyzed by Illumina/Solexa high-throughput sequencing technique.The miRNA lentiviral plasmids were constructed and injected into the model of AD rats.The cognitive function of rats was analyzed with water maze test.The mimics and inhibitors were synthesized and transfected into SH-SY5Y cell lines.The protein expression of β-amyloid precursor protein cleavage enzyme 1 (BACE1) was detected by western blot.Results Compared with the NC group,the expression of miR-146a-5p (log2AD/Control =2.3),miR-195-5p (log2 AD/Control =10.2),miR-20b-5p(log2AD/Control =15.1) in serum of AD group was up-regulated and the expression of miR-19b-3p (log2 AD/Control =-8.0) and miR-125b-3p (log2 AD/Control =-15.3) was down-regulated (P＜ 0.05).Compared with the rats in the AD group((26.50±3.12) s),the rats in the AD+miR-19b-3p group ((15.33±2.78) s) showed significantly shorter escape latencies(P＜0.05).Compared with the NC group,the protein expression of BACE1 in miR-19b-3p mimetic group was decreased and the protein expression of BACE1 in inhibitor group was increased.Conclusion miR-19b-3p may improve the cognitive function of AD patients via regulating the expression of BACE1.

Brain injury and β-amyloid precursor protein, β-amyloid protein / 国际外科学杂志

As a transmembrane protein, β-amyloid precursor protein(β-APP) distributes extensively in the central nervous system, has the effect of neurotrophic, and neuroprotective, promote neurite growth and synaptogenesis, β-amyloid(Aβ) is the digestion products of its precursor-APP in the pathological conditions, and it is the main component of senile plaques-the main pathological changes of Alzheimer' s disease (AD), its toxic effects can also induce neuronal apoptosis, The expression of the two proteins after brain injuried has a close relationship with the injury, cognitive dysfunction, Alzheimer' s disease and the pathophysiological changes of central nervous system. To explore its expression in the brain after traumatic brain injury can determine the degree of injury, assess the prognosis and open up new avenues for the treatment of traumatic brain injury.

The Dynamical Changes of Serum β-APP in Early Stage of Rabbit Traumatic Brain Injury / 中国医科大学学报

Objective To explore the relationship of serum levels of β-amyloid precursor protein (β-APP) with degree of traumatic brain injury (TBI) and the traumatic time.Methods Sprague-Dawley (SD) rats were randomly divided into normal control and injury group.The rats in injury groups suffered from TBI after free-falling percussion with different pressure (wild-injury,moderate-injury and severe-injury group).Then serum was collected at 0.5 h,2 h,6 h,and 24 h and subject to β-APP detection by ELISA.All data were analyzed statistically with completely randomized design multiple factor repeated measure of variance analysis and least significant difference (LSD) test.Results The serum levels of β-APP were higher after injury.The serum levels of β-APP were significantly higher in moderate-injury or severe-injury group than those in normal group or slight-injury group (P＜0.05).The serum levels of β-APP were higher in severe-injury group than that in moderate-injury group with no statistical difference (P＞0.05).There was no statistical difference in serum β-APP levels between normal control and slight-injury group (P＞0.05).Conclusion The serum level of P-APP is increasingly higher with traumatic brain injury more serious and could be employed as an indicator of TBI degree.It implies that β-APP has the potential as an early diagnosis marker for TBI.

Up-regulation of β-amyloid precursor protein cleavage enzyme 1 and β-amyloid protein in brain with diabetic rats / 解剖学报

Objective To investigate expression of β-site APP-cleaving enzymel(BACE1) and Aβ in brain of diabetes mellitus of Wistar rats,to study pathophysiological mechanism of Alzheimer disease from diabetic metabolic disorder. Methods Animal model of diabetes mellitus was established by streptozocin with intraperitoneal injection. Wistar rats were randomly divided into normal group (N), sham-operation group (S), 4 weeks diabetes mellitus model group (M4), 6 weeks diabetes mellitus model group (M6) and 8 weeks diabetes mellitus model group (M8). Behaviour was tested with Morris water maze and shuttle box test. Expression of Aβ was measured by enzyme linked immunosorbent assay and BACE1 by immunohistochemistry, enzyme linked immunosorbent assay, Western blotting and RT-PCR. The absorbance value was measured by imaging analysis. Results The electric times and latancy of memory and study were more increased in model group than that in N and S group but the times of escape more decreased(P<0.01). The expression of Aβ_(1-40) increased from (64.13±6.76)pg/mg in normal group to (86.43±7.03)pg/mg in model group by ELISA(P<0.001) and Aβ_(1-42) from (67.43±5.12 )pg/mg in normal group to (89.45±5.28) pg/mg (P<0.001) in model group. The expression of BACE1 increased from (116.46±8.10)pg/mg in normal group to (158.73±6.24)pg/mg in model group by ELISA and from 0.61±0.11 in normal group to 1.52±0.16 by Western blotting absorbance valule and from 1.62±0.26 in normal group to 3.61±0.32 by RT-PCR absorbance valule and from 0.81±0.21 in normal group to 2.01±0.36 by immunohistochemistry absorbance valule (P<0.001). The expression of BACE1 and Aβ in MT group was higher than that of in N and S group (P<0.01). The level of BACE1 and Aβ had positive correlation with cognitive impairment.Conclusion The expression of BACE1 and Aβ is increased in diabetes mellitus rats. Diabetes mellitus contributes to Alzheimer diseases that.

Fluorescence imaging of APP in Alzheimer's disease with quantum dot or Cy3: a comparative study / 中南大学学报(医学版)

Objective To compare the fluorescence intensity and duration of qdots streptavidin conjugate (QDs-SA) with Cy3 as the molecular probe of β amyloid precursor protein (APP), and to provide evidence for early molecular imaging and diagnosis of Alzheimer's dissease (AD). Methods With the help of laser scanning confocal microscope and flow cytometry, the flurescence probe based on the QDs-SA was used to detect APP in HEK293 cells stably transfected pcDNA3.1/APP, and to compare with conventional fluroimmunoassay Cy3. Results The immunofluorescence staining detection indicated APP expression was mainly located in the plasma membrane. The mean fluorescence intensity of QDs-SA (34.2336±4.2455) was greater than that of Cy3 (21.6023±3.0102)under the confocal fluorescence microscope (P<0.05). After persistent exciting for 12 min, the fluorescence intensity of APP stained by QDs-SA decreased by 27.87%. The other stained by Cy3 decreased by 79.60%. The positive rate of APP staining had no significant difference between the QDs-SA(54.4700±3.4433)% and Cy3 (54.3800±8.5229)% by flow cytometry, but the mean fluorescence intensity had statistical significance(P<0.05). The QDs-SA (1 045.4167±47.3623) was significantly higher than the mean fluorescence intensity of Cy3 (658.5467±55.0591). Conclusion QDs-SA fluorescence probes can effectively recognize APP and are sensitive and exceptionally photostable, suggesting that QDs-SA fluorescence probes could be a potential method in APP detection and offer a novel way for the diagnosis of Alzheimer's disease.