AD Model Mice Research Articles

Novel imaging agents for β-amyloid plaque based on the N-benzoylindole core

We report the synthesis and evaluation of a series of N-benzoylindole derivatives as novel potential imaging agents for β-amyloid plaques. In vitro binding studies using Aβ(1-40) aggregates versus [(125)I]TZDM showed that all these derivatives demonstrated high binding affinities (K(i) values ranged from 8.4 to 121.6 nM). Moreover, two radioiodinated compounds [(125)I]7 and [(125)I]14 were prepared. Autoradiography for [(125)I]14 displayed intense and specific labeling of Aβ plaques in the brain sections of AD model mice (C57, APP/PS1) with low background. In vivo biodistribution in normal mice exhibited sufficient initial brain uptake for imaging (2.19% and 2.00%ID/g at 2 min postinjection for [(125)I]7 and [(125)I]14, respectively). Although additional modifications are necessary to improve brain uptake and clearance from the brain, the N-benzoylindole may be served as a backbone structure to develop novel β-amyloid imaging probes.

Compliance of patients with Alzheimer's disease to treatment with transdermal rivastigmine and its effectiveness and tolerability in daily practice

currently using mouse models of AD to test the therapeutic out come of these peptides to rescue the AD phenotypes. Methods: A modified peptide P5 derived from P35, an activator of Cdk5 was constructed and used to study the regulated and deregulated activity of Cdk5 in in vitro and in vivo using the AD model mice. Results: P5 peptide rescued the in situ phenotypes induced by neuronal stress in hippocampal and cortical neurons Conclusions: These studies suggest that P5 may be a potential candidate as a therapeutic reagent for AD and other neurodegenerative diseases induced due to Cdk5 deregulation.

Synthesis and evaluation of benzofuran-2-yl(phenyl)methanone derivatives as ligands for β-amyloid plaques

A series of benzofuran-2-yl(phenyl)methanone derivatives were synthesized and evaluated as novel probes for β-amyloid plaques. These derivatives were produced by a Rap–Stoermer condensation reaction. Compounds with a N, N-dimethylamino group displayed high affinity for Aβ 1–42 aggregates with K i values in the nanomolar range. Autoradiography with brain sections of AD model mice (APP/PS1) revealed that a radioiodinated probe, [ 125I] 10, labeled β-amyloid plaques selectively and displayed good brain uptake (3.53% ID/g) at 2 min. The results suggest that benzofuran-2-yl(phenyl)methanone derivatives should be investigated further as potential probes for detecting β-amyloid plaques in the AD brain.

Novel ( E)-5-styryl-2,2′-bithiophene derivatives as ligands for β-amyloid plaques

In continuation of our investigation on the bithiophene structure as potential β-amyloid probes, a series of ( E)-5-styryl-2,2′-bithiophene (SBTP) derivatives was designed and synthesized. In vitro binding showed that all of them displayed high binding affinities to A β 1–42 aggregates ( K i = 0.10–41.05 nM). Moreover, two radio-iodinated probes, [ 125I]-( E)-5-(4-iodostyryl)-2,2′-bithiophene ([ 125I] 8) and [ 125I]-( E)-5-iodo-5′-(4-methoxystyryl)-2,2′-bithiophene ([ 125I] 31) were prepared. Both of them displayed specific labeling of A β plaques in the brain sections of AD model mice with low background. In vivo biodistribution in normal mice indicated that [ 125I] 8 exhibited high initial brain uptake (2.11% ID/g at 2 min) and rapid clearance (0.41% ID/g at 30 min). These preliminary results suggest that SBTP derivatives may be served as novel β-amyloid imaging probes.

99mTc- and Re-labeled 6-dialkylamino-2-naphthylethylidene derivatives as imaging probes for β-amyloid plaques

Based on the conjugate strategy, two neutral (99m)Tc labeled 2-(1-(6-(dialkylamino)naphthalen-2-yl)ethylidene)malononitrile (DDNP) and 1-(6-(dialkylamino)naphthalen-2-yl)ethanone (ENE) derivatives, and their corresponding rhenium complexes were synthesized. In vitro fluorescent staining indicated that the corresponding rhenium derivatives selectively stained the β-amyloid (Aβ) plaques in the brain sections of AD model mice with low background. Compared with FDDNP and FENE, the affinities of the corresponding rhenium derivatives to Aβ aggregates decreased about 10-14-fold. In vivo biodistribution experiments in normal mice showed that (99m)Tc-MAMA-ENE displayed medium initial brain uptake (0.65%ID/g at 2min) with a reasonable washout from the brain (0.19%ID/g at 2h) while (99m)Tc-MAMA-DDNP showed a low brain uptake (0.28%ID/g at 2 min). Further optimize these (99m)Tc-labeled tracers in order to improve their binding affinities to Aβ plaques and diffusion through the blood brain barrier may generate useful imaging agents for SPECT.

Reduction of amyloid β-peptide accumulation in Tg2576 transgenic mice by oral vaccination

Alzheimer’s disease (AD) is pathologically characterized by the presence of extracellular senile plaques and intracellular neurofibrillary tangles. Amyloid β-peptide (Aβ) is the main component of senile plaques, and the pathological load of Aβ in the brain has been shown to be a marker of the severity of AD. Aβ is produced from the amyloid precursor protein by membrane proteases and is known to aggregate. Recently, immune-mediated cerebral clearance of Aβ has been studied extensively as potential therapeutic strategy. In previous studies that used a purified Aβ challenge in a mouse model of AD, symptomatic improvement was reported. However, a clinical Alzheimer’s vaccine trial in the United States was stopped because of severe side effects. Immunization with the strong adjuvant used in these trials might have activated an inflammatory Th1 response. In this study, to establish a novel, safer, lower-cost therapy for AD, we tested an oral vaccination in a wild-type and a transgenic mouse model of AD administered via green pepper leaves expressing GFP-Aβ. Anti-Aβ antibodies were effectively induced after oral immunization. We examined the immunological effects in detail and identified no inflammatory reactions. Furthermore, we demonstrated a reduction of Aβ in the immunized AD-model mice. These results suggest this edible vehicle for Aβ vaccination has a potential clinical application in the treatment of AD.

Consumption of Hydrogen Water Prevents the Age-dependent Impairment in Learning and Memory Tasks in AD Model Mice

Oxidative stress is a causative factor in the pathogenesis of major neurodegenerative diseases including AD and PD. Recently, we demonstrated that hydrogen molecule (H2) selectively reduced the hydroxyl radical, the most cytotoxic ROS, without affecting the other oxygen-derived free radicals (Ohsawa et al., Nat. Med. 2007). Inhalation of H2 gas markedly suppressed brain injury caused by ischemia-reperfusion. Moreover, consumption of water with dissolved H2 to a saturated level (H2 water) prevented stress-induced cognitive decline in mouse and 6-hydroxydopamine-induced neurodegeneration in a rat model of PD. To evaluate the effect of H2 on AD, we used transgenic mice (DAL and Tg2576). DAL mice express a dominant-negative form of mitochondrial aldehyde dehydrogenase2 in the brain and decrease ability to detoxify 4-hydroxynonenal (HNE) in their cortical neurons. Consequently, age-dependent neurodegeneration with cognitive impairment is observed (Ohsawa et al., J. Neurosci. 2008). H2 was dissolved in water under to a saturated level (0.8 mM). Each day, H2 water was placed into a closed glass vessel equipped with an outlet line containing two ball bearings, which kept the water from being degassed. H2 water degassed by gentle stirring was used for control. Mice were allowed to freely drink water with or without H2. Providing H2 water prevented the deficits in learning and memory of DAL mice at age 12 month (P < 0.0244) and age 16 month (P < 0.044), as examined by object recognition task. In the blood, providing H2 water decreased HNE, indicating that H2 water prevented the accumulation of oxidative stress. H2 has no known toxic effect on human body. A clinical trial showed the decrease in modifying low-density lipoprotein by drinking H2 water. Thus, we propose that H2, one of the most well-known molecules, could be widely used in medical applications in neurodegenerative diseases as a safe and effective antioxidant.

Effect of Synthetic Cannabinoid HU210 on Memory Deficits and Neuropathology in Alzheimers Disease Mouse Model

Cannabinoids have been shown to increase neurogenesis in adult brain, as well as protect neurons from excitotoxicity, calcium influx, inflammation, and ischemia. Recent studies have shown that synthetic cannabinoids can alleviate water maze impairments in rats treated with intracranial amyloid beta protein (Abeta); however it is unknown whether this effect is due to the cannabinoids' anti-inflammatory properties or whether it affects Abeta processing. Here we investigate whether cannabinoids have any effect on Alzheimer's disease in vivo. We found that HU210, a potent synthetic cannabinoid, did not improve water maze performance or a contextual fear conditioning task in an APP23/PS45 double transgenic mouse model of AD. HU210 had no effect on APP processing and Abeta generation, as well as neuritic plaque formation in the brains of AD transgenic mice. Our study showed that synthetic cannabinoid HU210 had no beneficial effects on AD neuropathology and behavioral deficits of AD model mice, which advises caution of such drug's application in AD therapies.

Synthesis and evaluation of novel benzothiazole derivatives based on the bithiophene structure as potential radiotracers for β-amyloid plaques in Alzheimer’s disease

In this study, six novel benzothiazole derivatives based on the bithiophene structure were developed as potential β-amyloid probes. In vitro binding studies using Aβ aggregates showed that all of them demonstrated high binding affinities with Ki values ranged from 0.11 to 4.64nM. In vitro fluorescent staining results showed that these compounds can intensely stained Aβ plaques within brain sections of APP/PS1 transgenic mice, animal model for AD. Two radioiodinated compounds [125I]-2-(5′-iodo-2,2′-bithiophen-5-yl)-6-methoxybenzo[d]thiazole [125I]10 and [125I]-2-(2,2′-bithiophen-5-yl)-6-iodobenzo[d]thiazole [125I]13 were successfully prepared through an iododestannylation reaction. Furthermore, in vitro autoradiography of the AD model mice brain sections showed that both [125I]10 and [125I]13 labeled the Aβ plaques specifically with low background. In vivo biodistribution studies in normal mice indicated that [125I]13 exhibited high brain uptake (3.42% ID/g at 2min) and rapid clearance from the brain (0.53% ID/g at 60min), while [125I]10 showed lower brain uptake (0.87% ID/g at 2min). In conclusion, these preliminary results of this study suggest that the novel radioiodinated benzothiazole derivative [125I]13 may be a candidate as an in vivo imaging agent for detecting β-amyloid plaques in the brain of AD patients.

Computed tomography of amyloid plaques in a mouse model of Alzheimer's disease using diffraction enhanced imaging

Our understanding of early development in Alzheimer's disease (AD) is clouded by the scale at which the disease progresses; amyloid beta (Aβ) plaques, a hallmark feature of AD, are small (~50 μm) and low contrast in diagnostic clinical imaging techniques. Diffraction enhanced imaging (DEI), a phase contrast x-ray imaging technique, has greater soft tissue contrast than conventional radiography and generates higher resolution images than magnetic resonance microimaging. Thus, in this proof of principle study, DEI in micro-CT mode was performed on the brains of AD-model mice to determine if DEI can visualize Aβ plaques. Results revealed small nodules in the cortex and hippocampus of the brain. Histology confirmed that the features seen in the DEI images of the brain were Aβ plaques. Several anatomical structures, including hippocampal subregions and white matter tracks, were also observed. Thus, DEI has strong promise in early diagnosis of AD, as well as general studies of the mouse brain.

In vitro and in vivo Staining Characteristics of Small, Fluorescent, Aβ42‐Binding D‐Enantiomeric Peptides in Transgenic AD Mouse Models

One of the characteristic pathological hallmarks of Alzheimer's disease (AD) are neuritic plaques that consist of amyloid peptide (Abeta). To improve diagnosis and treatment evaluation, neuroimaging tools that make use of Abeta-binding ligands to visualise amyloid plaques are being developed. We investigate the in vitro and in vivo characteristics of a series of three D-enantiomeric peptides (D1-D3) that were developed to specifically bind amyloid beta1-42 (Abeta42) in the brains of transgenic AD-model mice. We stained brain sections of the mice, injected and infused the mice with these small D-peptides, and examined their staining of Abeta42 in the brain. The experiments demonstrate that the D-peptides label all plaques that contain Abeta42 in the brain. In contrast, diffuse amyloid beta deposits (which do not contain Abeta42) are not stained by any of the D-peptides. The in vivo and in vitro studies demonstrate that the D-peptides label all Abeta42 in the brain, and none of the D-peptides causes inflammation or is taken up by astrocytes or microglia. Furthermore, long-term infusion of the peptides does not cause inflammation. Together, this demonstrates that these D-peptides might be suitable for use as molecular probes to measure Abeta plaque load in the living brain for early diagnosis of Alzheimer's disease, or to monitor Abeta42 plaque load during disease progression or during treatment.

Effects of Chinese herbal medicine fuzhisan on aged rats

Fuzhisan (FZS), a Chinese herbal complex prescription, has been used in the treatment of Alzheimer’s disease (AD) for more than 15 years. Previous studies showed that FZS enhanced the cognitive ability in AD patients and AD model rats. FZS modulated the impaired cellular functions, and attenuated the damage caused by β-amyloid protein, dose-dependently regulated and ameliorated the cholinergic functions of the Aβ 25–35-induced AD-model mice. The SPECT imaging revealed that FZS improved the blood flow of the frontal and temporal lobes and the callosal gyrus in AD patients. However, little investigation of the effects of FZS on the naturally aged rats was reported. The underlying mechanism also remains to be explored. Recently we investigated the effects of the aqueous extract of FZS on the cognitive functions of the aged rats and the pharmacological basis for its therapeutic efficacy. The results showed a significant improvement made by FZS (0.3, 0.6, and 1.2 g/kg/d) for impaired cognitive functions of the aged rats. The rats manifested a shortened latency in Morris water maze test after intra-gavage administration (ig) of FZS for 30 consecutive days. The micro-positron emission tomography (microPET) using 18F-2-fluoro-2-deoxy- d-glucose ( 18F-FDG) as the tracer demonstrated that FZS promoted the glucose metabolism in the whole brains especially the temporal and parietal regions in the aged rats. The spectrophotometry and Western blot showed that FZS obviously increased the activity and the production of choline O-acetyltransferase (ChAT, EC 2.3.1.6) and the acetylcholine (ACh) contents in the hippocampus, thus regulated and ameliorated the impaired cholinergic functions of the aged rats. The therapeutical effects of FZS on the learning and memory of the aged rats were dose-dependent. The mechanism of action of FZS in ameliorating the memory dysfunction of the aged rats is ascribed to the reinforcement of the function of the cholinergic system and the enhancement of the glucose metabolism in the brains. The results of this study, together with a survey of the findings in the clinical treatment with FZS suggest that FZS may not merely alleviate the symptoms of the dementia, but may also augment the production of the neurotransmitter ACh and the energy supply in the brain to build up fitness of the patients. FZS may be beneficial for the treatment of Alzheimer’s disease or cognitive impairment in old people.

P2-325: In VIVO labeling of amyloid β plaques in AD model mice using blood-brain barrier permeable styryl dyes

P2-325: In VIVO labeling of amyloid β plaques in AD model mice using blood-brain barrier permeable styryl dyes

P4-067: Development of in vitro Abeta sink assay, to identify the dissociation of soluble Abeta from fibrils

Abeta is one of the primary therapeutic targets for Alzheimer disease. Abeta vaccination induces the disappearance of Abeta deposits in AD model mice. Since few reports have focused on the reverse phase of Abeta aggregation, we established a new screening system, the in vitro Abeta sink assay, to clarify the difference in the process of dissociation of soluble forms from fibrils between Abeta40 and Abeta42. We analyzed soluble Abeta from fibrils by Abeta ELISA and size exclusion chromatography. We further employed ultrasonic irradiation and small molecules to enhance the dissociation of soluble Abeta from fibrils. Abeta42 was more resistant to dissociation from fibrils to monomers and/or low molecular weight (LMW) soluble oligomers than Abeta40. Ultrasound irradiation significantly enhanced the dissociation of soluble Abeta from fibrils, while ultrasound experiments also confirmed the difference between Abeta40 and Abeta42. We screened 7 known small molecules that are reported to inhibit Abeta aggregation. Some of them significantly enhanced dissociation of soluble Abeta42 from fibrils using this system. We propose that Abeta42 is more resistant to dissociation from fibrils to monomers and/or LMW soluble oligomers than Abeta40, and this system might be useful to identify dissociation of soluble Abeta from fibrils.

Neuropeptides concentrations in the skin of a murine (NC/Nga mice) model of atopic dermatitis

Background: It has been reported that the expression of neuropeptides (NPs), and the density and structure of peripheral nerves in atopic dermatitis (AD) are different from those in normal skin. Objective: We investigated the role of NPs, in the development of AD with quantitative study of substance P (SP) and calcitonin gene-related peptide (CGRP) in the skin of AD-model mice. Methods: We measured the NPs in the skin of mice (NC/Nga as AD-model mice, BALB/c and C57BL/6 as control) by enzyme-linked immunosorbentassay (ELISA). Peripheral nerve fibers and SP in the skin were stained by immunohistochemical staining, using anti-PGP9.5 antibody and anti-SP antibody. Results: Under conventional condition, SP concentration in AD-like skin lesions of NC/Nga mice was higher than that in non-affected skin of the same mice. Under specific pathogen-free condition, SP concentration in the skin of NC/Nga mice was higher than that in the skin of BALB/c and C57BL/6 mice. In contrast, CGRP concentration in the skin lesions was lower than that in non-affected skin of NC/Nga mice. SP was detected not only in the nerve fibers in the dermis but also in mast cells in the inflammatory areas. Conclusions: The skin of NC/Nga mice contains more SP congenitally, and environmental factors may aggravate this abnormal condition. We hypothesize that increase of SP accompanied with a decrease of CGRP in the skin may play important roles in the pathogenesis and development of AD.

Diffuse amyloid deposition, but not plaque number, is reduced in amyloid precursor protein/presenilin 1 double-transgenic mice by pathway lesions

Alzheimer's disease (AD) is the most common form of dementia in the elderly, and the characteristic pathological hallmarks of the disease are neuritic plaques and neurofibrillary tangles. The sequence of events leading to the extracellular deposition of amyloidβ (Aβ) peptides in plaques or in diffuse deposits is not clear. Here we investigate the relation between disrupted axonal transport of amyloid precursor protein (APP) and/or Aβ and the deposition of Aβ in the deafferented terminal fields in APP/presenilin 1 double-transgenic AD-model mice. In the first experiment we ablated entorhinal cortex neurons and examined the subsequent changes in amyloid deposition in the hippocampus 1 month later. We show that there is a substantial reduction in the amount of diffuse amyloid deposits in the denervated areas of the hippocampus. Further, to investigate the effects of long-term deafferentation, in a second experiment we cut the fimbria-fornix and analyzed the brains 11 months post-lesion. Diffuse amyloid deposits in the deafferented terminal fields of area CA1 and subiculum were dramatically reduced as assessed by image analysis of the Aβ load. Our findings indicate that neuronal ablations decrease diffuse amyloid deposits in the terminal fields of these neurons, and, further, that pathway lesions similarly decrease the amount of diffuse amyloid deposits in the terminal fields of the lesioned axons. Together, this suggests that the axonal transport of APP and/or Aβ and subsequent secretion of Aβ at terminals plays an important role in the deposition of Aβ protein in Alzheimer's disease, and, further, that diffuse deposits do not develop into plaques.py>