Dimerization of the transmembrane domain of amyloid precursor proteins and familial Alzheimer's disease mutants

Background  

Amyloid precursor protein (APP) is enzymatically cleaved by γ-secretase to form two peptide products, either Aβ40 or the more neurotoxic Aβ42. The Aβ42/40 ratio is increased in many cases of familial Alzheimer's disease (FAD). The transmembrane domain (TM) of APP contains the known dimerization motif GXXXA. We have investigated the dimerization of both wild type and FAD mutant APP transmembrane domains.     

Chronic administration of R-flurbiprofen attenuates learning impairments in transgenic amyloid precursor protein mice

Background  

Long-term use of non-steroidal anti-inflammatory drugs (NSAIDs) is associated with a reduced incidence of Alzheimer's disease (AD). We and others have shown that certain NSAIDs reduce secretion of Aβ42 in cell culture and animal models, and that the effect of NSAIDs on Aβ42 is independent of the inhibition of cyclooxygenase by these compounds. Since Aβ42 is hypothesized to be the initiating pathologic molecule in AD, the ability of these compounds to lower Aβ42 selectively may be associated with their protective effect. We have previously identified R-flurbiprofen (tarenflurbil) as a selective Aβ42 lowering agent with greatly reduced cyclooxygenase activity that shows promise for testing this hypothesis. In this study we report the effect of chronic R-flurbiprofen treatment on cognition and Aβ loads in Tg2576 APP mice.     

Improvement of a low pH antigen-antibody dissociation procedure for ELISA measurement of circulating anti-Aβ antibodies

Background  

Prior work from our group found that acid dissociation (pH 2.5 incubation) of serum from APP transgenic mice vaccinated against Aβ increased the apparent anti-Aβ titers, suggesting antibody masking by antigen in the ELISA assay. Subsequently, we found that pH 2.5 incubation of serum from unvaccinated non-transgenic mice showed antibody binding to Aβ1–42, but no increase when other proteins, including shorter Aβ peptides, coated the ELISA plate. To investigate further the effects of low pH incubation on apparent anti-Aβ1–42 signals, we examined normal sera from nonTg unvaccinated mice, nonTg mice vaccinated with Aβ peptide (to produce authentic anti-Aβ antibodies) or a monoclonal antibody against Aβ (6E10) using competitive-inhibition ELISA and Aβ epitope mapping assays. In addition, we examined use of a less stringent low pH procedure at pH 3.5, to ascertain if it had the same effects as the pH 2.5 procedure.     

Overcoming antigen masking of anti-amyloidbeta antibodies reveals breaking of B cell tolerance by virus-like particles in amyloidbeta immunized amyloid precursor protein transgenic mice

Background  

In prior work we detected reduced anti-Aβ antibody titers in Aβ-vaccinated transgenic mice expressing the human amyloid precursor protein (APP) compared to nontransgenic littermates. We investigated this observation further by vaccinating APP and nontransgenic mice with either the wild-type human Aβ peptide, an Aβ peptide containing the "Dutch Mutation", E22Q, or a wild-type Aβ peptide conjugated to papillomavirus virus-like particles (VLPs).     

Anti-PrP<Superscript>C</Superscript> monoclonal antibody infusion as a novel treatment for cognitive deficits in an alzheimer's disease model mouse

Background  

Alzheimer's Disease (AD) is the most common of the conformational neurodegenerative disorders characterized by the conversion of a normal biological protein into a β-sheet-rich pathological isoform. In AD the normal soluble Aβ (sAβ) forms oligomers and fibrils which assemble into neuritic plaques. The most toxic form of Aβ is thought to be oligomeric. A recent study reveals the cellular prion protein, PrP^C, to be a receptor for Aβ oligomers. Aβ oligomers suppress LTP signal in murine hippocampal slices but activity remains when pretreated with the PrP monoclonal anti-PrP antibody, 6D11. We hypothesized that targeting of PrP^C to prevent Aβ oligomer-related cognitive deficits is a potentially novel therapeutic approach. APP/PS1 transgenic mice aged 8 months were intraperitoneally (i.p.) injected with 1 mg 6D11 for 5 days/week for 2 weeks. Two wild-type control groups were given either the same 6D11 injections or vehicle solution. Additional groups of APP/PS1 transgenic mice were given either i.p. injections of vehicle solution or the same dose of mouse IgG over the same period. The mice were then subjected to cognitive behavioral testing using a radial arm maze, over a period of 10 days. At the conclusion of behavioral testing, animals were sacrificed and brain tissue was analyzed biochemically or immunohistochemically for the levels of amyloid plaques, PrP^C, synaptophysin, Aβ40/42 and Aβ oligomers.     

Long-term neprilysin gene transfer is associated with reduced levels of intracellular Abeta and behavioral improvement in APP transgenic mice

Background  

Proteolytic degradation has emerged as a key pathway involved in controlling levels of the Alzheimer's disease (AD)-associated amyloid-β (Aβ) peptide in the brain. The endopeptidase, neprilysin, has been implicated as a major Aβ degrading enzyme in mice and humans. Previous short and intermediate term studies have shown the potential therapeutic application of neprilysin by delivering this enzyme into the brain of APP transgenic mice using gene transfer with viral vectors. However the effects of long-term neprilysin gene transfer on other aspects of Aβ associated pathology have not been explored yet in APP transgenic mice.     

Soy isoflavone glycitein protects against beta amyloid-induced toxicity and oxidative stress in transgenic <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis>

Background  

Epidemiological studies have associated estrogen replacement therapy with a lower risk of developing Alzheimer's disease, but a higher risk of developing breast cancer and certain cardiovascular disorders. The neuroprotective effect of estrogen prompted us to determine potential therapeutic impact of soy-derived estrogenic compounds. Transgenic C. elegans, that express human beta amyloid (Aβ), were fed with soy derived isoflavones genistein, daidzein and glycitein (100 μg/ml) and then examined for Aβ-induced paralysis and the levels of reactive oxygen species.     

Acceleration and inhibition of amyloid-β fibril formation by peptide-conjugated fluorescent-maghemite nanoparticles

The formation of amyloid aggregates by association of peptides into ordered structures is hallmark of certain neurodegenerative disorders. Exploring the effect of specific nanoparticles on the formation of amyloid fibrils may contribute toward a mechanistic understanding of the aggregation processes, leading to design nanoparticles that modulate the formation of toxic amyloid plaques. Uniform maghemite (γ-Fe₂O₃) magnetic nanoparticles, containing fluorescein covalently encapsulated within (F-γ-Fe₂O₃)_, were prepared. These F-γ-Fe₂O₃ nanoparticles of 14.0 ± 4.0 nm were then coated with human serum albumin (HSA) via a precipitation process. Covalent conjugation of the spacer arm succinimidyl polyethylene glycol succinimidyl ester (NHS–PEG–NHS) to the F-γ-Fe₂O₃~HSA nanoparticles was then accomplished by interacting the primary amine groups of the HSA coating with excess NHS–PEG–NHS molecules. Covalent conjugation of the peptides amyloid-β 40 (Aβ₄₀) or Leu-Pro-Phe-Phe-Asp (LPFFD) onto the surface of the former fluorescent nanoparticles was then performed, by interacting the terminal activated NHS groups of the PEG derivatized F-γ-Fe₂O₃~HSA nanoparticles with primary amino groups of the peptides. Kinetics of the Aβ₄₀ fibrillation process in the absence and presence of varying concentrations of the Aβ₄₀ or LPFFD conjugated nanoparticles were also elucidated. The non-peptide conjugated fluorescent nanoparticles do not affect the Aβ₄₀ fibrillation process significantly. However, the Aβ₄₀-conjugated nanoparticles (F-γ-Fe₂O₃~HSA–PEG–Aβ₄₀) accelerate the fibrillation process while the LPFFD-conjugated nanoparticles (F-γ-Fe₂O₃~HSA–PEG–LPFFD) inhibit it. By applying MRI and fluorescence imaging techniques simultaneously these bioactive fluorescent magnetic iron oxide nanoparticles can be used as an efficient tool to study and control the Aβ₄₀ amyloid fibril formation process.     

Excitability of Aβ sensory neurons is altered in an animal model of peripheral neuropathy

Background  

Causes of neuropathic pain following nerve injury remain unclear, limiting the development of mechanism-based therapeutic approaches. Animal models have provided some directions, but little is known about the specific sensory neurons that undergo changes in such a way as to induce and maintain activation of sensory pain pathways. Our previous studies implicated changes in the Aβ, normally non-nociceptive neurons in activating spinal nociceptive neurons in a cuff-induced animal model of neuropathic pain and the present study was directed specifically at determining any change in excitability of these neurons. Thus, the present study aimed at recording intracellularly from Aβ-fiber dorsal root ganglion (DRG) neurons and determining excitability of the peripheral receptive field, of the cell body and of the dorsal roots.     

Adrenalectomy promotes a permanent decrease of plasma corticoid levels and a transient increase of apoptosis and the expression of Transforming Growth Factor β1 (TGF-β1) in hippocampus: effect of a TGF-β1 oligo-antisense

Background  

Corticosterone reduction produced by adrenalectomy (ADX) induces apoptosis in dentate gyrus (DG) of the hippocampus, an effect related to an increase in the expression of the pro-apoptotic gene bax. However it has been reported that there is also an increase of the anti-apoptotic gene bcl-2, suggesting the promotion of a neuroprotective phenomenon, perhaps related to the expression of transforming growth factor β1 (TGF-β1). Thus, we have investigated whether TGF-β1 levels are induced by ADX, and whether apoptosis is increased by blocking the expression of TGF-β1 with an antisense oligonucleotide (ASO) administered intracerebrally in corticosterone depleted rats.     

Diffusion and aggregation of Alzheimer’s Aβ1–40 peptide in aqueous trifluoroethanol solutions as studied by pulsed field gradient NMR

Pulsed field gradient nuclear magnetic resonance technique was applied to measure the self-diffusion coefficient of Aβ_1–40 peptide in trifluoroethanol (TFE) and mixed solvent TFE-water (D₂O) buffer (pD 7.8) at 293 K. The data were analyzed on the basis of the Stokes model and the hardsphere approach was used to estimate self-diffusion coefficients. It was found that the extent of the Aβ_1–40 aggregation in TFE solutions depends on the concentration of the peptide and the sample preparation protocol. After soft mixing, i.e., without any additional mechanical pretreatment of the peptide, the peptide is present in the monomeric form in TFE solutions. However, the additional water-bath sonication of the sample during the dissolution of Aβ_1–40 in TFE enforces oligomerization of the peptide with the size of aggregates ranging from tetra- to hexamers. An increase of D₂O in the mixed TFE-D₂O solvent of up to 75% leads to the aggregation of the larger part of the peptide. However, the components of self-diffusion coefficients related to low-mass Aβ_1–40 oligomers (dimers and trimers) were not observed in the diffusion decay curves. The most probable explanation is that dimers and trimers are not the principal intermediate species in the aggregation of Aβ_1–40 peptide.     

Central administration of dipeptides,beta-alanyl-BCAAs,induces hyperactivity in chicks

Background  

Carnosine (β-alanyl-L-histidine) is a putative neurotransmitter and has a possible role in neuron-glia cell interactions. Previously, we reported that carnosine induced hyperactivity in chicks when intracerebroventricularly (i.c.v.) administered. In the present study, we focused on other β-alanyl dipeptides to determine if they have novel functions.     

One-pot synthesis of benzochromeno-pyrazole derivatives

Abstract  

One-pot synthesis of different benzochromeno-pyrazole derivatives has been reported via reaction of aldehydes, 3-methyl-1H-pyrazol-5(4H)-one and α-naphthol/β-naphthol in the presence of sulfamic acid.     

TGF-β1 induction of the adenine nucleotide translocator 1 in astrocytes occurs through Smads and Sp1 transcription factors

Background  

The adenine nucleotide translocator 1 (Ant1) is an inner mitochondrial membrane protein involved with energy mobilization during oxidative phosphorylation. We recently showed that rodent Ant1 is upregulated by transforming growth factor-beta (TGF-β) in reactive astrocytes following CNS injury. In the present study, we describe the molecular mechanisms by which TGF-β1 regulates Ant1 gene expression in cultured primary rodent astrocytes.     

New insight into the role of the β3 subunit of the GABA<Subscript>A</Subscript>-R in development,behavior, body weight regulation,and anesthesia revealed by conditional gene knockout

Background  

The β3 subunit of the γ-aminobutyric acid type A receptor (GABA_A-R) has been reported to be important for palate formation, anesthetic action, and normal nervous system function. This subunit has also been implicated in the pathogenesis of Angelman syndrome and autism spectrum disorder. To further investigate involvement of this subunit, we previously produced mice with a global knockout of β3. However, developmental abnormalities, compensation, reduced viability, and numerous behavioral abnormalities limited the usefulness of that murine model. To overcome many of these limitations, a mouse line with a conditionally inactivated β3 gene was engineered.     

Protection of cortical cells by equine estrogens against glutamate-induced excitotoxicity is mediated through a calcium independent mechanism

Background  

High concentrations of glutamate can accumulate in the brain and may be involved in the pathogenesis of neurodegenerative disorders such as Alzheimer's disease. This form of neurotoxicity involves changes in the regulation of cellular calcium (Ca²⁺) and generation of free radicals such as peroxynitrite (ONOO^-). Estrogen may protect against glutamate-induced cell death by reducing the excitotoxic Ca²⁺ influx associated with glutamate excitotoxicity. In this study, the inhibition of N-methyl-D-aspartate (NMDA) receptor and nitric oxide synthase (NOS) along with the effect of 17β-estradiol (17β-E₂) and a more potent antioxidant Δ⁸, 17β-estradiol (Δ⁸, 17β-E₂) on cell viability and intracellular Ca²⁺ ([Ca²⁺]_i), following treatment of rat cortical cells with glutamate, was investigated.     

Intramolecular Charge Transfer Effects on Flutamide Drug

Spectral characteristics of flutamide drug have been studied in various solvents and β-cyclodextrin (β-CD). The inclusion complex of flutamide with β-CD is analysed by UV-visible, fluorimetry, FT-IR, 1H NMR, SEM, DSC and AM1 methods. In all solvents, flutamide exhibits a dual fluorescence. The longer wavelength emission (A band ∼380 nm) is due to intramolecular charge transfer state (ICT) and the shorter wavelength emission (B band ∼285 nm) originates from a locally excited state. In β-CD, the increase in the fluorescence intensity of ‘A’ band indicates ICT emission enhanced in the β-CD medium. β-CD studies shows isopropyl group is present in the interior part of the β-CD cavity whereas amino and CF₃ groups are present in the outside of the β-CD cavity. A mechanism is proposed to explain the 1:1 inclusion process.     

Equine estrogens differentially inhibit DNA fragmentation induced by glutamate in neuronal cells by modulation of regulatory proteins involved in programmed cell death

Background  

Recent data indicate that excitotoxicity of high levels of neurotransmitter glutamate may be mediated via programmed cell death (apoptosis) and that it can be prevented in HT22 mouse hippocampal cells by various equine estrogens with Δ⁸,17β-estradiol (Δ⁸,17β-E₂) being the most potent. In order to delineate the mechanism(s), glutamate-induced cell death of HT22 cells was assessed by measuring (a) DNA fragmentation in the presence or absence of 11 equine estrogens (components of the drug CEE); (b) cell death and (c) levels of anti-apoptotic (Bcl-2) and proapoptotic (Bax) proteins in the presence or absence of two equine estrogens, Δ⁸,17β-E₂ and 17β-estradiol (17β-E₂) by LDH release assay and Western blot analysis respectively.     

Different methods for administering 17β-estradiol to ovariectomized rats result in opposite effects on ischemic brain damage

Background  

Numerous stroke studies have controversially shown estrogens to be either neuroprotective or neurodamaging. The discordant results observed in rat brain ischemia models may be a consequence of discrepancies in estrogen administration modes resulting in plasma concentration profiles far from those intended. To test this hypothesis we reproduced in detail and extended an earlier study from our lab using a different mode of 17β-estradiol administration; home-made silastic capsules instead of commercial slow-release 17β-estradiol pellets. Four groups of female rats (n = 12) were ovariectomized and administered 17β-estradiol or placebo via silastic capsules. All animals underwent MCAo fourteen days after ovariectomy and were sacrificed three days later.