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Disease-modifying therapies, such as neuroprotective and neurorestorative interventions, are strongly desired for Alzheimer’s disease (AD) treatment. Several studies have suggested that histone deacetylase 2 (HDAC2) inhibition can exhibit disease-modifying effects in AD patients. However, whether HDAC2 inhibition shows neuroprotective and neurorestorative effects under neuropathic conditions, such as amyloid β (Aβ)-elevated states, remains poorly understood. Here, we performed HDAC2-specific knockdown in CA1 pyramidal cells and showed that HDAC2 knockdown increased the length of dendrites and the number of mushroom-like spines of CA1 basal dendrites in APP/PS1 transgenic mouse model. Furthermore, HDAC2 knockdown also ameliorated the deficits in hippocampal CA1 long-term potentiation and memory impairment in contextual fear conditioning tests. Taken together, our results support the notion that specific inhibition of HDAC2 has the potential to slow the disease progression of AD through ameliorating Aβ-induced neuronal impairments.

Impulsive choice behavior, which can be assessed using the delay discounting task, is a characteristic of various psychiatric disorders, including attention-deficit/hyperactivity disorder (ADHD). Guanfacine is a selective α2A-adrenergic receptor agonist that is clinically effective in treating ADHD. However, there is no clear evidence that systemic guanfacine administration reduces impulsive choice behavior in the delay discounting task in rats. In the present study, we examined the effect of systemic guanfacine administration on food-motivated impulsive choice behavior in rats and the neuronal mechanism underlying this effect. Repeated administration of either guanfacine, methylphenidate, or atomoxetine significantly enhanced impulse control, increasing the number of times the rats chose a large but delayed reward in a dose-dependent manner. The effect of guanfacine was significantly blocked by pretreatment with an α2A-adrenergic receptor antagonist. Furthermore, the effect of guanfacine remained unaffected in rats pretreated with a selective noradrenergic neurotoxin, consistent with a post-synaptic action. In contrast, the effect of atomoxetine on impulsive choice behavior was attenuated by pretreatment with the noradrenergic neurotoxin. These results provide the first evidence that systemically administered guanfacine reduces impulsive choice behavior in rats and that direct stimulation of postsynaptic, rather than presynaptic, α2A-adrenergic receptors is involved in this effect.

Summary γ-secretase inhibitors (GSI) are drugs developed to decrease amyloid-β peptide (Aβ) production by inhibiting intramembranous cleavage of β-amyloid protein precursor (βAPP). However, a large phase 3 trial of semagacestat, a potential non-transition state analog (non-TSA) GSI, in patients with Alzheimer's disease (AD) was terminated due to unexpected aggravation of cognitive deficits and side effects. Here, we show that some semagacestat effects are clearly different from a phenotype caused by a loss of function of presenilins, core proteins in the γ-secretase complex. Semagacestat increases intracellular byproduct peptides, produced along with Aβ through serial γ-cleavage of βAPP, as well as intracellular long Aβ species, in cell-based and in vivo studies of AD model mice. Other potential non-TSA GSIs, but not L685,458, a TSA GSI, have similar effects. Furthermore, semagacestat inhibits release of de novo intramembranous γ-byproducts to the soluble space. Thus, semagacestat is a pseudo-GSI, and therefore, the semagacestat clinical trial did not truly test the Aβ hypothesis.

We propose a novel semi-automatic approach to design biomarkers for capturing pharmacodynamic effects induced by pharmacological agents on the spectral power of electroencephalography (EEG) recordings. We apply this methodology to investigate the pharmacodynamic effects of methylphenidate (MPH) and atomoxetine (ATX) on attention deficit/hyperactivity disorder (ADHD), using rodent models. We inject the two agents into the spontaneously hypertensive rat (SHR) model of ADHD, the Wistar-Kyoto rat (WKY), and the Wistar rat (WIS), and record their EEG patterns. To assess individual EEG patterns quantitatively, we use an integrated methodological approach, which consists of calculating the mean, slope and intercept parameters of temporal records of EEG spectral power using a smoothing filter, outlier truncation, and linear regression. We apply Fisher discriminant analysis (FDA) to identify dominant discriminants to be heuristically consolidated into several new composite biomarkers. Results of the analysis of variance (ANOVA) and t-test show benefits in pharmacodynamic parameters, especially the slope parameter. Composite biomarker evaluation confirms their validity for genetic model stratification and the effects of the pharmacological agents used. The methodology proposed is of generic use as an approach to investigating thoroughly the dynamics of the EEG spectral power.

Impulsive choice behavior, which can be assessed using the delay discounting task, is a characteristic of various psychiatric disorders, including attention-deficit/hyperactivity disorder (ADHD). Guanfacine is a selective α

Background We analyze the dynamics of rodent EEG amplitude in an experiment accompanied by video recordings. Brain activity of animals is commonly acquired together with a video of behavior, but recordings are rarely combined in analysis. The data acquired is most commonly analyzed separately. To our knowledge, no study has used behavior to improve the analysis of EEG waveforms, specifically for artifact removal – other than through manual editing. Comparison with existing method(s) We explore two approaches: a traditional approach that relies on data preprocessing and artifact rejection by an expert; and an alternative approach that combines analysis of EEG with behavior extracted from video recordings. New method We use the level of activity extracted from the behavioral video as a measure of confidence in the acquired EEG waveform, and as a weighting factor in averaging and statistical comparisons. Results We find in analysis of the EEG that the two approaches lead to similar conclusions, but the analysis leveraging behavioral data achieves this while avoiding many subjective choices often required for artifact rejection and data preprocessing. Conclusions The methods we describe allow for the inclusion of all recorded data in the analysis, thereby making statistical tests more friendly to interpretation, and making the data processing transparent and reproducible.

Gram-positive bacteria of the genus Streptomyces are industrially important microorganisms, producing >70% of commercially important antibiotics. The production of these compounds is often regulated by low-molecular-weight bacterial hormones called autoregulators. Although 60% of Streptomyces strains may use γ-butyrolactone–type molecules as autoregulators and some use furan-type molecules, little is known about the signaling molecules used to regulate antibiotic production in many other members of this genus. Here, we purified a signaling molecule (avenolide) from Streptomyces avermitilis —the producer of the important anthelmintic agent avermectin with annual world sales of $850 million—and determined its structure, including stereochemistry, by spectroscopic analysis and chemical synthesis as (4 S ,10 R )-10-hydroxy-10-methyl-9-oxo-dodec-2-en-1,4-olide, a class of Streptomyces autoregulator. Avenolide is essential for eliciting avermectin production and is effective at nanomolar concentrations with a minimum effective concentration of 4 nM. The aco gene of S. avermitilis, which encodes an acyl-CoA oxidase, is required for avenolide biosynthesis, and homologs are also present in Streptomyces fradiae , Streptomyces ghanaensis , and Streptomyces griseoauranticus , suggesting that butenolide-type autoregulators may represent a widespread and another class of Streptomyces autoregulator involved in regulating antibiotic production.

Surrogate markers for the Alzheimer disease (AD)-associated 42-amino acid form of amyloid-beta (Abeta42) have been sought because they may aid in the diagnosis of AD and for clarification of disease pathogenesis. Here, we demonstrate that human cerebrospinal fluid (CSF) contains three APLP1-derived Abeta-like peptides (APL1beta) that are generated by beta- and gamma-cleavages at a concentration of approximately 4.5 nM. These novel peptides, APL1beta25, APL1beta27 and APL1beta28, were not deposited in AD brains. Interestingly, most gamma-secretase modulators (GSMs) and familial AD-associated presenilin1 mutants that up-regulate the relative production of Abeta42 cause a parallel increase in the production of APL1beta28 in cultured cells. Moreover, in CSF from patients with pathological mutations in presenilin1 gene, the relative APL1beta28 levels are higher than in non-AD controls, while the relative Abeta42 levels are unchanged or lower. Most strikingly, the relative APL1beta28 levels are higher in CSF from sporadic AD patients (regardless of whether they are at mild cognitive impairment or AD stage), than those of non-AD controls. Based on these results, we propose the relative level of APL1beta28 in the CSF as a candidate surrogate marker for the relative level of Abeta42 production in the brain.

Alzheimer-disease-associated beta-amyloid (Abeta) is produced by sequential endoproteolysis of beta-amyloid protein precursor (betaAPP): the extracellular portion is shed by cleavage in the juxtamembrane region by beta-amyloid-cleaving enzyme (BACE)/beta-secretase, after which it is cleaved by presenilin (PS)/gamma-secretase near the middle of the transmembrane domain. Thus, inhibition of either of the secretases reduces Abeta generation and is a fundamental strategy for the development of drugs to prevent Alzheimer disease. However, it is not clear how small compounds reduce Abeta production without inhibition of the secretases. Such compounds are expected to avoid some of the side effects of secretase inhibitors. Here, we report that destruxin E (Dx-E), a natural cyclic hexadepsipeptide, reduces Abeta generation without affecting BACE or PS/gamma-secretase activity. In agreement with this, Dx-E did not inhibit Notch signaling. We found that Dx-E decreases colocalization of BACE1 and betaAPP, which reduces beta-cleavage of betaAPP. Therefore, the data demonstrate that Dx-E represents a novel Abeta-reducing process which could have fewer side effects than secretase inhibitors.

Secretion by glandular trichomes is thought to be a strategy to retain moisture water on leaves under drought conditions. In the wild tomato species Lycopersicon pennellii exudates accumulate at the apical terminal of the trichome, forming a bulbous head. During a 30 s mist treatment, the exudates condensed moisture water to form water drops on the trichome heads, and water drops were absorbed through trichomes by the plants. However, a longer mist treatment (40 s) caused the drops on all trichome heads to drop down to the leaf surface. Water drops containing the exudates were amphiphilic and spread over the leaf surface in a thin layer. This is an additional mechanism of the glandular trichomes for covering leaves with the exudates as a chemical boundary layer. Exudates of type IV glandular trichomes of L. pennellii showed antifungal activity against conidia of the tomato powdery mildew pathogen Oidium neolycopersici. The exudates covering the leaves completely suppressed germination of inoculated conidia, which prevented subsequent infection events such as formation of appressoria and haustoria. On non-misted leaves almost all conidia germinated and formed appressoria, resulting in successful infection. L. pennellii is susceptible to tomato powdery mildew, but the mechanisms for condensing mist water by type IV trichome-derived exudates can contribute to suppressing foliar infection by the powdery mildew pathogen.

Background: The process of aggregation of brain amyloid-β peptides (Aβ) is thought to be associated with the pathogenesis of Alzheimer's disease (AD). Amyloid-β peptides are produced by sequential endoproteolysis by β-site amyloid-β protein precursor-cleaving enzyme (BACE) followed by presenilin (PS)/γ-secretase. There are several species of Aβ due to cleavage diversity of PS/γ-secretase. The predominant species in human cerebrospinal fluid (CSF) or plasma is Aβ40, whereas Aβ42 is much more aggregatable and accumulated in senile plaques. The level of Aβ in the brain is determined by the balance between the generation and clearance of Aβ, including transport across the brain–blood barrier (BBB). Although the processes of Aβ generation and degradation have been studied in some detail, knowledge of the Aβ transport process across the BBB is limited. So far, low-density lipoprotein receptor-related protein (LRP1), P-glycoprotein (P-gp), and insulin-like growth factor-1 (IGF-1) have been identified to modify the excretion of brain Aβ to the blood. Methods: To investigate whether macrophage colony stimulating factor (M-CSF) has a role in the Aβ transport process, human Aβ was injected into the lateral ventricle of the brain of M-CSF-deficient (op/op) mice. Then, plasma and brain Aβ levels were measured by ELISA to determine the time-course of Aβ movement from the brain to the plasma. Result: When human Aβ40 was injected into mouse lateral ventricles, the efflux of Aβ from the CSF to the blood was transiently decreased and delayed in M-CSF-deficient mice. Moreover, endogenous plasma Aβ40 levels were lower in M-CSF-deficient mice. Conclusion: The results indicate that M-CSF deficiency impairs excretion of human-type Aβ40 from the CSF to blood. We propose that M-CSF may be a novel factor that facilitates the excretion of Aβ from the CSF to the blood via the BBB.

Accumulation of amyloid beta (Abeta) is a pathological hallmark of Alzheimer's disease, and lowering Abeta is a promising therapeutic approach. Intact anti-Abeta antibodies reduce brain Abeta through two pathways: enhanced microglial phagocytosis and Abeta transfer from the brain to the periphery (Abeta sequestration). While activation of microglia, which is essential for microglial phagocytosis, is necessarily accompanied by undesired neuroinflammatory events, the capacity for sequestration does not seem to be linked to such effects. We and other groups have found that simple Abeta binding agents are sufficient to reduce brain Abeta through the sequestration pathway. In this study, we aimed to eliminate potentially deleterious immune activation from antibodies without affecting the ability to induce sequestration. The glycan portion of immunoglobulin is critically involved in interactions with immune effectors including the Fc receptor and complement c1q; deglycosylation eliminates these interactions, while antigen (Abeta)-binding affinity is maintained. In this study, we investigated whether deglycosylated anti-Abeta antibodies reduce microglial phagocytosis and neuroinflammation without altering the capacity to induce Abeta sequestration. Deglycosylated antibodies maintained Abeta binding affinity. Deglycosylated antibodies did not enhance Abeta phagocytosis or cytokine release in primary cultured microglia, whereas intact antibodies did so significantly. Intravenous injection of deglycosylated antibodies elevated plasma Abeta levels and induced Abeta sequestration to a similar or greater degree compared with intact antibodies in an Alzheimer's transgenic mouse model without or with Abeta plaque pathology. We conclude that deglycosylated antibodies effectively induced Abeta sequestration without provoking neuroinflammation; thus, these deglycosylated antibodies may be optimal for sequestration therapy for Alzheimer's disease.

Accumulation of amyloid beta peptide (Abeta) in brain is a hallmark of Alzheimer's disease (AD). Inhibition of beta-site amyloid precursor protein (APP)-cleaving enzyme-1 (BACE1), the enzyme that initiates Abeta production, and other Abeta-lowering strategies are commonly tested in transgenic mice overexpressing mutant APP. However, sporadic AD cases, which represent the majority of AD patients, are free from the mutation and do not necessarily have overproduction of APP. In addition, the commonly used Swedish mutant APP alters APP cleavage. Therefore, testing Abeta-lowering strategies in transgenic mice may not be optimal. In this study, we investigated the impact of BACE1 inhibition in non-transgenic mice with physiologically relevant APP expression. Existing Abeta ELISAs are either relatively insensitive to mouse Abeta or not specific to full-length Abeta. A newly developed ELISA detected a significant reduction of full-length soluble Abeta 1-40 in mice with the BACE1 homozygous gene deletion or BACE1 inhibitor treatment, while the level of x-40 Abeta was moderately reduced due to detection of non-full-length Abeta and compensatory activation of alpha-secretase. These results confirmed the feasibility of Abeta reduction through BACE1 inhibition under physiological conditions. Studies using our new ELISA in non-transgenic mice provide more accurate evaluation of Abeta-reducing strategies than was previously feasible.

Alzheimer’s disease (AD) is characterized by the accumulation of amyloid-β (Aβ). The genes that govern this process, however, have remained elusive. To this end, we combined distinct mouse strains with transcriptomics to directly identify disease-relevant genes. We show that AD model mice (APP-Tg) with DBA/2 genetic backgrounds have significantly lower levels of Aβ accumulation compared with SJL and C57BL/6 mice. We then applied brain transcriptomics to reveal the genes in DBA/2 that suppress Aβ accumulation. To avoid detecting secondarily affected genes by Aβ, we used non-Tg mice in the absence of Aβ pathology and selected candidate genes differently expressed in DBA/2 mice. Additional transcriptome analysis of APP-Tg mice with mixed genetic backgrounds revealed kinesin light chain-1 (Klc1) as an Aβ modifier, indicating a role for intracellular trafficking in Aβ accumulation. Aβ levels correlated with the expression levels of Klc1 splice variant E and the genotype of Klc1 in these APP-Tg mice. In humans, the expression levels of KLC1 variant E in brain and lymphocyte were significantly higher in AD patients compared with unaffected individuals. Finally, functional analysis using neuroblastoma cells showed that overexpression or knockdown of KLC1 variant E increases or decreases the production of Aβ, respectively. The identification of KLC1 variant E suggests that the dysfunction of intracellular trafficking is a causative factor of Aβ pathology. This unique combination of distinct mouse strains and model mice with transcriptomics is expected to be useful for the study of genetic mechanisms of other complex diseases.

Effects of cement composition and silica content on the microstructure and compressive strength of autoclaved mortar with silica powder were studied. The strength of autoclaved mortars prepared by mixing with portland cement and silica powder with low water cement ratio (W/S=0.25) was affected by the amount of silica powder. The maximum strength was obtained for the mortar with silica content where calcium hydroxide was completely consumed by the reaction with silica powder added. The correlation between formation of tobermorite and compressive strength of autocraved mortar was not clear. The excessive addition of silica powder resulted in the formation of gap between the silica particles and matrix, with decreasing in the strength of mortars. A correlation between pore volume and compressive strength was obtained when these gap did not exist.

The appressorial shapes of the powdery mildews are an important clue to the taxonomy of the powdery mildew fungi, but the conidia of the tomato powdery mildew Oidium neolycopersici KTP-01 develop non-lobed, nipple-shaped, and moderately lobed or multilobed appressoria on the same leaves. To remove this ambiguity, we performed consecutive observations of sequential appressorial development of KTP-01 conidia with a high-fidelity digital microscope. Highly germinative conidia of KTP-01, collected from conidial pseudochains formed on the tomato leaves, were inoculated into host tomato and nonhost barley leaves or an artificial hydrophobic membrane (Parafilm). Events from germination initiation to appressorium formation were synchronous in all conidia on all materials used for inoculation, but post-appressorial behaviors varied among the materials. Appressoria on the membrane-stuck glass slide formed several projections at different portions of the appressoria to repeat unsuccessful penetration attempts. Similar unsuccessful penetration behavior by KTP-01 conidia was observed in the inoculations into leaves of barley plants, wild tomato species Lycopersicon peruvianum LA2172 (carrying the Ol-4 gene for powdery mildew resistance), and a susceptible host tomato (Lycopersicon esculentum) that had been inoculated with the barley powdery mildew (Blumeria graminis f. sp. hordei, race 1) conidia. On the barley leaves, all penetrations of KTP-01 were impeded by the papillae formed beneath the sites of the appressorial projections. On both the wild tomato and the race 1-inoculated cultivated tomato plants, KTP-01 conidia were prevented from forming functional haustoria by hypersensitive epidermal cell death; this hypersensitive reaction involved the Ol-4 gene in the wild tomato plants or the ‘induced resistance’ acquired by the nonpathogenic conidia previously inoculated into the cultivated tomato plants. All these KTP-01 conidia produced several projections on the appressoria during the repeated unsuccessful penetration attempts and eventually exhibited multilobed appressoria. On the host tomato leaves inoculated singly with KTP-01 conidia, fewer than 20 % of the conidia located appressoria on the central part of target epidermal cells and succeeded in forming functional haustoria at the first penetration attempt without forming an appressorial projection. These conidia exhibited non-lobed appressoria. The remaining conidia, however, whose appressoria were located on/near the border of the target epidermal cells, were more likely to fail to penetrate at the first penetration, and then to develop additional projections for subsequent penetrations. Most conidia succeeded in forming functional haustoria at the second to fourth penetration attempts, but a few conidia failed to produce haustoria at all attempted penetrations. Eventually, the conidia that succeeded at the second penetration possessed a single appressorial projection (exhibiting the nipple-shaped appressoria), whereas the remaining conidia exhibited moderately lobed appressoria with two to four appressorial projections and multilobed appressoria, with more projections. Thus, the present study revealed that the basic shape of appressoria of KTP-01 was the non-lobed type, and that polymorphic changes of the appressoria occurred as a result of successive production of projections during repeated unsuccessful penetration attempts.