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Background: Pyroglutamate-3 Aβ (pGlu-3 Aβ) is an N-terminally truncated and post-translationally modified Aβ species found in Alzheimer's disease (AD) brain. Its increased peptide aggregation propensity and toxicity make it an attractive emerging treatment strategy for AD. We address the question of how the effector function of an anti-pGlu-3 Aβ antibody influences the efficacy of immunotherapy in mouse models with AD-like pathology., Methods: We compared two different immunoglobulin (Ig) isotypes of the same murine anti-pGlu-3 Aβ mAb (07/1 IgG1 and 07/2a IgG2a) and a general N-terminal Aβ mAb (3A1 IgG1) for their ability to clear Aβ and protect cognition in a therapeutic passive immunotherapy study in aged, plaque-rich APP SWE /PS1ΔE9 transgenic (Tg) mice. We also compared the ability of these antibodies and a CDC-mutant form of 07/2a (07/2a-k), engineered to avoid complement activation, to clear Aβ in an ex vivo phagocytosis assay and following treatment in APP SL xhQC double Tg mice, and to activate microglia using longitudinal microPET imaging with TSPO-specific 18 F-GE180 tracer following a single bolus antibody injection in young and old Tg mice., Results: We demonstrated significant cognitive improvement, better plaque clearance, and more plaque-associated microglia in the absence of microhemorrhage in aged APP SWE /PS1ΔE9 Tg mice treated with 07/2a, but not 07/1 or 3A1, compared to PBS in our first in vivo study. All mAbs cleared plaques in an ex vivo assay, although 07/2a promoted the highest phagocytic activity. Compared with 07/2a, 07/2a-k showed slightly reduced affinity to Fcγ receptors CD32 and CD64, although the two antibodies had similar binding affinities to pGlu-3 Aβ. Treatment of APP SL xhQC mice with 07/2a and 07/2a-k mAbs in our second in vivo study showed significant plaque-lowering with both mAbs. Longitudinal 18 F-GE180 microPET imaging revealed different temporal patterns of microglial activation for 3A1, 07/1, and 07/2a mAbs and no difference between 07/2a-k and PBS-treated Tg mice., Conclusion: Our results suggest that attenuation of behavioral deficits and clearance of amyloid is associated with strong effector function of the anti-pGlu-3 Aβ mAb in a therapeutic treatment paradigm. We present evidence that antibody engineering to reduce CDC-mediated complement binding facilitates phagocytosis of plaques without inducing neuroinflammation in vivo. Hence, the results provide implications for tailoring effector function of humanized antibodies for clinical development.

Space travel will expose people to high-energy, heavy particle radiation, and the cognitive deficits induced by this exposure are not well understood. To investigate the short-term effects of space radiation, we irradiated 4-month-old Alzheimer's disease (AD)-like transgenic (Tg) mice and wildtype (WT) littermates with a single, whole-body dose of 10 or 50 cGy 56 Fe ions (1 GeV/u) at Brookhaven National Laboratory. At ~1.5 months post irradiation, behavioural testing showed sex-, genotype-, and dose-dependent changes in locomotor activity, contextual fear conditioning, grip strength, and motor learning, mainly in Tg but not WT mice. There was little change in general health, depression, or anxiety. Two months post irradiation, microPET imaging of the stable binding of a translocator protein ligand suggested no radiation-specific change in neuroinflammation, although initial uptake was reduced in female mice independently of cerebral blood flow. Biochemical and immunohistochemical analyses revealed that radiation reduced cerebral amyloid-β levels and microglia activation in female Tg mice, modestly increased microhemorrhages in 50 cGy irradiated male WT mice, and did not affect synaptic marker levels compared to sham controls. Taken together, we show specific short-term changes in neuropathology and behaviour induced by 56 Fe irradiation, possibly having implications for long-term space travel.

Both aging and Alzheimer's disease (AD) are associated with widespread epigenetic changes, with most evidence suggesting global hypomethylation in AD. It is, however, unclear how these age-related epigenetic changes are linked to molecular aberrations as expressed in animal models of AD. Here, we investigated age-related changes of epigenetic markers of DNA methylation and hydroxymethylation in a range of animal models of AD, and their correlations with amyloid plaque load. Three transgenic mouse models, including the J20, APP/PS1dE9 and 3xTg-AD models, as well as Caribbean vervets (a non-transgenic non-human primate model of AD) were investigated. In the J20 mouse model, an age-related decrease in DNA methylation was found in the dentate gyrus (DG) and a decrease in the ratio between DNA methylation and hydroxymethylation was found in the DG and cornu ammonis (CA) 3. In the 3xTg-AD mice, an age-related increase in DNA methylation was found in the DG and CA1-2. No significant age-related alterations were found in the APP/PS1dE9 mice and non-human primate model. In the J20 model, hippocampal plaque load showed a significant negative correlation with DNA methylation in the DG, and with the ratio a negative correlation in the DG and CA3. For the APP/PS1dE9 model a negative correlation between the ratio and plaque load was observed in the CA3, as well as a negative correlation between DNA methyltransferase 3A (DNMT3A) levels and plaque load in the DG and CA3. Thus, only the J20 model showed an age-related reduction in global DNA methylation, while DNA hypermethylation was observed in the 3xTg-AD model. Given these differences between animal models, future studies are needed to further elucidate the contribution of different AD-related genetic variation to age-related epigenetic changes., (Copyright © 2017. Published by Elsevier Inc.)

The complement cascade not only is an innate immune response that enables removal of pathogens but also plays an important role in microglia-mediated synaptic refinement during brain development. Complement C3 is elevated in Alzheimer's disease (AD), colocalizing with neuritic plaques, and appears to contribute to clearance of Aβ by microglia in the brain. Previously, we reported that C3-deficient C57BL/6 mice were protected against age-related and region-specific loss of hippocampal synapses and cognitive decline during normal aging. Furthermore, blocking complement and downstream iC3b/CR3 signaling rescued synapses from Aβ-induced loss in young AD mice before amyloid plaques had accumulated. We assessed the effects of C3 deficiency in aged, plaque-rich APPswe/PS1dE9 transgenic mice (APP/PS1; C3 KO). We examined the effects of C3 deficiency on cognition, Aβ plaque deposition, and plaque-related neuropathology at later AD stages in these mice. We found that 16-month-old APP/PS1; C3 KO mice performed better on a learning and memory task than did APP/PS1 mice, despite having more cerebral Aβ plaques. Aged APP/PS1; C3 KO mice also had fewer microglia and astrocytes localized within the center of hippocampal Aβ plaques compared to APP/PS1 mice. Several proinflammatory cytokines in the brain were reduced in APP/PS1; C3 KO mice, consistent with an altered microglial phenotype. C3 deficiency also protected APP/PS1 mice against age-dependent loss of synapses and neurons. Our study suggests that complement C3 or downstream complement activation fragments may play an important role in Aβ plaque pathology, glial responses to plaques, and neuronal dysfunction in the brains of APP/PS1 mice., (Copyright © 2017, American Association for the Advancement of Science.)

To investigate the effects of starvation and re-feeding on survival rate, behavior, growth and digestive enzyme activities of juvenile Sepia pharaonis, the manipulative laboratory experiments were conducted with 0-6 days of starvation and 15 days of re-feeding to reveal the mechanism of compensatory growth. The results showed that starvation exerted a significant negative effect on the survival rate, growth, hepatosomatic index and digestive enzyme activities (P<0.05). The survival rate and digestive enzyme activities declined during the whole starvation process. After 3 days, the survival rate began to decline significantly, the decrease rate of body mass increased obviously and juveniles experienced some abnormal behaviors, such as inkjet, fighting and so on. The effects of starvation on digestive enzyme activities of juveniles were significant, and the digestive enzyme activities declined and then rose during the whole starvation process. The lowest amylase activity occurred after 4 days of starvation (0.07±0.02 U·mg -1 ·prot -1 ). The lowest lipase activity occurred after 2 days of starvation (18.47±2.07 U·g -1 ·prot -1 ), and the highest after 6 days (57.60±3.98 U·g -1 ·prot -1 ). The lowest pepsin and trypsin activities occurred after 5 days (1.98±0.59 U·mg -1 ·prot -1 ) and 4 days (186.68±20.72 U·mg -1 ·prot -1 ) of starvation, respectively. The effects of re-feeding on survival rate, growth, hepatosomatic index and digestive enzyme activities of juveniles were significant. The survival rate, specific growth rate, hepatosomatic index and feeding rate were negatively correlated with hunger processing duration. The survival rate, specific growth rate and hepatosomatic index showed no significant difference between 1 day starvation group and 2 days starvation group (P>0.05). The survival rate, specific growth rate and hepatosomatic index of 3 days to 6 days starvation groups were significantly lower than the control group. The feeding rates of 1 day and 2 days starvation groups were obviously higher than that of the control group. The feeding rate of 6 days starvation group was significantly lower than that of the control group. The amylase and lipase activities were not significantly different among different starvation treatments, whereas the pepsin and trypsin activities were significantly different, with the highest value in the control group (pepsin 7.06±0.64 U·mg -1 ·prot -1 , trypsin 914.67±26.54 U·mg -1 ·prot -1 ) and the lo-west value in the group with 6 days of starvation (pepsin 3.21±0.57 U·mg -1 ·prot -1 , trypsin 660.04±37.92 U·mg -1 ·prot -1 ). Six days of starvation would be the point of no-return for the juveniles, without any compensatory effect after starvation and re-feeding.

Pyroglutamate-3 amyloid-beta (pGlu-3 Aβ) is an N-terminally truncated Aβ isoform likely playing a decisive role in Alzheimer's disease pathogenesis. Here, we describe a prophylactic passive immunization study in APPswe/PS1ΔE9 mice using a novel pGlu-3 Aβ immunoglobulin G1 (IgG1) monoclonal antibody, 07/1 (150 and 500 μg, intraperitoneal, weekly) and compare its efficacy with a general Aβ IgG1 monoclonal antibody, 3A1 (200 μg, intraperitoneal, weekly) as a positive control. After 28 weeks of treatment, plaque burden was reduced and cognitive performance of 07/1-immunized Tg mice, especially at the higher dose, was normalized to wild-type levels in 2 hippocampal-dependent tests and partially spared compared with phosphate-buffered saline-treated Tg mice. Mice that received 3A1 had reduced plaque burden but showed no cognitive benefit. In contrast with 3A1, treatment with 07/1 did not increase the concentration of Aβ in plasma, suggesting different modes of Aβ plaque clearance. In conclusion, early selective targeting of pGlu-3 Aβ by immunotherapy may be effective in lowering cerebral Aβ plaque burden and preventing cognitive decline in the clinical setting. Targeting this pathologically modified form of Aβ thereby is unlikely to interfere with potential physiologic function(s) of Aβ that have been proposed., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Alzheimer's disease (AD) is the most common cause of dementia. Neuroinflammation appears to play an important role in AD pathogenesis. Ligands of the 18 kDa translocator protein (TSPO), a marker for activated microglia, have been used as positron emission tomography (PET) tracers to reflect neuroinflammation in humans and mouse models. Here, we used the novel TSPO-targeted PET tracer (18)F-GE180 (flutriciclamide) to investigate differences in neuroinflammation between young and old WT and APP/PS1dE9 transgenic (Tg) mice. In vivo PET scans revealed an overt age-dependent elevation in whole-brain uptake of (18)F-GE180 in both WT and Tg mice, and a significant increase in whole-brain uptake of (18)F-GE180 (peak-uptake and retention) in old Tg mice compared with young Tg mice and all WT mice. Similarly, the (18)F-GE180 binding potential in hippocampus was highest to lowest in old Tg > old WT > young Tg > young WT mice using MRI coregistration. Ex vivo PET and autoradiography analysis further confirmed our in vivo PET results: enhanced uptake and specific binding (SUV75%) of (18)F-GE180 in hippocampus and cortex was highest in old Tg mice followed by old WT, young Tg, and finally young WT mice. (18)F-GE180 specificity was confirmed by an in vivo cold tracer competition study. We also examined (18)F-GE180 metabolites in 4-month-old WT mice and found that, although total radioactivity declined over 2 h, of the remaining radioactivity, ∼90% was due to parent (18)F-GE180. In conclusion, (18)F-GE180 PET scans may be useful for longitudinal monitoring of neuroinflammation during AD progression and treatment., Significance Statement: Microglial activation, a player in Alzheimer's disease (AD) pathogenesis, is thought to reflect neuroinflammation. Using in vivo microPET imaging with a novel TSPO radioligand, (18)F-GE180, we detected significantly enhanced neuroinflammation during normal aging in WT mice and in response to AD-associated pathology in APP/PS1dE9 Tg mice, an AD mouse model. Increased uptake and specific binding of (18)F-GE180 in whole brain and hippocampus were confirmed by ex vivo PET and autoradiography. The binding specificity and stability of (18)F-GE180 was further confirmed by a cold tracer competition study and a metabolite study, respectively. Therefore, (18)F-GE180 PET imaging may be useful for longitudinal monitoring of neuroinflammation during AD progression and treatment and may also be useful for other neurodegenerative diseases., (Copyright © 2015 the authors 0270-6474/15/3515717-15$15.00/0.)

The complement system is part of the innate immune response responsible for removing pathogens and cellular debris, in addition to helping to refine CNS neuronal connections via microglia-mediated pruning of inappropriate synapses during brain development. However, less is known about the role of complement during normal aging. Here, we studied the role of the central complement component, C3, in synaptic health and aging. We examined behavior as well as electrophysiological, synaptic, and neuronal changes in the brains of C3-deficient male mice (C3 KO) compared with age-, strain-, and gender-matched C57BL/6J (wild-type, WT) control mice at postnatal day 30, 4 months, and 16 months of age. We found the following: (1) region-specific and age-dependent synapse loss in aged WT mice that was not observed in C3 KO mice; (2) age-dependent neuron loss in hippocampal CA3 (but not in CA1) that followed synapse loss in aged WT mice, neither of which were observed in aged C3 KO mice; and (3) significantly enhanced LTP and cognition and less anxiety in aged C3 KO mice compared with aged WT mice. Importantly, CA3 synaptic puncta were similar between WT and C3 KO mice at P30. Together, our results suggest a novel and prominent role for complement protein C3 in mediating aged-related and region-specific changes in synaptic function and plasticity in the aging brain. Significance statement: The complement cascade, part of the innate immune response to remove pathogens, also plays a role in synaptic refinement during brain development by the removal of weak synapses. We investigated whether complement C3, a central component, affects synapse loss during aging. Wild-type (WT) and C3 knock-out (C3 KO) mice were examined at different ages. The mice were similar at 1 month of age. However, with aging, WT mice lost synapses in specific brain regions, especially in hippocampus, an area important for memory, whereas C3 KO mice were protected. Aged C3 KO mice also performed better on learning and memory tests than aged WT mice. Our results suggest that complement C3, or its downstream signaling, is detrimental to synapses during aging., (Copyright © 2015 the authors 0270-6474/15/3513029-14$15.00/0.)

In this report we present a study of a series of Re(CO)3 pyridine-imine complexes with pendant phenol groups. We investigated the effects of the position of the phenol hydroxyl group (para, meta or ortho to the imine) on the steric and electronic characteristics of a series of Re(CO)3X(pyca-C6H4OH) compounds, where X = Cl, Br and pyca = pyridine-2-carbaldehyde imine. These compounds can be generated either via ligand synthesis followed by metal chelation (compound 4) or via a one-pot method (compounds 2, 3, 5 and 6). All six compounds show striking differences in pH-dependent UV-visible absorption based on the position of the phenol hydroxyl group.

Amyloid-β (Aβ) peptides, starting with pyroglutamate at the third residue (pyroGlu-3 Aβ), are a major species deposited in the brain of Alzheimer disease (AD) patients. Recent studies suggest that this isoform shows higher toxicity and amyloidogenecity when compared to full-length Aβ peptides. Here, we report the first comprehensive and comparative IHC evaluation of pyroGlu-3 Aβ deposition in humans and animal models. PyroGlu-3 Aβ immunoreactivity (IR) is abundant in plaques and cerebral amyloid angiopathy of AD and Down syndrome patients, colocalizing with general Aβ IR. PyroGlu-3 Aβ is further present in two nontransgenic mammalian models of cerebral amyloidosis, Caribbean vervets, and beagle canines. In addition, pyroGlu-3 Aβ deposition was analyzed in 12 different AD-like transgenic mouse models. In contrast to humans, all transgenic models showed general Aβ deposition preceding pyroGlu-3 Aβ deposition. The findings varied greatly among the mouse models concerning age of onset and cortical brain region. In summary, pyroGlu-3 Aβ is a major species of β-amyloid deposited early in diffuse and focal plaques and cerebral amyloid angiopathy in humans and nonhuman primates, whereas it is deposited later in a subset of focal and vascular amyloid in AD-like transgenic mouse models. Given the proposed decisive role of pyroGlu-3 Aβ peptides for the development of human AD pathology, this study provides insights into the usage of animal models in AD studies., (Copyright © 2013 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Avian influenza virus vaccines produced in oil-emulsified inactivated form with antigen content of at least 160 hemagglutinin units (HAU) induced immunity in birds. However, in addition to enhancing the effect of the adjuvant(s), other additional supplemented biological compounds included in inactivated vaccines could produce higher levels of antibody. We examined in chickens, Vietnamese ducks, and muscovy ducks the adjuvant effect of Sophy β-glucan (SBG), a β-1,3-1,6 glucan produced by the black yeast Aureobasidium pollulans strain AF0-202, when administered with an avian influenza H5 subtype vaccine. In Experiment 1, 40 chickens (ISA Brown hybrid), allocated to four groups of ten each, were immunized with Oil-H5N1(VN), Oil-H5N1(CN), Oil-H5N2(CN), and saline (control group), respectively. In Experiment 2, chickens (ISA Brown hybrid), muscovy ducks (French hybrid), and Vietnamese ducks (indigenous Vietnamese) were used to further assess the effect of SBG on immunogenicity of the Oil-H5N1(VN) Vietnamese vaccine. ELISA and hemagglutination inhibition (HI) assays were used to assess the antibody response. The H5 subtype vaccines initiated significantly higher immune responses in the animals dosed with SBG, with 1.0-1.5 log2 higher HI titers and 10-20% ELISA seroconversion, compared with those not dosed with β-glucan. Notably, some of the animals dosed with SBG induced HI titers higher than 9.0 log2 following boosting immunization. Taken together, our serial studies indicated that SBG is a potential effector, such as enhancing the immune response to the H5 vaccines tested.