Chronic Anatabine Treatment Reduces Alzheimer's Disease (AD)-Like Pathology and Improves Socio-Behavioral Deficits in a Transgenic Mouse Model of AD.
ABSTRACT: Anatabine is a minor tobacco alkaloid, which is also found in plants of the Solanaceae family and displays a chemical structure similarity with nicotine. We have shown previously that anatabine displays some anti-inflammatory properties and reduces microgliosis and tau phosphorylation in a pure mouse model of tauopathy. We therefore investigated the effects of a chronic oral treatment with anatabine in a transgenic mouse model (Tg PS1/APPswe) of Alzheimer's disease (AD) which displays pathological A? deposits, neuroinflammation and behavioral deficits. In the elevated plus maze, Tg PS1/APPswe mice exhibited hyperactivity and disinhibition compared to wild-type mice. Six and a half months of chronic oral anatabine treatment, suppressed hyperactivity and disinhibition in Tg PS1/APPswe mice compared to Tg PS1/APPswe receiving regular drinking water. Tg PS1/APPswe mice also elicited profound social interaction and social memory deficits, which were both alleviated by the anatabine treatment. We found that anatabine reduces the activation of STAT3 and NF?B in the vicinity of A? deposits in Tg PS1/APPswe mice resulting in a reduction of the expression of some of their target genes including Bace1, iNOS and Cox-2. In addition, a significant reduction in microgliosis and pathological deposition of A? was observed in the brain of Tg PS1/APPswe mice treated with anatabine. This is the first study to investigate the impact of chronic anatabine treatment on AD-like pathology and behavior in a transgenic mouse model of AD. Overall, our data show that anatabine reduces ?-amyloidosis, neuroinflammation and alleviates some behavioral deficits in Tg PS1/APPswe, supporting further exploration of anatabine as a possible disease modifying agent for the treatment of AD.
Project description:Insulin-like growth factor-1 (IGF-1) is a pleiotropic molecule with neurotrophic and immunomodulatory functions. Knowing the capacity of chronically activated microglia to produce IGF-1 may therefore show essential to promote beneficial microglial functions in Alzheimer's disease (AD). Here, we investigated the expression of IGF-1 mRNA and IGF-1 along with the expression of tumor necrosis factor (TNF) mRNA, and the amyloid-? (A?) plaque load in the hippocampus of 3- to 24-month-old APPswe/PS1?E9 transgenic (Tg) and wild-type (WT) mice. As IGF-1, in particular, is implicated in neurogenesis we also monitored the proliferation of cells in the subgranular zone (sgz) of the dentate gyrus. We found that the A? plaque load reached its maximum in aged 21- and 24-month-old APPswe/PS1?E9 Tg mice, and that microglial reactivity and hippocampal IGF-1 and TNF mRNA levels were significantly elevated in aged APPswe/PS1?E9 Tg mice. The sgz cell proliferation decreased with age, regardless of genotype and increased IGF-1/TNF mRNA levels. Interestingly, IGF-1 mRNA was expressed in subsets of sgz cells, likely neuroblasts, and neurons in both genotypes, regardless of age, as well as in glial-like cells. By double in situ hybridization these were shown to be IGF1 mRNA+ CD11b mRNA+ cells, i.e., IGF-1 mRNA-expressing microglia. Quantification showed a 2-fold increase in the number of microglia and IGF-1 mRNA-expressing microglia in the molecular layer of the dentate gyrus in aged APPswe/PS1?E9 Tg mice. Double-immunofluorescence showed that IGF-1 was expressed in a subset of A? plaque-associated CD11b+ microglia and in several subsets of neurons. Exposure of primary murine microglia and BV2 cells to A?42 did not affect IGF-1 mRNA expression. IGF-1 mRNA levels remained constant in WT mice with aging, unlike TNF mRNA levels which increased with aging. In conclusion, our results suggest that the increased IGF-1 mRNA levels can be ascribed to a larger number of IGF-1 mRNA-expressing microglia in the aged APPswe/PS1?E9 Tg mice. The finding that subsets of microglia retain the capacity to express IGF-1 mRNA and IGF-1 in the aged APPswe/PS1?E9 Tg mice is encouraging, considering the beneficial therapeutic potential of modulating microglial production of IGF-1 in AD.
Project description:Alzheimer's disease (AD) pathology occurs in part as the result of excessive production of ?-amyloid (A?). Metabotropic glutamate receptor 5 (mGluR5) is now considered a receptor for A? and consequently contributes to pathogenic A? signaling in AD.Genetic deletion of mGluR5 rescues the spatial learning deficits observed in APPswe/PS1?E9 AD mice. Moreover, both A? oligomer formation and A? plaque number are reduced in APPswe/PS1?E9 mice lacking mGluR5 expression. In addition to the observed increase in A? oligomers and plaques in APPswe/PS1?E9 mice, we found that both mTOR phosphorylation and fragile X mental retardation protein (FMRP) expression were increased in these mice. Genetic deletion of mGluR5 reduced A? oligomers, plaques, mTOR phosphorylation and FMRP expression in APPswe/PS1?E9 mice.Thus, we propose that A? activation of mGluR5 appears to initiate a positive feedback loop resulting in increased A? formation and AD pathology in APPswe/PS1?E9 mice via mechanism that is regulated by FMRP.
Project description:While amyloid-beta (A?) peptides play a central role in the development of Alzheimer's disease (AD), recent evidence also implicates altered metabolism of L-arginine in the pathogenesis of AD. The present study systematically investigated how behavioural function and the brain and plasma arginine metabolic profiles changed in a chronic A? accumulation model using male APPswe/PS1?E9 transgenic (Tg) mice at 7 and 13 months of age. As compared to their wild-type (WT) littermates, Tg mice displayed age-related deficits in spatial water maze tasks and alterations in brain arginine metabolism. Interestingly, the plasma arginine metabolic profile was markedly altered in 7-month Tg mice prior to major behavioural impairment. Receiver operating characteristic curve analysis revealed that plasma putrescine and spermine significantly differentiated between Tg and WT mice. These results demonstrate the parallel development of altered brain arginine metabolism and behavioural deficits in Tg mice. The altered plasma arginine metabolic profile that preceded the behavioural and brain profile changes suggests that there may be merit in an arginine-centric set of ante-mortem biomarkers for AD.
Project description:BACKGROUND:The pathological hallmarks of Alzheimer's disease (AD) involve alterations in the expression of numerous genes associated with transcriptional levels, which are determined by chromatin accessibility. Here, the landscape of chromatin accessibility was studied to understand the outline of the transcription and expression of AD-associated metabolism genes in an AD mouse model. METHODS:The assay for transposase-accessible chromatin by sequencing (ATAC-seq) was used to investigate the AD-associated chromatin reshaping in the APPswe/PS1dE9 (APP/PS1) mouse model. ATAC-seq data in the hippocampus of 8-month-old APP/PS1 mice were generated, and the relationship between chromatin accessibility and gene expression was analyzed in combination with RNA sequencing. Gene ontology (GO) analysis was applied to elucidate biological processes and signaling pathways altered in APP/PS1 mice. Critical transcription factors were identified; alterations in chromatin accessibility were further confirmed using chromatin immunoprecipitation assays. RESULTS:We identified 1690 increased AD-associated chromatin-accessible regions in the hippocampal tissues of APP/PS1 mice. These regions were enriched in genes related to diverse signaling pathways, including the PI3K-Akt, Hippo, TGF-?, and Jak-Stat signaling pathways, which play essential roles in regulating cell proliferation, apoptosis, and inflammatory responses. A total of 1003 decreased chromatin-accessible regions were considered to be related with declined AD-associated biological processes including cellular response to hyperoxia and insulin stimulus, synaptic transmission, and positive regulation of autophagy. In the APP/PS1 hippocampus, 1090 genes were found to be upregulated and 1081 downregulated. Interestingly, enhanced ATAC-seq signal was found in approximately 740 genes, with 43 exhibiting upregulated mRNA levels. Several genes involved in AD development were found to have a significantly increased expression in APP/PS1 mice compared to controls, including Sele, Clec7a, Cst7, and Ccr6. The signatures of numerous transcription factors, including Olig2, NeuroD1, TCF4, and NeuroG2, were found enriched in the AD-associated accessible chromatin regions. The transcription-activating marks of H3K4me3 and H3K27ac were also found increased in the promoters of these genes. These results indicate that the mechanism for the upregulation of genes could be attributed to the enrichment of open chromatin regions with transcription factors motifs and the histone marks H3K4me3 and H3K27ac. CONCLUSION:Our study reveals that alterations in chromatin accessibility may be an initial mechanism in AD pathogenesis.
Project description:BACKGROUND:Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by an abnormal accumulation of amyloid-? (A?) plaques, neuroinflammation, and impaired neurogenesis. Urolithin A (UA), a gut-microbial metabolite of ellagic acid, has been reported to exert anti-inflammatory effects in the brain. However, it is unknown whether UA exerts its properties of anti-inflammation and neuronal protection in the APPswe/PS1?E9 (APP/PS1) mouse model of AD. METHODS:Morris water maze was used to detect the cognitive function. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay was performed to detect neuronal apoptosis. Immunohistochemistry analyzed the response of glia, A? deposition, and neurogenesis. The expression of inflammatory mediators were measured by enzyme-linked immunosorbent assay (ELISA) and quantitative real-time polymerase chain reaction (qRT-PCR). The modulating effects of UA on cell signaling pathways were assayed by Western blotting. RESULTS:We demonstrated that UA ameliorated cognitive impairment, prevented neuronal apoptosis, and enhanced neurogenesis in APP/PS1 mice. Furthermore, UA attenuated A? deposition and peri-plaque microgliosis and astrocytosis in the cortex and hippocampus. We also found that UA affected critical cell signaling pathways, specifically by enhancing cerebral AMPK activation, decreasing the activation of P65NF-?B and P38MAPK, and suppressing Bace1 and APP degradation. CONCLUSIONS:Our results indicated that UA imparted cognitive protection by protecting neurons from death and triggering neurogenesis via anti-inflammatory signaling in APP/PS1 mice, suggesting that UA might be a promising therapeutic drug to treat AD.
Project description:Back-translation of clinical imaging biomarkers of Alzheimer's disease (AD), such as alterations in cerebral glucose metabolism detected by [18F]FDG positron emission tomography (PET), would be valuable for preclinical studies evaluating new disease-modifying drugs for AD. However, previous confounding results have been difficult to interpret due to differences in mouse models and imaging protocols between studies. We used an equivalent study design and [18F]FDG µPET imaging protocol to compare changes in cerebral glucose metabolism in commercial transgenic APPSwe-PS1dE9 (n = 12), Tg2576 (n = 15), and wild-type mice (n = 15 and 9). Dynamic [18F]FDG scans were performed in young (6 months) and aged (12 or 17 months) mice and the results verified by ex vivo methods (i.e., tissue counting, digital autoradiography, and beta-amyloid and Iba-1 immunohistochemistry). [18F]FDG uptake exhibited significant regional differences between genotypes (TG < WT) and ages (6 months <12 months) in the APPSwe-PS1dE9 model, whereas similar differences were not present in Tg2576 mice. In both models, only weak correlations were detected between regional beta-amyloid deposition or microgliosis and [18F]FDG uptake. By using equivalent methodology, this study demonstrated differences in cerebral glucose metabolism dysfunction detected with [18F]FDG PET between two widely used commercial AD mouse models.
Project description:Despite compelling evidence that the accumulation of amyloid-beta (A?) promotes neocortical MAPT (tau) aggregation in familial and idiopathic Alzheimer's disease (AD), murine models of cerebral amyloidosis are not considered to develop tau-associated pathology. In the present study, we show that tau can accumulate spontaneously in aged transgenic APPswe/PS1?E9 mice. Tau pathology is abundant around A? deposits, and further characterized by accumulation of Gallyas and thioflavin-S-positive inclusions, which were detected in the APPswe/PS1?E9 brain at 18 months of age. Age-dependent increases in argyrophilia correlated positively with binding levels of the paired helical filament (PHF) tracer [18F]Flortaucipir, in all brain areas examined. Sarkosyl-insoluble PHFs were visualized by electron microscopy. Quantitative proteomics identified sequences of hyperphosphorylated and three-repeat tau in transgenic mice, along with signs of RNA missplicing, ribosomal dysregulation and disturbed energy metabolism. Tissue from the frontal gyrus of human subjects was used to validate these findings, revealing primarily quantitative differences between the tau pathology observed in AD patient vs. transgenic mouse tissue. As physiological levels of endogenous, 'wild-type' tau aggregate secondarily to A? in APPswe/PS1?E9 mice, this study suggests that amyloidosis is both necessary and sufficient to drive tauopathy in experimental models of familial AD.
Project description:BACKGROUND:Pyroglutamate-3 A? (pGlu-3 A?) is an N-terminally truncated and post-translationally modified A? 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 A? 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 A? mAb (07/1 IgG1 and 07/2a IgG2a) and a general N-terminal A? mAb (3A1 IgG1) for their ability to clear A? and protect cognition in a therapeutic passive immunotherapy study in aged, plaque-rich APPSWE/PS1?E9 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 A? in an ex vivo phagocytosis assay and following treatment in APPSLxhQC double Tg mice, and to activate microglia using longitudinal microPET imaging with TSPO-specific 18F-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 APPSWE/PS1?E9 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 Fc? receptors CD32 and CD64, although the two antibodies had similar binding affinities to pGlu-3 A?. Treatment of APPSLxhQC mice with 07/2a and 07/2a-k mAbs in our second in vivo study showed significant plaque-lowering with both mAbs. Longitudinal 18F-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 A? 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.
Project description:Recent evidence suggests the commensal microbiome regulates host immunity and influences brain function; findings that have ramifications for neurodegenerative diseases. In the context of Alzheimer's disease (AD), we previously reported that perturbations in microbial diversity induced by life-long combinatorial antibiotic (ABX) selection pressure in the APPSWE/PS1?E9 mouse model of amyloidosis is commensurate with reductions in amyloid-? (A?) plaque pathology and plaque-localised gliosis. Considering microbiota-host interactions, specifically during early post-natal development, are critical for immune- and neuro-development we now examine the impact of microbial community perturbations induced by acute ABX exposure exclusively during this period in APPSWE/PS1?E9 mice. We show that early post-natal (P) ABX treatment (P14-P21) results in long-term alterations of gut microbial genera (predominantly Lachnospiraceae and S24-7) and reduction in brain A? deposition in aged APPSWE/PS1?E9 mice. These mice exhibit elevated levels of blood- and brain-resident Foxp3+ T-regulatory cells and display an alteration in the inflammatory milieu of the serum and cerebrospinal fluid. Finally, we confirm that plaque-localised microglia and astrocytes are reduced in ABX-exposed mice. These findings suggest that ABX-induced microbial diversity perturbations during post-natal stages of development coincide with altered host immunity mechanisms and amyloidosis in a murine model of AD.
Project description:[18F]FPEB is a positron emission tomography (PET) radiopharmaceutical used for imaging the abundance and distribution of mGluR5 in the central nervous system (CNS). Efficient radiolabeling of the aromatic ring of [18F]FPEB has been an ongoing challenge. Herein, five metal-free precursors for the radiofluorination of [18F]FPEB were compared, namely, a chloro-, nitro-, sulfonium salt, and two spirocyclic iodonium ylide (SCIDY) precursors bearing a cyclopentyl (SPI5) and a new adamantyl (SPIAd) auxiliary. The chloro- and nitro-precursors resulted in a low radiochemical yield (<10% RCY), whereas both SCIDY precursors and the sulfonium salt precursor produced [18F]FPEB in the highest RCYs of 25% and 36%, respectively. Preliminary PET/CT imaging studies with [18F]FPEB were conducted in a transgenic model of Alzheimer's Disease (AD) using B6C3-Tg(APPswe,PSEN1dE9)85Dbo/J (APP/PS1) mice, and data were compared with age-matched wild-type (WT) B6C3F1/J control mice. In APP/PS1 mice, whole brain distribution at 5 min post-injection showed a slightly higher uptake (SUV = 4.8 ± 0.4) than in age-matched controls (SUV = 4.0 ± 0.2). Further studies to explore mGluR5 as an early biomarker for AD are underway.