The ability to investigate therapeutic interventions in animal models of neurodegenerative diseases depends on extensive characterization of the model(s) being used. There are numerous models that ...have been generated to study Alzheimer's disease (AD) and the underlying pathogenesis of the disease. While transgenic models have been instrumental in understanding AD mechanisms and risk factors, they are limited in the degree of characteristics displayed in comparison with AD in humans, and the full spectrum of AD effects has yet to be recapitulated in a single mouse model. The Model Organism Development and Evaluation for Late-Onset Alzheimer's Disease (MODEL-AD) consortium was assembled by the National Institute on Aging (NIA) to develop more robust animal models of AD with increased relevance to human disease, standardize the characterization of AD mouse models, improve preclinical testing in animals, and establish clinically relevant AD biomarkers, among other aims toward enhancing the translational value of AD models in clinical drug design and treatment development. Here we have conducted a detailed characterization of the 5XFAD mouse, including transcriptomics, electroencephalogram,
imaging, biochemical characterization, and behavioral assessments. The data from this study is publicly available through the AD Knowledge Portal.
Variants identified in genome‐wide association studies have implicated immune pathways in the development of Alzheimer’s disease (AD). Here, we investigated the mechanistic basis for protection from ...AD associated with PLCγ2 R522, a rare coding variant of the PLCG2 gene. We studied the variant's role in macrophages and microglia of newly generated PLCG2‐R522‐expressing human induced pluripotent cell lines (hiPSC) and knockin mice, which exhibit normal endogenous PLCG2 expression. In all models, cells expressing the R522 mutation show a consistent non‐redundant hyperfunctionality in the context of normal expression of other PLC isoforms. This manifests as enhanced release of cellular calcium ion stores in response to physiologically relevant stimuli like Fc‐receptor ligation or exposure to Aβ oligomers. Expression of the PLCγ2‐R522 variant resulted in increased stimulus‐dependent PIP2 depletion and reduced basal PIP2 levels in vivo. Furthermore, it was associated with impaired phagocytosis and enhanced endocytosis. PLCγ2 acts downstream of other AD‐related factors, such as TREM2 and CSF1R, and alterations in its activity directly impact cell function. The inherent druggability of enzymes such as PLCγ2 raises the prospect of PLCγ2 manipulation as a future therapeutic approach in AD.
SYNOPSIS
The Alzheimer’s disease protective variant of PLCG2, R522, is hyperfunctional and alters endocytic and phagocytic uptake by macrophages and microglia. Hyperfunctionality of the R522 variant can lead to substrate depletion, which would have implications for therapeutic approaches.
We observed a clear consistent increase in enzymatic activation in both human and mouse microglia and macrophages due to the Alzheimer’s disease protective variant of PLCG2 (R522).
Hyperactivity of PLCγ2‐R522 results in substrate depletion in macrophages and microglia.
Cells expressing the R522 variant exhibit alterations in the phagocytic and endocytic activities.
Expression analyses in hIPSCs and knock‐in mice implicate enhanced Ca2+ release and stimulus‐dependent PIP2 depletion in macrophages and microglia in Alzheimer's disease protection associated with a rare PLCG2 coding variant.
Affymetrix and spotted oligonucleotide microarrays were used to assess global differential gene expression comparing normal human melanocytes with six independent melanoma cell strains from advanced ...lesions. The data, validated at the protein level for selected genes, confirmed the overexpression in melanoma cells relative to normal melanocytes of several genes in the growth factor/receptor family that confer growth advantage and metastasis. In addition, novel pathways and patterns of associated expression in melanoma cells not reported before emerged, including the following: (a) activation of the NOTCH pathway; (b) increased Twist expression and altered expression of additional transcriptional regulators implicated in embryonic development and epidermal/mesenchymal transition; (c) coordinated activation of cancer/testis antigens; (d) coordinated down-regulation of several immune modulation genes, in particular in the IFN pathways; (e) down-regulation of several genes implicated in membrane trafficking events; and (f) down-regulation of growth suppressors, such as the Prader-Willi gene NECDIN, whose function was confirmed by overexpression of ectopic Flag-necdin. Validation of differential expression using melanoma tissue microarrays showed that reduced ubiquitin COOH-terminal esterase L1 in primary melanoma is associated with worse outcome and that increased expression of the basic helix-loop-helix protein Twist is associated with worse outcome. Some differentially expressed genes reside on chromosomal regions displaying common loss or gain in melanomas or are known to be regulated by CpG promoter methylation. These results provide a comprehensive view of changes in advanced melanoma relative to normal melanocytes and reveal new targets that can be used in assessing prognosis, staging, and therapy of melanoma patients.
Our objective was to identify if changes in serum protein concentrations occur in hyperthyroidism and to assess their association with the development of azotaemia following treatment.
Initially ...non-azotaemic hyperthyroid cats and healthy older cats were included. Serum concentrations of protein fractions were determined by agarose gel electrophoresis and compared between; hyperthyroid and control cats, initially non-azotaemic hyperthyroid cats which developed azotaemia in a 4month follow up period (masked-azotaemic) and those which remained non-azotaemic, and hyperthyroid cats before and at the time of restoration of euthyroidism. Data are presented as median 25th, 75th percentiles.
Hyperthyroid cats (n=56) had higher serum α2 globulin concentrations (12.5 10.9, 13.1 g/L vs. 9.8 3.0, 11.4 g/L; P<0.001) and lower serum γ globulin concentrations (11.4 9.1, 13.3 g/L vs. 14.0 12.4, 16.8 g/L; P=0.001) than control cats (n=26). Following treatment, serum total globulin concentration increased (from 38.6 35.4, 42.8 g/L to 42.3 39.0, 45.7 g/L; P<0.001), serum α2 globulin concentration decreased (from 12.5 10.9, 13.9 g/L to 11.5 10.1, 12.6 g/L; P<0.001) and serum γ globulin concentration increased (from 11.4 9.0, 13.3 g/L to 14.0 12.4, 16.8 g/L; P<0.001). Serum concentrations of total globulin or globulin fractions were not significantly different between masked-azotaemic and non azotaemic groups.
In conclusion, hyperthyroidism is associated with altered serum concentrations of the α2 and γ globulin fractions, however these changes were not associated with the development of azotaemic chronic kidney disease following treatment.
•Biomarkers of concurrent, but masked, azotaemic chronic kidney disease (CKD) in hyperthyroid cats are lacking.•Hyperthyroid cats demonstrate mild changes in the serum alpha-2 and gamma globulins that resolve following treatment.•Serum concentrations of all globulin fractions and total globulin are not associated with the development of azotaemia.•Serum concentrations of globulin fractions and total globulin are not biomarkers of masked azotaemic CKD in hyperthyroidism.
Introduction
Hyperexcitability and epileptiform activity are commonplace in Alzheimer's disease (AD) patients and associated with impaired cognitive function. The anti‐seizure drug levetiracetam ...(LEV) is currently being evaluated in clinical trials for ability to reduce epileptiform activity and improve cognitive function in AD. The purpose of our studies was to establish a pharmacokinetic/pharmacodynamic (PK/PD) relationship with LEV in an amyloidogenic mouse model of AD to enable predictive preclinical to clinical translation, using the rigorous preclinical testing pipeline of the Model Organism Development and Evaluation for Late‐Onset Alzheimer's Disease Preclinical Testing Core.
Methods
A multi‐tier approach was applied that included quality assurance and quality control of the active pharmaceutical ingredient, PK/PD modeling, positron emission tomography/magnetic resonance imaging (PET/MRI), functional outcomes, and transcriptomics. 5XFAD mice were treated chronically with LEV for 3 months at doses in line with those allometrically scaled to the clinical dose range.
Results
Pharmacokinetics of LEV demonstrated sex differences in Cmax, AUC0‐∞, and CL/F, and a dose dependence in AUC0‐∞. After chronic dosing at 10, 30, 56 mg/kg, PET/MRI tracer 18F‐AV45, and 18F‐fluorodeoxyglucose (18F‐FDG) showed specific regional differences with treatment. LEV did not significantly improve cognitive outcomes. Transcriptomics performed by nanoString demonstrated drug‐ and dose‐related changes in gene expression relevant to human brain regions and pathways congruent with changes in 18F‐FDG uptake.
Discussion
This study represents the first report of PK/PD assessment of LEV in 5XFAD mice. Overall, these results highlighted non‐linear kinetics based on dose and sex. Plasma concentrations of the 10 mg/kg dose in 5XFAD overlapped with human plasma concentrations used for studies of mild cognitive impairment, while the 30 and 56 mg/kg doses were reflective of doses used to treat seizure activity. Post‐treatment gene expression analysis demonstrated LEV dose‐related changes in immune function and neuronal‐signaling pathways relevant to human AD, and aligned with regional 18F‐FDG uptake. Overall, this study highlights the importance of PK/PD relationships in preclinical studies to inform clinical study design.
Highlights
Significant sex differences in pharmacokinetics of levetiracetam were observed in 5XFAD mice.
Plasma concentrations of 10 mg/kg levetiracetam dose in 5XFAD overlapped with human plasma concentration used in the clinic.
Drug‐ and dose‐related differences in gene expression relevant to human brain regions and pathways were also similar to brain region–specific changes in 18F‐fluorodeoxyglucose uptake.
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