The 5xFAD mouse is a popular model of familial Alzheimer's disease (AD) that is characterized by early beta-amyloid (Aβ) deposition and cognitive decrements. Despite numerous studies, the 5xFAD mouse ...has not been comprehensively phenotyped for vascular and metabolic perturbations over its lifespan.
Male and female 5xFAD and wild type (WT) littermates underwent
F-fluorodeoxyglucose (FDG) positron emission tomography (PET) imaging at 4, 6, and 12 months of age to assess regional glucose metabolism. A separate cohort of mice (4, 8, 12 months) underwent "vessel painting" which labels all cerebral vessels and were analyzed for vascular characteristics such as vessel density, junction density, vessel length, network complexity, number of collaterals, and vessel diameter.
With increasing age, vessels on the cortical surface in both 5xFAD and WT mice showed increased vessel length, vessel and junction densities. The number of collateral vessels between the middle cerebral artery (MCA) and the anterior and posterior cerebral arteries decreased with age but collateral diameters were significantly increased only in 5xFAD mice. MCA total vessel length and junction density were decreased in 5xFAD mice compared to WT at 4 months. Analysis of
F-FDG cortical uptake revealed significant differences between WT and 5xFAD mice spanning 4-12 months. Broadly, 5xFAD males had significantly increased
F-FDG uptake at 12 months compared to WT mice. In most cortical regions, female 5xFAD mice had reduced
F-FDG uptake compared to WT across their lifespan.
While the 5xFAD mouse exhibits AD-like cognitive deficits as early as 4 months of age that are associated with increasing Aβ deposition, we only found significant differences in cortical vascular features in males, not in females. Interestingly, 5xFAD male and female mice exhibited opposite effects in
F-FDG uptake. The MCA supplies blood to large portions of the somatosensory cortex and portions of motor and visual cortex and increased vessel length alongside decreased collaterals which coincided with higher metabolic rates in 5xFAD mice. Thus, a potential mismatch between metabolic demand and vascular delivery of nutrients in the face of increasing Aβ deposition could contribute to the progressive cognitive deficits seen in the 5xFAD mouse model.
Vascular contributions to cognitive impairment and dementia (VCID) particularly Alzheimer’s disease and related dementias (ADRDs) are increasing; however, mechanisms driving cerebrovascular decline ...are poorly understood. Methylenetetrahydrofolate reductase (MTHFR) is a critical enzyme in the folate and methionine cycles. Variants in MTHFR, notably 677 C > T, are associated with dementias, but no mouse model existed to identify mechanisms by which MTHFR677C > T increases risk. Therefore, MODEL-AD created a novel knock-in (KI) strain carrying the Mthfr677C > T allele on the C57BL/6J background (Mthfr677C > T) to characterize morphology and function perturbed by the variant. Consistent with human clinical data, Mthfr677C > T mice have reduced enzyme activity in the liver and elevated plasma homocysteine levels. MTHFR enzyme activity is also reduced in the Mthfr677C > T brain. Mice showed reduced tissue perfusion in numerous brain regions by PET/CT as well as significantly reduced vascular density, pericyte number and increased GFAP-expressing astrocytes in frontal cortex. Electron microscopy revealed cerebrovascular damage including endothelial and pericyte apoptosis, reduced luminal size, and increased astrocyte and microglial presence in the microenvironment. Collectively, these data support a mechanism by which variations in MTHFR perturb cerebrovascular health laying the foundation to incorporate our new Mthfr677C > T mouse model in studies examining genetic susceptibility for cerebrovascular dysfunction in ADRDs.
Lead (Pb) is an environmental factor has been suspected of contributing to the dementia including Alzheimer’s disease (AD). Our previous studies have shown that Pb exposure at the subtoxic dose ...increased brain levels of beta-amyloid (Aβ) and amyloid plaques, a pathological hallmark for AD, in amyloid precursor protein (APP) transgenic mice, and is hypothesized to inhibit Aβ clearance in the blood- cerebrospinal fluid (CSF) barrier. However, it remains unclear how different levels of Pb affect Aβ clearance in the whole blood-brain barrier system. This study was designed to investigate whether chronic exposure of Pb affected the permeability of the blood-brain barrier system by using the Dynamic Contrast-Enhanced Computerized Tomography (DCE-CT) method.
DEC-CT was used to investigate whether chronic exposure of toxic Pb affected the permeability of the real-time blood brain barrier system.
Data showed that Pb exposure increased permeability surface area product, and also significantly induced brain perfusion. However, Pb exposure did not alter extracellular volumes or fractional blood volumes of mouse brain.
Our data suggest that Pb exposure at subtoxic and toxic levels directly targets the brain vasculature and damages the blood brain barrier system.
Obesity is recognized as a significant risk factor for Alzheimer’s disease (AD). Studies have supported the notion that obesity accelerates AD-related pathophysiology in mouse models of AD. The ...majority of studies, to date, have focused on the use of early-onset AD models. Here, we evaluate the impact of genetic risk factors on late-onset AD (LOAD) in mice fed with a high fat/high sugar diet (HFD). We focused on three mouse models created through the IU/JAX/PITT MODEL-AD Center. These included a combined risk model with
APOE4
and a variant in triggering receptor expressed on myeloid cells 2 (
Trem2
R47H
). We have termed this model, LOAD1. Additional variants including the M28L variant in phospholipase C Gamma 2 (
Plcg2
M28L
) and the 677C > T variant in methylenetetrahydrofolate reductase (
Mthfr
677C >
T
) were engineered by CRISPR onto LOAD1 to generate LOAD1.
Plcg2
M28L
and LOAD1.
Mthfr
677C >
T
. At 2 months of age, animals were placed on an HFD that induces obesity or a control diet (CD), until 12 months of age. Throughout the study, blood was collected to assess the levels of cholesterol and glucose. Positron emission tomography/computed tomography (PET/CT) was completed prior to sacrifice to image for glucose utilization and brain perfusion. After the completion of the study, blood and brains were collected for analysis. As expected, animals fed a HFD, showed a significant increase in body weight compared to those fed a CD. Glucose increased as a function of HFD in females only with cholesterol increasing in both sexes. Interestingly, LOAD1.
Plcg2
M28L
demonstrated an increase in microglia density and alterations in regional brain glucose and perfusion on HFD. These changes were not observed in LOAD1 or LOAD1.
Mthfr
677C >
T
animals fed with HFD. Furthermore, LOAD1.
Plcg2
M28L
but not LOAD1.
Mthfr
677C >
T
or LOAD1 animals showed transcriptomics correlations with human AD modules. Our results show that HFD affects the brain in a genotype-specific manner. Further insight into this process may have significant implications for the development of lifestyle interventions for the treatment of AD.
Background
The Preclinical Testing Core (PTC) of the Model Organism Development for Evaluation of Late Onset Alzheimer’s Disease (MODEL‐AD) consortium established a rigorous preclinical drug testing ...strategy with go/no‐go decision points that permits unbiased assessments of therapeutic agents. As part of the pipeline validation, the chimeric murinized therapeutic antibody aducanumab (chAducanumab), was selected for evaluation in 5XFAD mice.
Methods
Initial PK modeling and simulation was guided by literature and Aβ reductions from a pilot cohort of 9 month aged 5XFAD mice following 1x/week treatment of 30 mg/kg chAducanumab for 4 weeks. These pilot data were used to inform the chronic dosing regimen for the PD study which started at an age in 5XFAD mice where significant amyloid plaque accumulation was present (9 mos). PD endpoints (n=10‐12/sex/genotype/treatment) were assessed at the conclusion of chronic treatment, and included: 18‐FDG and 18F‐AV45 PET/CT, autoradiography, immunohistochemistry, AB40 and AB42 in plasma and brain fractions, and a behavioral battery. An additional cohort was enrolled for comprehensive cognitive testing using touchscreen learning and pattern separation tasks and evaluated for electroencephalography (EEG) activity using wireless telemetry.
Results
PK/PD modeling revealed slow clearance of chAducanumab following IP dosing with a T1/2 of ∼2.5 days. Therefore, the dose regimen for chronic PD studies included 0.1, 1.56, and 30 mg/kg administered 1x weekly for 12 weeks. Treatment with chAducanumab resulted in dose‐ and sex‐dependent reduction in amyloid deposition via 18F‐AV45 PET. Glucose uptake via 18F‐FDG PET similarly showed a dose dependent reversal of glycolytic loss in key brain regions. Cognitive assessments indicated no effect on learning however an improvement in pattern separation was observed with chAducanumab in females but not males. EEG analysis revealed improvements in delta, alpha, and beta oscillations with chAducanumab treatment. Multi‐omics analysis are in progress.
Conclusions
chAducanumab treatment in 5XFAD mice resulted in the expected reductions in brain amyloid consistent with clinical findings. Moreover, chAducanumab showed a unique glycolytic restoration profile in 5XFAD mice and improvements in some aspects of cognitive function. Together these data positively support pipeline validation of the MODEL‐AD PTC for evaluating therapeutic antibodies.
Introduction
Alzheimer's disease (AD) is the most common form of dementia. Beta‐secretase (BACE) inhibitors have been proposed as potential therapeutic interventions; however, initiating treatment ...once disease has significantly progressed has failed to effectively stop or treat disease. Whether BACE inhibition may have efficacy when administered prophylactically in the early stages of AD has been under‐investigated. The present studies aimed to evaluate prophylactic treatment of the BACE inhibitor verubecestat in an AD mouse model using the National Institute on Aging (NIA) resources of the Model Organism Development for Late‐Onset Alzheimer's Disease (MODEL‐AD) Preclinical Testing Core (PTC) Drug Screening Pipeline.
Methods
5XFAD mice were administered verubecestat ad libitum in chow from 3 to 6 months of age, prior to the onset of significant disease pathology. Following treatment (6 months of age), in vivo imaging was conducted with 18F‐florbetapir (AV‐45/Amyvid) (18F‐AV45) and 18‐FDG (fluorodeoxyglucose)–PET (positron emission tomography)/MRI (magnetic resonance imaging), brain and plasma amyloid beta (Aβ) were measured, and the clinical and behavioral characteristics of the mice were assessed and correlated with the pharmacokinetic data.
Results
Prophylactic verubecestat treatment resulted in dose‐ and region‐dependent attenuations of 18F‐AV45 uptake in male and female 5XFAD mice. Plasma Aβ40 and Aβ42 were also dose‐dependently attenuated with treatment. Across the dose range evaluated, side effects including coat color changes and motor alterations were reported, in the absence of cognitive improvement or changes in 18F‐FDG uptake.
Discussion
Prophylactic treatment with verubecestat resulted in attenuated amyloid plaque deposition when treatment was initiated prior to significant pathology in 5XFAD mice. At the same dose range effective at attenuating Aβ levels, verubecestat produced side effects in the absence of improvements in cognitive function. Taken together these data demonstrate the rigorous translational approaches of the MODEL‐AD PTC for interrogating potential therapeutics and provide insight into the limitations of verubecestat as a prophylactic intervention for early‐stage AD.
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.
Background
As research efforts to discover and develop promising future therapeutics continues, the necessity to develop clinically significant Alzheimer’s disease (AD) mouse models and analysis is ...more important than ever.
Method
The study utilizes a novel MODEL‐AD (Model Organism Development and Evaluation for Late‐onset AD) mouse model that incorporates APOE4, Trem2*R47H, and humanized amyloid‐beta (Aβ) allele into C57BL/6J (B6) mice to produce LOAD2. Two cohorts of male and female LOAD2 mice received either control or high fat diet (HFD) followed by brain metabolism imaging using 18F‐FDG PET/CT. Brain regions were segmented using the Paxinos‐Franklin atlas and analyzed using a whole brain neurovascular uncoupling and connectivity approach. To do this, we computed z‐score statistics for all mice relative their control diet group, and conducted hierarchical modularization of all 28 brain regions and statically compared results across sex and high‐risk diet in 12‐month‐old mice.
Result
An initial network analysis detected metabolic hypo‐perfusion and metabolism for female mice, while males showed neurovascular uncoupling. Hierarchical analysis revealed primary modules within the whole brain network. These primary modules were then further modularized to yield a final breakdown of six secondary sub‐modules consisting of 3‐7 brain regions. Predominate functional associations of the sub‐modules’ brain regions were sensory (S) and learning (L), while sub‐module 2.1 had the strongest visual (V) representation. Statistical analysis of the six sub‐modules exhibited significant, module‐, sex‐, and dietary‐dependent effect for LOAD2 mice on high fat diet (HFD) by 12mo.
Conclusion
The incorporation of APOE4, Trem2*R47H, and humanized Ab sequence in combination with HFD induced age‐dependent LOAD‐relevant changes in neurovascular coupling and whole brain network connectivity consistent with known brain circuit variations observed clinically. These findings support application of this newer LOAD2 mouse model to improve knowledge regarding disease mechanism. Additionally, our connectivity analysis approach shows promise for implementation in future therapeutic development and testing.
Background
The Model Organism Development and Evaluation for Late‐Onset Alzheimer’s Disease (MODEL‐AD) Consortium seeks to develop the next generation of AD models based on human data. A popular ...model of familial AD is the 5xFAD mouse. It is characterized by early amyloid‐β deposition and cognitive decrements. Despite numerous studies, the 5xFAD mouse has not been comprehensively phenotyped for vascular and metabolic aspects over its lifespan.
Method
Males and females 5xFAD and WT littermates underwent in vivo 18F‐FDG‐PET imaging at 4, 6, and 12 months to evaluate regional glucose metabolism. A separate cohort of mice (4, 8, 12 months) underwent “vessel painting” that labels all cerebral vessels with a fluorescent dye. Brains were analyzed for vascular characteristics such as vessel and junction density, vessel length, network complexity, number and diameter of collaterals.
Result
Our analyses revealed that vessel length, vessel and junction densities increased from 4 to 12 months on the cortical surface in both 5xFAD and WT mice. The number of collateral vessels between the middle cerebral artery (MCA) and the anterior and posterior cerebral arteries decreased with age but interestingly their diameters were significantly increased only in 5xFAD mice. MCA average vessel length was significantly decreased at 8 and 12 months in 5xFAD mice compared to WT; primarily driven by males. Analysis of 18F‐FDG cortical uptake found significant interactions between WT and 5xFAD mice spanning 4‐12 months of age in retrosplenial somatosensory and visual cortices. Broadly, 5xFAD males had increased 18F‐FDG uptake at 12 months of age compared to WT mice. In most cortical regions, female 5xFAD mice had reduced FDG uptake compared to WT across the lifespan. In males these metabolic increases coincided with decreased vessel characteristics.
Conclusion
The 5xFAD mouse exhibits AD‐like cognitive deficits with age that are associated with increasing amyloid‐β deposition. No significant differences were found in cortical vascular features although males and females exhibited opposite effects in 18F‐FDG uptake. The MCA supplies blood to large portions of the motor cortex and increased vessel lengths and decreased collaterals along with higher metabolic rates in 5xFAD mice may be related to increasing behavioral deficits via metabolic insufficiency or other mechanisms.
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