Bisecting N-Acetylglucosamine Correlates with Phospho-Tau181 in Subjective Cognitive Decline but not in Control Cases.

Background: The N-glycan structure bisecting N-acetylglucosamine (bisecting GlcNAc) is present on several N-glycans that are elevated in Alzheimer's disease (AD), and previous studies have shown that bisecting GlcNAc levels correlate with total tau and phospho-tau181 in cerebrospinal fluid at early stages of AD. A recent population-based study showed that bisecting GlcNAc correlates with total tau also in blood and that this correlation could predict conversion to dementia. Objective: In this study, we have further investigated how bisecting GlcNAc relates to total tau and phospho-tau 181 in cerebrospinal fluid samples from controls and cases with early cognitive deficits, stratified by amyloid/tau status and gender. Methods: Relative levels of bisecting GlcNAc in cerebrospinal fluid were measured by an enzyme-linked lectin assay in individuals with subjective cognitive decline, mild cognitive impairment and controls from the Norwegian Dementia Disease Initiation cohort. Results: As in our previous study, the correlation between bisecting GlcNAc and total tau or phospho-tau181 was particularly strong in the subjective cognitive decline group. The correlation was observed in amyloid negative and tau negative as well as amyloid positive and tau positive individuals, both in females and in males. Interestingly, among the amyloid negative and tau negative individuals, the correlation was observed in individuals with subjective cognitive decline but not in the controls. Conclusions: Thus, bisecting GlcNAc could be a biomarker for early cognitive decline.

In-vivo neuronal dysfunction by Aß and tau overlaps with brain-wide inflammatory mechanisms in Alzheimer's disease.

The molecular mechanisms underlying neuronal dysfunction in Alzheimer's disease (AD) remain uncharacterized. Here, we identify genes, molecular pathways and cellular components associated with whole-brain dysregulation caused by amyloid-beta (Aß) and tau deposits in the living human brain. We obtained in-vivo resting-state functional MRI (rs-fMRI), Aß- and tau-PET for 47 cognitively unimpaired and 16 AD participants from the Translational Biomarkers in Aging and Dementia cohort. Adverse neuronal activity impacts by Aß and tau were quantified with personalized dynamical models by fitting pathology-mediated computational signals to the participant's real rs-fMRIs. Then, we detected robust brain-wide associations between the spatial profiles of Aß-tau impacts and gene expression in the neurotypical transcriptome (Allen Human Brain Atlas). Within the obtained distinctive signature of in-vivo neuronal dysfunction, several genes have prominent roles in microglial activation and in interactions with Aß and tau. Moreover, cellular vulnerability estimations revealed strong association of microglial expression patterns with Aß and tau's synergistic impact on neuronal activity (q < 0.001). These results further support the central role of the immune system and neuroinflammatory pathways in AD pathogenesis. Neuronal dysregulation by AD pathologies also associated with neurotypical synaptic and developmental processes. In addition, we identified drug candidates from the vast LINCS library to halt or reduce the observed Aß-tau effects on neuronal activity. Top-ranked pharmacological interventions target inflammatory, cancer and cardiovascular pathways, including specific medications undergoing clinical evaluation in AD. Our findings, based on the examination of molecular-pathological-functional interactions in humans, may accelerate the process of bringing effective therapies into clinical practice.

Neurofilaments and progranulin are related to atrophy in frontotemporal lobar degeneration - A transdiagnostic study cross-validating atrophy and fluid biomarkers.

INTRODUCTION: Frontotemporal lobar degeneration (FTLD) encompasses behavioral variant frontotemporal dementia (bvFTD), progressive supranuclear palsy, corticobasal syndrome/degeneration, and primary progressive aphasias (PPAs). We cross-validated fluid biomarkers and neuroimaging. METHODS: Seven fluid biomarkers from cerebrospinal fluid and serum were related to atrophy in 428 participants including these FTLD subtypes, logopenic variant PPA (lvPPA), Alzheimer's disease (AD), and healthy subjects. Atrophy was assessed by structural magnetic resonance imaging and atlas-based volumetry. RESULTS: FTLD subtypes, lvPPA, and AD showed specific profiles for neurofilament light chain, phosphorylated heavy chain, tau, phospho-tau, amyloid beta1-42 from serum/cerebrospinal fluid, and brain atrophy. Neurofilaments related to regional atrophy in bvFTD, whereas progranulin was associated with atrophy in semantic variant PPA. Ubiquitin showed no effects. DISCUSSION: Results specify biomarker and atrophy patterns in FTLD and AD supporting differential diagnosis. They identify neurofilaments and progranulin in interaction with structural imaging as promising candidates for monitoring disease progression and therapy. HIGHLIGHTS: Study cross-validated neuroimaging and fluid biomarkers in dementia. Five kinds of frontotemporal lobar degeneration and two variants of Alzheimer's disease. Study identifies disease-specific fluid biomarker and atrophy profiles. Fluid biomarkers and atrophy interact in a disease-specific way. Neurofilaments and progranulin are proposed as biomarkers for diagnosis and therapy.

Prognostic Power? Do the Plasma Biomarkers, Neurofilament Light and Phospho-Tau 181, Improve Prediction of Progression to Alzheimer's Disease Using a Machine Learning Approach in the ADNI Cohort?

With the advent of therapeutics with potential to slow Alzheimer's disease progression the necessity of understanding the diagnostic value of plasma biomarkers is critical, not only for understanding the etiology and progression of Alzheimer's disease, but also for access and response to potentially disease modifying therapeutic agents. Multiple studies are currently assessing the sensitivity and specificity of plasma biomarkers in large cohorts such as the Alzheimer's Disease Neuroimaging Initiative. This study uses machine learning to predict the progression from mild cognitive impairment using plasma biomarkers in conjunction with well-established cerebrospinal fluid and imaging biomarkers of disease progression.

Prenylated Flavanone Isolated from Dalea Species as a Potential Multitarget-Neuroprotector in an In Vitro Alzheimer's Disease Mice Model.

Alzheimer's disease (AD) involves a neurodegenerative process that has not yet been prevented, reversed, or stopped. Continuing with the search for natural pharmacological treatments, flavonoids are a family of compounds with proven neuroprotective effects and multi-targeting behavior. The American genus Dalea L. (Fabaceae) is an important source of bioactive flavonoids. In this opportunity, we tested the neuroprotective potential of three prenylated flavanones isolated from Dalea species in a new in vitro pre-clinical AD model previously developed by us. Our approach consisted in exposing neural cells to conditioned media (3xTg-AD ACM) from neurotoxic astrocytes derived from hippocampi and cortices of old 3xTg-AD mice, mimicking a local neurodegenerative microenvironment. Flavanone 1 and 3 showed a neuroprotective effect against 3xTg-AD ACM, being 1 more active than 3. The structural requirements to afford neuroprotective activity in this model are a 5'-dimethylallyl and 4'-hydroxy at the B ring. In order to search the mechanistic performance of the most active flavanone, we focus on the flavonoid-mediated regulation of GSK-3ß-mediated tau phosphorylation previously reported. Flavanone 1 treatment decreased the rise of hyperphosphorylated tau protein neuronal levels induced after 3xTg-AD ACM exposure and inhibited the activity of GSK-3ß. Finally, direct exposure of these neurotoxic 3xTg-AD astrocytes to flavanone 1 resulted in toxicity to these cells and reduced the neurotoxicity of 3xTg-AD ACM as well. Our results allow us to present compound 1 as a natural prenylated flavanone that could be used as a precursor to development and design of future drug therapies for AD.

Meta-Analysis in Transgenic Alzheimer's Disease Mouse Models Reveals Opposite Brain Network Effects of Amyloid-ß and Phosphorylated Tau Proteins.

Background: Cognitive deficits observed in Alzheimer's disease (AD) patients have been correlated with altered hippocampal activity. Although the mechanism remains under extensive study, neurofibrillary tangles and amyloid plaques have been proposed as responsible for brain activity alterations. Aiming to unveil the mechanism, researchers have developed several transgenic models of AD. Nevertheless, the variability in hippocampal oscillatory alterations found in different genetic backgrounds and ages remains unclear. Objective: To assess the oscillatory alterations in relation to animal developmental age and protein inclusion, amyloid-ß (Aß) load, and abnormally phosphorylated tau (pTau), we reviewed and analyzed the published data on peak power, frequency, and quantification of theta-gamma cross-frequency coupling (modulation index values). Methods: To ensure that the search was as current as possible, a systematic review was conducted to locate and abstract all studies published from January 2000 to February 2023 that involved in vivo hippocampal local field potential recording in transgenic mouse models of AD. Results: The presence of Aß was associated with electrophysiological alterations that are mainly reflected in power increases, frequency decreases, and lower modulation index values. Concomitantly, pTau accumulation was associated with electrophysiological alterations that are mainly reflected in power decreases, frequency decreases, and no significant alterations in modulation index values. Conclusions: In this study, we showed that electrophysiological parameters are altered from prodromal stages to the late stages of pathology. Thus, we found that Aß deposition is associated with brain network hyperexcitability, whereas pTau deposition mainly leads to brain network hypoexcitability in transgenic models.

Optimized Pre-analytical Handling Protocol for Blood-Based Biomarkers of Alzheimer's Disease.

The therapeutic management of patients with Alzheimer's disease (AD) has been hindered by poor diagnostic accuracy. As such, there is an unmet clinical need for tools that can detect and diagnose the disease in its early stages. Compared with cerebrospinal fluid (CSF)-based biomarkers or positron emission tomography (PET), the use of reliable blood-based biomarkers could offer an accessible and minimally invasive method of streamlining diagnosis in the clinical setting. However, the influence of pre-analytical processing and sample handling parameters on the accurate measurement of protein biomarkers is well established, especially for AD CSF-based biomarkers. In this chapter, we provide recommendations for an optimal sample handling protocol for the analysis of blood-based biomarkers specifically for amyloid pathology in AD.

The impact of kidney function on plasma neurofilament light and phospho-tau 181 in a community-based cohort: the Shanghai Aging Study.

BACKGROUND: The blood-based biomarkers are approaching the clinical practice of Alzheimer's disease (AD). Chronic kidney disease (CKD) has a potential confounding effect on peripheral protein levels. It is essential to characterize the impact of renal function on AD markers. METHODS: Plasma phospho-tau181 (P-tau181), and neurofilament light (NfL) were assayed via the Simoa HD-X platform in 1189 dementia-free participants from the Shanghai Aging Study (SAS). The estimated glomerular filter rate (eGFR) was calculated. The association between renal function and blood NfL, P-tau181 was analyzed. An analysis of interactions between various demographic and comorbid factors and eGFR was conducted. RESULTS: The eGFR levels were negatively associated with plasma concentrations of NfL and P-tau181 (B = - 0.19, 95% CI - 0.224 to - 0.156, P < 0.001; B = - 0.009, 95% CI - 0.013 to -0.005, P < 0.001, respectively). After adjusting for demographic characteristics and comorbid diseases, eGFR remained significantly correlated with plasma NfL (B = - 0.010, 95% CI - 0.133 to - 0.068, P < 0.001), but not with P-tau181 (B = - 0.003, 95% CI - 0.007 to 0.001, P = 0.194). A significant interaction between age and eGFR was found for plasma NfL (Pinteraction < 0.001). In participants ≥ 70 years and with eGFR < 60 ml/min/1.73 m2, the correlation between eGFR and plasma NfL was significantly remarkable (B = - 0.790, 95% CI - 1.026 to - 0,554, P < 0.001). CONCLUSIONS: Considering renal function and age is crucial when interpreting AD biomarkers in the general aging population.

Phospho-Tau 181 quantification method for Alzheimer's disease based on an array 2D biosensor combined with surface plasmon resonance imaging.

Alzheimer's disease is among the neurodegenerative diseases for which there is a lack of rapid, effective, and non-invasive diagnostic methods. The development of a phospho-Tau 181 assay biosensor is therefore a response to the need for methods to diagnose AD. The present work was aimed at developing a fast, selective, and repeatable method for the quantitative determination of phospho-Tau 181, which could be used even during routine blood tests. Our method is a form of what is called liquid biopsy. The developed method underwent validation, as a result of which its analytical parameters were determined. An LOQ of 3.35 pg mL-1 was obtained, confirming the possibility of trace analysis of phospho-Tau 181 in human plasma. Relative percentage error values below 15 % and CVs in the range 1.47-7.09 % attest to the high accuracy and precision of the presented method. Also, the sample matrix was not found to significantly affect the results obtained for phospho-Tau 181 concentrations. The new SPRi biosensor provides reproducible measurements of the analyte under study (CV = 3.18-4.26 %). Although the method requires absolute adherence to the recommendations of the analytical procedure protocol, it achieves high selectivity and provides 90 % certainty of the correctness of the diagnosis based on measurements of phospho-Tau 181 concentration.

Potential Utility of Plasma Biomarker Panels in Differential Diagnosis of Alzheimer's Disease.

Blood tests are in need, in the clinical diagnosis of Alzheimer's disease (AD) as minimally invasive and less expensive alternatives to cerebrospinal fluid and neuroimaging methods. On these lines, single molecule array (Simoa) analysis of amyloid-ß (Aß42), total tau (t-tau), phospho-tau (p-tau 181), and neurofilament L (NfL) in the plasma samples of AD subjects, healthy controls (HC), and non-AD subjects was conducted. Findings from this study suggest that a panel of multiple plasma biomarkers (NfL, Aß42, t-tau, and p-tau 181) combined with the clinical assessments could support differential diagnosis of AD and other dementias from healthy controls.

Alzheimer's Disease CSF Biomarkers as Possible Indicators of Tap-Test Response in Idiopathic Normal Pressure Hydrocephalus.

The aim of the present study is the evaluation of established Alzheimer's disease (AD) cerebrospinal fluid (CSF) biomarkers in patients with idiopathic normal-pressure hydrocephalus (iNPH), both individually and as a total profile, and the investigation of their use as potential predictors of Tap-test responsiveness. Fifty-three patients with iNPH participated in the study. Aß42, Aß40, total Tau and phospho-Tau proteins were measured in duplicate with double-sandwich ELISA assays. Clinical evaluation involved a 10 m timed walk test before an evacuative lumbar puncture (LP) and every 24 h for three consecutive days afterwards. Neuropsychological assessment involved a mini-mental state examination, frontal assessment battery, 5-word test and CLOX drawing test 1 and 2, which were also performed before and 48 h after LP. Response in the Tap-test was defined as a 20% improvement in gait and/or a 10% improvement in neuropsychological tests. The Aß42/Aß40 ratio was found to be significantly higher in Tap-test responders than non-responders. Total Tau and phospho-Tau CSF levels also differed significantly between these two groups, with Tap-test responders presenting with lower levels compared to non-responders. Regarding the AD CSF biomarker profile (decreased amyloid and increased Tau proteins levels), patients with a non-AD profile were more likely to have a positive response in the Tap-test than patients with an AD profile.

Deleterious and protective effects of epothilone-D alone and in the context of amyloid ß- and tau-induced alterations.

Amyloid-ß (Aß) and hyperphosphorylated tau (P-tau) are Alzheimer's disease (AD) biomarkers that interact in a complex manner to induce most of the cognitive and brain alterations observed in this disease. Since the neuronal cytoskeleton is a common downstream pathological target of tau and Aß, which mostly lead to augmented microtubule instability, the administration of microtubule stabilizing agents (MSAs) can protect against their pathological actions. However, the effectiveness of MSAs is still uncertain due to their state-dependent negative effects; thus, evaluating their specific actions in different pathological or physiological conditions is required. We evaluated whether epothilone-D (Epo-D), a clinically used MSA, rescues from the functional and behavioral alterations produced by intracerebroventricular injection of Aß, the presence of P-tau, or their combination in rTg4510 mice. We also explored the side effects of Epo-D. To do so, we evaluated hippocampal-dependent spatial memory with the Hebb-Williams maze, hippocampal CA1 integrity and the intrinsic and synaptic properties of CA1 pyramidal neurons with the patch-clamp technique. Aß and P-tau mildly impaired memory retrieval, but produced contrasting effects on intrinsic excitability. When Aß and P-tau were combined, the alterations in excitability and spatial reversal learning (i.e., cognitive flexibility) were exacerbated. Interestingly, Epo-D prevented most of the impairments induced Aß and P-tau alone and combined. However, Epo-D also exhibited some side effects depending on the prevailing pathological or physiological condition, which should be considered in future preclinical and translational studies. Although we did not perform extensive histopathological evaluations or measured microtubule stability, our findings show that MSAs can rescue the consequences of AD-like conditions but otherwise be harmful if administered at a prodromal stage of the disease.

Severe acute respiratory syndrome coronavirus 2 infection leads to Tau pathological signature in neurons.

COVID-19 has represented an issue for global health since its outbreak in March 2020. It is now evident that the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection results in a wide range of long-term neurological symptoms and is worryingly associated with the aggravation of Alzheimer's disease. Little is known about the molecular basis of these manifestations. Here, several strain variants were used to infect SH-SY5Y neuroblastoma cells and K18-hACE C57BL/6J mice. The Tau phosphorylation profile and aggregation propensity upon infection were investigated on cellular extracts, subcellular fractions, and brain tissue. The viral proteins spike, nucleocapsid, and membrane were overexpressed in SH-SY5Y cells, and the direct interaction and effect on Tau phosphorylation were checked using immunoblot experiments. Upon infection, Tau is phosphorylated at several pathological epitopes associated with Alzheimer's disease and other tauopathies. Moreover, this event increases Tau's propensity to form insoluble aggregates and alters its subcellular localization. Our data support the hypothesis that SARS-CoV-2 infection in the central nervous system triggers downstream effects altering Tau function, eventually leading to the impairment of neuronal function.

PPARɑ Ligand Caudatin Improves Cognitive Functions and Mitigates Alzheimer's Disease Defects By Inducing Autophagy in Mice Models.

The autophagy-lysosomal pathway (ALP) is a major cellular machinery involved in the clearance of aggregated proteins in Alzheimer disease (AD). However, ALP is dramatically impaired during AD pathogenesis via accumulation of toxic amyloid beta (Aß) and phosphorylated-Tau (phospho-Tau) proteins in the brain. Therefore, activation of ALP may prevent the increased production of Aß and phospho-Tau in AD. Peroxisome proliferator-activated receptor alpha (PPARα), a transcription factor that can activate autophagy, and transcriptionally regulate transcription factor EB (TFEB) which is a key regulator of ALP. This suggests that targeting PPARα, to reduce ALP impairment, could be a viable strategy for AD therapy. In this study, we investigated the anti-AD activity of Caudatin, an active constituent of Cynanchum otophyllum (a traditional Chinese medicinal herb, Qing Yang Shen; QYS). We found that Caudatin can bind to PPARα as a ligand and augment the expression of ALP in microglial cells and in the brain of 3XTg-AD mice model. Moreover, Caudatin could activate PPARα and transcriptionally regulates TFEB-augmented lysosomal degradation of Aß and phosphor-Tau aggregates in AD cell models. Oral administration of Caudatin decreased AD pathogenesis and ameliorated the cognitive dysfunction in 3XTg-AD mouse model. Conclusively, Caudatin can be a potential AD therapeutic agent via activation of PPARα-dependent ALP.

Early ß-amyloid accumulation in the brain is associated with peripheral T cell alterations.

INTRODUCTION: Fast and minimally invasive approaches for early diagnosis of Alzheimer's disease (AD) are highly anticipated. Evidence of adaptive immune cells responding to cerebral ß-amyloidosis has raised the question of whether immune markers could be used as proxies for ß-amyloid accumulation in the brain. METHODS: Here, we apply multidimensional mass-cytometry combined with unbiased machine-learning techniques to immunophenotype peripheral blood mononuclear cells from a total of 251 participants in cross-sectional and longitudinal studies. RESULTS: We show that increases in antigen-experienced adaptive immune cells in the blood, particularly CD45RA-reactivated T effector memory (TEMRA) cells, are associated with early accumulation of brain ß-amyloid and with changes in plasma AD biomarkers in still cognitively healthy subjects. DISCUSSION: Our results suggest that preclinical AD pathology is linked to systemic alterations of the adaptive immune system. These immunophenotype changes may help identify and develop novel diagnostic tools for early AD assessment and better understand clinical outcomes.

Alzheimer's disease biomarkers in cerebrospinal fluid are stable with the Elecsys immunoassay to most pre-analytical influencing factors except freezing at -80 °C.

BACKGROUND: Alzheimer´s disease is considered a neurodegenerative disease and is diagnosed by exclusion, while the detection of specific cerebrospinal fluid (CSF) biomarkers, namely amyloid-beta (Aß) peptides Aß1-42 (Aß42), phospho-tau (181P; P-tau), and total-tau (T-tau), has been shown to improve diagnostic accuracy. Recently, a new generation of sample tubes (Sarstedt false-bottom tubes) for the Elecsys CSF immunoassay for the determination of Alzheimer´s disease biomarkers in CSF was introduced, promising better measurability. However, the pre-analytic influencing factors have not yet been sufficiently investigated. METHODS: In 29 patients without Alzheimer's disease diagnosis, CSF concentrations of Aß42, P-tau and T-tau were examined in native CSF and after different influencing interventions using the Elecsys immunoassay test method. The following influencing factors were analyzed: contamination with blood (10,000 and 20,000 erythrocytes/µl CSF), 14-day storage at 4 °C, blood contamination of CSF and 14-day storage at 4 °C, 14-day freezing at -80 °C in Sarstedt tubes or glass vials, 3-month intermediate storage at -80 °C in glass vials. RESULTS: Both storage at -80 °C for 14 days in Sarstedt false-bottom tubes and in glass vials and storage at -80 °C for 3 months in glass vials resulted in significant decreases in Aß42 (13% after 14 days in Sarstedt and 22% in glass vials, 42% after 3 months in glass vials), P-tau (9% after 14 days in Sarstedt and 13% in glass vials, 12% after 3 months in glass vials) and T-tau (12% after 14 days in Sarstedt and 19% in glass vials, 20% after 3 months in glass vials) concentrations in CSF. No significant differences were found for the other pre-analytical influencing factors. CONCLUSIONS: Measurements of the concentrations of Aß42, P-tau, and T-tau in CSF with use of the Elecsys immunoassay are robust to the pre-analytical influencing factors of blood contamination and duration of storage. Freezing at -80 °C results in significant reduction of biomarker concentrations regardless of the storage tube and must be considered in retrospective analysis.

Yeast biopanning against site-specific phosphorylations in tau.

The detection of site-specific phosphorylation in the microtubule-associated protein tau is emerging as a means to diagnose and monitor the progression of Alzheimer's Disease and other neurodegenerative diseases. However, there is a lack of phospho-specific monoclonal antibodies and limited validation of their binding specificity. Here, we report a novel approach using yeast biopanning against synthetic peptides containing site-specific phosphorylations. Using yeast cells displaying a previously validated phospho-tau (p-tau) single-chain variable region fragment (scFv), we show selective yeast cell binding based on single amino acid phosphorylation on the antigen. We identify conditions that allow phospho-specific biopanning using scFvs with a wide range of affinities (KD = 0.2 to 60 nM). Finally, we demonstrate the capability of screening large libraries by performing biopanning in 6-well plates. These results show that biopanning can effectively select yeast cells based on phospho-site specific antibody binding, opening doors for the facile identification of high-quality monoclonal antibodies.

Alzheimer's Disease-Related Phospho-Tau181 Signals Are Localized to Demyelinated Axons of Parvalbumin-Positive GABAergic Interneurons in an App Knock-In Mouse Model of Amyloid-ß Pathology.

BACKGROUND: The tau protein phosphorylated at Thr181 (p-tau181) in cerebrospinal fluid and blood is a sensitive biomarker for Alzheimer's disease (AD). Increased p-tau181 levels correlate well with amyloid-ß (Aß) pathology and precede neurofibrillary tangle formation in the early stage of AD; however, the relationship between p-tau181 and Aß-mediated pathology is less well understood. We recently reported that p-tau181 represents axonal abnormalities in mice with Aß pathology (AppNLGF). However, from which neuronal subtype(s) these p-tau181-positive axons originate remains elusive. OBJECTIVE: The main purpose of this study is to differentiate neuronal subtype(s) and elucidate damage associated with p-tau181-positive axons by immunohistochemical analysis of AppNLGF mice brains. METHODS: Colocalization between p-tau181 and (1) unmyelinated axons positive for vesicular acetylcholine transporter or norepinephrine transporter and (2) myelinated axons positive for vesicular glutamate transporter, vesicular GABA transporter, or parvalbumin in the brains of 24-month-old AppNLGF and control mice without Aß pathology were analyzed. The density of these axons was also compared. RESULTS: Unmyelinated axons of cholinergic or noradrenergic neurons did not overlap with p-tau181. By contrast, p-tau181 signals colocalized with myelinated axons of parvalbumin-positive GABAergic interneurons but not of glutamatergic neurons. Interestingly, the density of unmyelinated axons was significantly decreased in AppNLGF mice, whereas that of glutamatergic, GABAergic, or p-tau181-positive axons was less affected. Instead, myelin sheaths surrounding p-tau181-positive axons were significantly reduced in AppNLGF mice. CONCLUSION: This study demonstrates that p-tau181 signals colocalize with axons of parvalbumin-positive GABAergic interneurons with disrupted myelin sheaths in the brains of a mouse model of Aß pathology.

The Effects of a Blood-Brain Barrier Penetrating Erythropoietin in a Mouse Model of Tauopathy.

Erythropoietin (EPO), a hematopoietic neurotrophin, is a potential therapeutic for Alzheimer's disease (AD) but has limited blood-brain barrier (BBB) permeability. EPO fused to a chimeric transferrin receptor monoclonal antibody (cTfRMAb) enters the brain via TfR-mediated transcytosis across the BBB. We previously showed that cTfRMAb-EPO is protective in a mouse model of amyloidosis, but its effects on tauopathy are not known. Given that amyloid and tau pathology are characteristics of AD, the effects of cTfRMAb-EPO were studied in a tauopathy mouse model (PS19). Six-month-old PS19 mice were injected intraperitoneally with either saline (PS19-Saline; n = 9) or cTfRMAb-EPO (PS19-cTfRMAb-EPO, 10 mg/kg; n = 10); every two or three days on alternate weeks for 8 weeks. Age-matched, saline-treated, wildtype littermates (WT-Saline; n = 12) were injected using the same protocol. After 8 weeks, locomotion, hyperactivity, and anxiety were assessed via the open-field test, and brains were harvested and sectioned. Cerebral cortex, hippocampus, amygdala, and entorhinal cortex sections were analyzed for phospho-tau (AT8) and microgliosis (Iba1). Hippocampal cellular density (H&E) was also assessed. PS19-Saline mice were hyperactive and less anxious compared to WT-Saline mice, and these behavioral phenotypes were significantly reduced in the PS19-cTfRMAb-EPO mice compared to the PS19-Saline mice. cTfRMAb-EPO significantly reduced AT8 load by ≥50% in all of the brain regions analyzed and microgliosis in the entorhinal cortex and amygdala compared to the PS19-Saline mice. Hippocampal pyramidal and granule cell layer density did not differ significantly between the PS19-cTfRMAb-EPO and PS19-Saline mice. This proof-of-concept study demonstrates the therapeutic effects of the BBB-penetrating cTfRMAb-EPO in PS19 mice.

Impact of pre-analytical sample handling factors on plasma biomarkers of Alzheimer's disease.

An unmet need exists for reliable plasma biomarkers of amyloid pathology, in the clinical laboratory setting, to streamline diagnosis of Alzheimer's disease (AD). For routine clinical use, a biomarker must provide robust and reliable results under pre-analytical sample handling conditions. We investigated the impact of different pre-analytical sample handling procedures on the levels of seven plasma biomarkers in development for potential routine use in AD. Using (1) fresh (never frozen) and (2) previously frozen plasma, we evaluated the effects of (A) storage time and temperature, (B) freeze/thaw (F/T) cycles, (C) anticoagulants, (D) tube transfer, and (E) plastic tube types. Blood samples were prospectively collected from patients with cognitive impairment undergoing investigation in a memory clinic. ß-amyloid 1-40 (Aß40), ß-amyloid 1-42 (Aß42), apolipoprotein E4, glial fibrillary acidic protein, neurofilament light chain, phosphorylated-tau (phospho-tau) 181, and phospho-tau-217 were measured using Elecsys® plasma prototype immunoassays. Recovery signals for each plasma biomarker and sample handling parameter were calculated. For all plasma biomarkers measured, pre-analytical effects were comparable between fresh (never frozen) and previously frozen samples. All plasma biomarkers tested were stable for ≤24 h at 4°C when stored as whole blood and ethylenediaminetetraacetic acid (EDTA) plasma. Recovery signals were acceptable for up to five tube transfers, or two F/T cycles, and in both polypropylene and low-density polyethylene tubes. For all plasma biomarkers except Aß42 and Aß40, analyte levels were largely comparable between EDTA, lithium heparin, and sodium citrate tubes. Aß42 and Aß40 were most sensitive to pre-analytical handling, and the effects could only be partially compensated by the Aß42/Aß40 ratio. We provide recommendations for an optimal sample handling protocol for analysis of plasma biomarkers for amyloid pathology AD, to improve the reproducibility of future studies on plasma biomarkers assays and for potential use in routine clinical practice.