Neuropathological assessment of the olfactory bulb and tract in individuals with COVID-19.

The majority of patients with Parkinson disease (PD) experience a loss in their sense of smell and accumulate insoluble α-synuclein aggregates in their olfactory bulbs (OB). Subjects affected by a SARS-CoV-2-linked illness (COVID-19) also frequently experience hyposmia. We previously postulated that microglial activation as well as α-synuclein and tau misprocessing can occur during host responses following microbial encounters. Using semiquantitative measurements of immunohistochemical signals, we examined OB and olfactory tract specimens collected serially at autopsies between 2020 and 2023. Deceased subjects comprised 50 adults, which included COVID19 + patients (n = 22), individuals with Lewy body disease (e.g., PD; dementia with Lewy bodies (n = 6)), Alzheimer disease (AD; n = 3), and other neurodegenerative disorders (e.g., progressive supranuclear palsy (n = 2); multisystem atrophy (n = 1)). Further, we included neurologically healthy controls (n = 9), and added subjects with an inflammation-rich brain disorder as neurological controls (NCO; n = 7). When probing for microglial and histiocytic reactivity in the anterior olfactory nuclei (AON) by anti-CD68 immunostaining, scores were consistently elevated in NCO and AD cases. In contrast, microglial signals on average were not significantly altered in COVID19 + patients relative to healthy controls, although anti-CD68 reactivity in their OB and tracts declined with progression in age. Mild-to-moderate increases in phospho-α-synuclein and phospho-tau signals were detected in the AON of tauopathy- and synucleinopathy-afflicted brains, respectively, consistent with mixed pathology, as described by others. Lastly, when both sides were available for comparison in our case series, we saw no asymmetry in the degree of pathology of the left versus right OB and tracts. We concluded from our autopsy series that after a fatal course of COVID-19, microscopic changes in the rostral, intracranial portion of the olfactory circuitry -when present- reflected neurodegenerative processes seen elsewhere in the brain. In general, microglial reactivity correlated best with the degree of Alzheimer's-linked tauopathy and declined with progression of age in COVID19 + patients.

In Vivo Assessment of Astrocyte Reactivity in Patients with Progressive Supranuclear Palsy.

OBJECTIVE: Although astrocytic pathology is a pathological hallmark of progressive supranuclear palsy (PSP), its pathophysiological role remains unclear. This study aimed to assess astrocyte reactivity in vivo in patients with PSP. Furthermore, we investigated alterations in brain lactate levels and their relationship with astrocyte reactivity. METHODS: We included 30 patients with PSP-Richardson syndrome and 30 healthy controls; in patients, tau deposition was confirmed through 18F-florzolotau positron emission tomography. Myo-inositol, an astroglial marker, and lactate were quantified in the anterior cingulate cortex through magnetic resonance spectroscopy. We measured plasma biomarkers, including glial fibrillary acidic protein as another astrocytic marker. The anterior cingulate cortex was histologically assessed in postmortem samples of another 3 patients with PSP with comparable disease durations. RESULTS: The levels of myo-inositol and plasma glial fibrillary acidic protein were significantly higher in patients than those in healthy controls (p < 0.05); these increases were significantly associated with PSP rating scale and cognitive function scores (p < 0.05). The lactate level was high in patients, and correlated significantly with high myo-inositol levels. Histological analysis of the anterior cingulate cortex in patients revealed reactive astrocytes, despite mild tau deposition, and no marked synaptic loss. INTERPRETATION: We discovered high levels of astrocyte biomarkers in patients with PSP, suggesting astrocyte reactivity. The association between myo-inositol and lactate levels suggests a link between reactive astrocytes and brain energy metabolism changes. Our results indicate that astrocyte reactivity in the anterior cingulate cortex precedes pronounced tau pathology and neurodegenerative processes in that region, and affects brain function in PSP. ANN NEUROL 2024;96:247-261.

Kinetic Ensemble of Tau Protein through the Markov State Model and Deep Learning Analysis.

The ordered assembly of Tau protein into filaments characterizes Alzheimer's and other neurodegenerative diseases, and thus, stabilization of Tau protein is a promising avenue for tauopathies therapy. To dissect the underlying aggregation mechanisms on Tau, we employ a set of molecular simulations and the Markov state model to determine the kinetics of ensemble of K18. K18 is the microtubule-binding domain of Tau protein and plays a vital role in the microtubule assembly, recycling processes, and amyloid fibril formation. Here, we efficiently explore the conformation of K18 with about 150 µs lifetimes in silico. Our results observe that all four repeat regions (R1-R4) are very dynamic, featuring frequent conformational conversion and lacking stable conformations, and the R2 region is more flexible than the R1, R3, and R4 regions. Additionally, it is worth noting that residues 300-310 in R2-R3 and residues 319-336 in R3 tend to form sheet structures, indicating that K18 has a broader functional role than individual repeat monomers. Finally, the simulations combined with Markov state models and deep learning reveal 5 key conformational states along the transition pathway and provide the information on the microsecond time scale interstate transition rates. Overall, this study offers significant insights into the molecular mechanism of Tau pathological aggregation and develops novel strategies for both securing tauopathies and advancing drug discovery.

Sildenafil as a Candidate Drug for Alzheimer's Disease: Real-World Patient Data Observation and Mechanistic Observations from Patient-Induced Pluripotent Stem Cell-Derived Neurons.

Background: Alzheimer's disease (AD) is a chronic neurodegenerative disease needing effective therapeutics urgently. Sildenafil, one of the approved phosphodiesterase-5 inhibitors, has been implicated as having potential effect in AD. Objective: To investigate the potential therapeutic benefit of sildenafil on AD. Methods: We performed real-world patient data analysis using the MarketScan® Medicare Supplemental and the Clinformatics® databases. We conducted propensity score-stratified analyses after adjusting confounding factors (i.e., sex, age, race, and comorbidities). We used both familial and sporadic AD patient induced pluripotent stem cells (iPSC) derived neurons to evaluate the sildenafil's mechanism-of-action. Results: We showed that sildenafil usage is associated with reduced likelihood of AD across four new drug compactor cohorts, including bumetanide, furosemide, spironolactone, and nifedipine. For instance, sildenafil usage is associated with a 54% reduced incidence of AD in MarketScan® (hazard ratio [HR] = 0.46, 95% CI 0.32- 0.66) and a 30% reduced prevalence of AD in Clinformatics® (HR = 0.70, 95% CI 0.49- 1.00) compared to spironolactone. We found that sildenafil treatment reduced tau hyperphosphorylation (pTau181 and pTau205) in a dose-dependent manner in both familial and sporadic AD patient iPSC-derived neurons. RNA-sequencing data analysis of sildenafil-treated AD patient iPSC-derived neurons reveals that sildenafil specifically target AD related genes and pathobiological pathways, mechanistically supporting the beneficial effect of sildenafil in AD. Conclusions: These real-world patient data validation and mechanistic observations from patient iPSC-derived neurons further suggested that sildenafil is a potential repurposable drug for AD. Yet, randomized clinical trials are warranted to validate the causal treatment effects of sildenafil in AD.

A comparison of visual assessment and semi-quantification for the diagnostic and prognostic use of [18F]flortaucipir PET in a memory clinic cohort.

PURPOSE: [18F]Flortaucipir PET is a powerful diagnostic and prognostic tool for Alzheimer's disease (AD). Tau status definition is mainly based in the literature on semi-quantitative measures while in clinical settings visual assessment is usually preferred. We compared visual assessment with established semi-quantitative measures to classify subjects and predict the risk of cognitive decline in a memory clinic population. METHODS: We included 245 individuals from the Geneva Memory Clinic who underwent [18F]flortaucipir PET. Amyloid status was available for 207 individuals and clinical follow-up for 135. All scans were blindly evaluated by three independent raters who visually classified the scans according to Braak stages. Standardized uptake value ratio (SUVR) values were obtained from a global meta-ROI to define tau positivity, and the Simplified Temporo-Occipital Classification (STOC) was applied to obtain semi-quantitatively tau stages. The agreement between measures was tested using Cohen's kappa (k). ROC analysis and linear mixed-effects models were applied to test the diagnostic and prognostic values of tau status and stages obtained with the visual and semi-quantitative approaches. RESULTS: We found good inter-rater reliability in the visual interpretation of tau Braak stages, independently from the rater's expertise (k>0.68, p<0.01). A good agreement was equally found between visual and SUVR-based classifications for tau status (k=0.67, p<0.01). All tau-assessment modalities significantly discriminated amyloid-positive MCI and demented subjects from others (AUC>0.80) and amyloid-positive from negative subjects (AUC>0.85). Linear mixed-effect models showed that tau-positive individuals presented a significantly faster cognitive decline than the tau-negative group (p<0.01), independently from the classification method. CONCLUSION: Our results show that visual assessment is reliable for defining tau status and stages in a memory clinic population. The high inter-rater reliability, the substantial agreement, and the similar diagnostic and prognostic performance of visual rating and semi-quantitative methods demonstrate that [18F]flortaucipir PET can be robustly assessed visually in clinical practice.

Identification of Aggregation Mechanism of Acetylated PHF6* and PHF6 Tau Peptides Based on Molecular Dynamics Simulations and Markov State Modeling.

The microtubule-associated protein tau (MAPT) has a critical role in the development and preservation of the nervous system. However, tau's dysfunction and accumulation in the human brain can lead to several neurodegenerative diseases, such as Alzheimer's disease, Down's syndrome, and frontotemporal dementia. The microtubule binding (MTB) domain plays a significant, important role in determining the tau's pathophysiology, as the core of paired helical filaments PHF6* (275VQIINK280) and PHF6 (306VQIVYK311) of R2 and R3 repeat units, respectively, are formed in this region, which promotes tau aggregation. Post-translational modifications, and in particular lysine acetylation at K280 of PHF6* and K311 of PHF6, have been previously established to promote tau misfolding and aggregation. However, the exact aggregation mechanism is not known. In this study, we established an atomic-level nucleation-extension mechanism of the separated aggregation of acetylated PHF6* and PHF6 hexapeptides, respectively, of tau. We show that the acetylation of the lysine residues promotes the formation of ß-sheet enriched high-ordered oligomers. The Markov state model analysis of ac-PHF6* and ac-PHF6 aggregation revealed the formation of an antiparallel dimer nucleus which could be extended from both sides in a parallel manner to form mixed-oriented and high-ordered oligomers. Our study describes the detailed mechanism for acetylation-driven tau aggregation, which provides valuable insights into the effect of post-translation modification in altering the pathophysiology of tau hexapeptides.

GTP1 metabolic stability assessment: A study of the tau PET tracer [18F]GTP1.

Tau PET imaging using the tau specific PET tracer [18F]GTP1 has been and is part of therapeutic trials in Alzheimer's disease to monitor the accumulation of tau aggregates in the brain. Herein, we examined the metabolic processes of GTP1 and assessed the influence of smoking on its metabolism through in vitro assays. The tracer metabolic profile was assessed by incubating GTP1 with human liver microsomes (HLM) and human hepatocytes. Since smoking strongly stimulates the CYP1A2 enzyme activity, we incubated GTP1 with recombinant CYP1A2 to evaluate the role of the enzyme in tracer metabolism. It was found that GTP1 could form up to eleven oxidative metabolites with higher polarity than the parent. Only a small amount (2.6 % at 60 min) of a defluorinated metabolite was detected in HLM and human hepatocytes incubations highlighting the stability of GTP1 with respect to enzymatic defluorination. Moreover, the major GTP1 metabolites were not the product of CYP1A2 activity suggesting that smoking may not impact in vivo tracer metabolism and subsequently GTP1 brain kinetics.

The interaction of global small vessel disease burden and Alzheimer's disease pathologies do not change the independent association of amyloid-beta with hippocampal volume: A longitudinal study on mild cognitive impairment subjects.

The purpose of this study was to investigate whether the co-existence of global small vessel disease (SVD) burdens and Alzheimer's disease (AD) pathologies change hippocampal volume (HV) and cognitive function of mild cognitive impairment (MCI) subjects. We obtained MRI images, cerebrospinal fluid biomarkers (Aß1-42 and p-tau), and neuropsychological tests of 310 MCI subjects from ADNI. The global SVD score was assessed. We used linear regression and linear mixing effect to analyze the effects of global SVD burdens, AD pathologies, and their interactions (SVD*AD) on baseline and longitudinal HV and cognition respectively. We used simple mediation effect to analyze the influencing pathways. After adjusting for global SVD and SVD*AD, Aß remained independently correlated with baseline and longitudinal HV (std ß = 0.294, p = .007; std ß = 0.292, p < .001), indicating that global SVD did not affect the correlation between Aß and HV. Global SVD score was correlated with longitudinal but not baseline HV (std ß = 0.470, p = .050), suggesting that global SVD may be more representative of long-term permanent impairment. Global SVD, AD pathologies, and SVD*AD were independently correlated with baseline and longitudinal cognitions, in which the association of Aß (B = 0.005, 95% CI: 0.005; 0.024) and p-tau (B = -0.002, 95% CI: -0.004; -0.000) with cognition were mediated by HV, suggesting that HV is more likely to explain the progression caused by AD pathology than SVD. The co-existence of global SVD and AD pathologies did not affect the individual association of Aß on HV; HV played a more important role in the influence of AD pathology on cognition than in SVD.

Assessment of tau phosphorylation and ß-amyloid pathology in human drug-resistant epilepsy.

OBJECTIVE: Epilepsy can be comorbid with cognitive impairments. Recent evidence suggests the possibility that cognitive decline in epilepsy may be associated with mechanisms typical of Alzheimer's disease (AD). Neuropathological hallmarks of AD have been found in brain biopsies surgically resected from patients with drug-resistant epilepsies. These include hyperphosphorylation of the tau protein (p-tau) that aggregates into neuropil threads (NT) or neurofibrillary tangles (NFT), as well as the presence of ß-amyloid (Aß) deposits. While recent studies agree on these AD neuropathological findings in epilepsy, some contrast in their correlation to cognitive decline. Thus, to further address this question we determined the abundance of p-tau and Aß proteins along with their association with cognitive function in 12 cases of refractory epilepsy. METHODS: Cortical biopsies surgically extracted from the temporal lobes of patients with refractory epilepsy were processed for immunohistology and enzyme-linked immunoassays to assess distribution and levels, respectively, of p-tau (Antibodies: Ser202/Thr205; Thr205; Thr181) and Aß proteins. In parallel, we measured the activation of mechanistic target of rapamycin (mTOR) via p-S6 (Antibodies: Ser240/244; Ser235/236). Pearson correlation coefficient analysis determined associations between these proteins and neurophysiological scores for full-scale intelligence quotient (FSIQ). RESULTS: We found a robust presence of p-tau (Ser202/Thr205)-related NT and NFT pathology, as well as Aß deposits, and p-S6 (Ser240/244; Ser235/236) in the epilepsy biopsies. We found no significant correlations between p-tau (Thr205; Thr181), Aß, or mTOR markers with FSIQ scores, although some correlation coefficients were modest to strong. SIGNIFICANCE: These findings strongly support the existence of hyperphosphorylated tau protein and Aß deposits in patients with human refractory epilepsy. However, their relation to cognitive decline is still unclear and requires further investigation.

Assessment of Lab4P Probiotic Effects on Cognition in 3xTg-AD Alzheimer's Disease Model Mice and the SH-SY5Y Neuronal Cell Line.

Aging and metabolic syndrome are associated with neurodegenerative pathologies including Alzheimer's disease (AD) and there is growing interest in the prophylactic potential of probiotic bacteria in this area. In this study, we assessed the neuroprotective potential of the Lab4P probiotic consortium in both age and metabolically challenged 3xTg-AD mice and in human SH-SY5Y cell culture models of neurodegeneration. In mice, supplementation prevented disease-associated deteriorations in novel object recognition, hippocampal neurone spine density (particularly thin spines) and mRNA expression in hippocampal tissue implying an anti-inflammatory impact of the probiotic, more notably in the metabolically challenged setting. In differentiated human SH-SY5Y neurones challenged with ß-Amyloid, probiotic metabolites elicited a neuroprotective capability. Taken together, the results highlight Lab4P as a potential neuroprotective agent and provide compelling support for additional studies in animal models of other neurodegenerative conditions and human studies.

Rodent Model Preclinical Assessment of PEGylated Block Copolymer Targeting Cognition and Oxidative Stress Insults of Alzheimer's Disease.

Misfolded peptide amyloid beta (Aß42), neurofibrillary tangles of hyper-phosphorylated tau, oxidative damage to the brain, and neuroinflammation are distinguished determinants of Alzheimer's disease (AD) responsible for disease progression. This multifaceted neurodegenerative disease is challenging to cure under a single treatment regime until the key disease determinants are traced for their sequential occurrence in disease progression. In an early report, a novel side-chain tripeptide containing PEGylated block copolymer has been tested thoroughly in vitro and in silico for the early inhibition of Aß42 aggregation as well as degradation of preformed Aß42 fibril deposits. The present study demonstrates a preclinical assessment of the PEGylated block copolymer in colchicine-induced AD-mimicking rodent model. The colchicine-induced Wistar rats receiving an intranasal delivery of the block copolymer at a daily dosage of 100 µg/kg and 200 µg/kg body weights, respectively, for 14 days manifested a notable attenuation of behavioral deficit pattern, oxidative stress, and neurotransmitters' deficiency as compared to the untreated ones. The current study also reports the ameliorative property of the PEGylated compound for progressive neuroinflammation and decreased mitochondrial bioenergetics in astrocytoma cell line, viz., U87. A closer look into the drug mechanism of action of a compact 3D PEGylated block copolymer confirmed its disintegrative interaction with Aß42 fibril via in silico simulation. The results obtained from this study signify the potential of the novel PEGylated block copolymer to ameliorate the cognitive decline and progressive oxidative insults in AD and may envision a successful clinical phase trial. The amelioration of disease condition of colchicine-induced AD rat. Initially the rat has given colchicine via stereotaxic surgery which led to a mimicking condition of AD including neuronal death in hippocampal CA1 region. After recovery from the surgery, the rat was treated with the PEGylated block copolymer through intranasal delivery, and this has led to the decrease in neuronal death in hippocampal CA1 region. The mechanism of drug action has shown by the separation of monomer chains of Aß42.

A simple genetic stratification method for lower cost, more expedient clinical trials in early Alzheimer's disease: A preliminary study of tau PET and cognitive outcomes.

INTRODUCTION: Identifying individuals who are most likely to accumulate tau and exhibit cognitive decline is critical for Alzheimer's disease (AD) clinical trials. METHODS: Participants (N = 235) who were cognitively normal or with mild cognitive impairment from the Alzheimer's Disease Neuroimaging Initiative were stratified by a cutoff on the polygenic hazard score (PHS) at 65th percentile (above as high-risk group and below as low-risk group). We evaluated the associations between the PHS risk groups and tau positron emission tomography and cognitive decline, respectively. Power analyses estimated the sample size needed for clinical trials to detect differences in tau accumulation or cognitive change. RESULTS: The high-risk group showed faster tau accumulation and cognitive decline. Clinical trials using the high-risk group would require a fraction of the sample size as trials without this inclusion criterion. DISCUSSION: Incorporating a PHS inclusion criterion represents a low-cost and accessible way to identify potential participants for AD clinical trials.

Assessment of the In Vivo Relationship Between Cerebral Hypometabolism, Tau Deposition, TSPO Expression, and Synaptic Density in a Tauopathy Mouse Model: a Multi-tracer PET Study.

Cerebral glucose hypometabolism is a typical hallmark of Alzheimer's disease (AD), usually associated with ongoing neurodegeneration and neuronal dysfunction. However, underlying pathological processes are not fully understood and reproducibility in animal models is not well established. The aim of the present study was to investigate the regional interrelation of glucose hypometabolism measured by [18F]FDG positron emission tomography (PET) with various molecular targets of AD pathophysiology using the PET tracers [18F]PI-2620 for tau deposition, [18F]DPA-714 for TSPO expression associated with neuroinflammation, and [18F]UCB-H for synaptic density in a transgenic tauopathy mouse model. Seven-month-old rTg4510 mice (n = 8) and non-transgenic littermates (n = 8) were examined in a small animal PET scanner with the tracers listed above. Hypometabolism was observed throughout the forebrain of rTg4510 mice. Tau pathology, increased TSPO expression, and synaptic loss were co-localized in the cortex and hippocampus and correlated with hypometabolism. In the thalamus, however, hypometabolism occurred in the absence of tau-related pathology. Thus, cerebral hypometabolism was associated with two regionally distinct forms of molecular pathology: (1) characteristic neuropathology of the Alzheimer-type including synaptic degeneration and neuroinflammation co-localized with tau deposition in the cerebral cortex, and (2) pathological changes in the thalamus in the absence of other markers of AD pathophysiology, possibly reflecting downstream or remote adaptive processes which may affect functional connectivity. Our study demonstrates the feasibility of a multitracer approach to explore complex interactions of distinct AD-pathomechanisms in vivo in a small animal model. The observations demonstrate that multiple, spatially heterogeneous pathomechanisms can contribute to hypometabolism observed in AD mouse models and they motivate future longitudinal studies as well as the investigation of possibly comparable pathomechanisms in human patients.

Tau Biomarkers in Dementia: Positron Emission Tomography Radiopharmaceuticals in Tauopathy Assessment and Future Perspective.

Abnormal accumulation of Tau protein is closely associated with neurodegeneration and cognitive impairment and it is a biomarker of neurodegeneration in the dementia field, especially in Alzheimer's disease (AD); therefore, it is crucial to be able to assess the Tau deposits in vivo. Beyond the fluid biomarkers of tauopathy described in this review in relationship with the brain glucose metabolic patterns, this review aims to focus on tauopathy assessment by using Tau PET imaging. In recent years, several first-generation Tau PET tracers have been developed and applied in the dementia field. Common limitations of first-generation tracers include off-target binding and subcortical white-matter uptake; therefore, several institutions are working on developing second-generation Tau tracers. The increasing knowledge about the distribution of first- and second-generation Tau PET tracers in the brain may support physicians with Tau PET data interpretation, both in the research and in the clinical field, but an updated description of differences in distribution patterns among different Tau tracers, and in different clinical conditions, has not been reported yet. We provide an overview of first- and second-generation tracers used in ongoing clinical trials, also describing the differences and the properties of novel tracers, with a special focus on the distribution patterns of different Tau tracers. We also describe the distribution patterns of Tau tracers in AD, in atypical AD, and further neurodegenerative diseases in the dementia field.

Assessment of Tau Pathology as Measured by 18F-THK5317 and 18F-Flortaucipir PET and Their Relation to Brain Atrophy and Cognition in Alzheimer's Disease.

BACKGROUND: In Alzheimer's disease (AD), the abnormal aggregation of hyperphosphorylated tau leads to synaptic dysfunction and neurodegeneration. Recently developed tau PET imaging tracers are candidate biomarkers for diagnosis and staging of AD. OBJECTIVE: We aimed to investigate the discriminative ability of 18F-THK5317 and 18F-flortaucipir tracers and brain atrophy at different stages of AD, and their respective associations with cognition. METHODS: Two cohorts, each including 29 participants (healthy controls [HC], prodromal AD, and AD dementia patients), underwent 18F-THK5317 or 18F-flortaucipir PET, T1-weighted MRI, and neuropsychological assessment. For each subject, we quantified regional 18F-THK5317 and 18F-flortaucipir uptake within six bilateral and two composite regions of interest. We assessed global brain atrophy for each individual by quantifying the brain volume index, a measure of brain volume-to-cerebrospinal fluid ratio. We then quantified the discriminative ability of regional 18F-THK5317, 18F-flortaucipir, and brain volume index between diagnostic groups, and their associations with cognition in patients. RESULTS: Both 18F-THK5317 and 18F-flortaucipir outperformed global brain atrophy in discriminating between HC and both prodromal AD and AD dementia groups. 18F-THK5317 provided the highest discriminative ability between HC and prodromal AD groups. 18F-flortaucipir performed best at discriminating between prodromal and dementia stages of AD. Across all patients, both tau tracers were predictive of RAVL learning, but only 18F-flortaucipir predicted MMSE. CONCLUSION: Our results warrant further in vivo head-to-head and antemortem-postmortem evaluations. These validation studies are needed to select tracers with high clinical validity as biomarkers for early diagnosis, prognosis, and disease staging, which will facilitate their incorporation in clinical practice and therapeutic trials.

Assessment of Amyloid Forming Tendency of Peptide Sequences from Amyloid Beta and Tau Proteins Using Force-Field, Semi-Empirical, and Density Functional Theory Calculations.

A wide variety of neurodegenerative diseases are characterized by the accumulation of protein aggregates in intraneuronal or extraneuronal brain regions. In Alzheimer's disease (AD), the extracellular aggregates originate from amyloid-ß proteins, while the intracellular aggregates are formed from microtubule-binding tau proteins. The amyloid forming peptide sequences in the amyloid-ß peptides and tau proteins are responsible for aggregate formation. Experimental studies have until the date reported many of such amyloid forming peptide sequences in different proteins, however, there is still limited molecular level understanding about their tendency to form aggregates. In this study, we employed umbrella sampling simulations and subsequent electronic structure theory calculations in order to estimate the energy profiles for interconversion of the helix to ß-sheet like secondary structures of sequences from amyloid-ß protein (KLVFFA) and tau protein (QVEVKSEKLD and VQIVYKPVD). The study also included a poly-alanine sequence as a reference system. The calculated force-field based free energy profiles predicted a flat minimum for monomers of sequences from amyloid and tau proteins corresponding to an α-helix like secondary structure. For the parallel and anti-parallel dimer of KLVFFA, double well potentials were obtained with the minima corresponding to α-helix and ß-sheet like secondary structures. A similar double well-like potential has been found for dimeric forms for the sequences from tau fibril. Complementary semi-empirical and density functional theory calculations displayed similar trends, validating the force-field based free energy profiles obtained for these systems.

Machine learning-based decision tree classifier for the diagnosis of progressive supranuclear palsy and corticobasal degeneration.

AIMS: This study aimed to clarify the different topographical distribution of tau pathology between progressive supranuclear palsy (PSP) and corticobasal degeneration (CBD) and establish a machine learning-based decision tree classifier. METHODS: Paraffin-embedded sections of the temporal cortex, motor cortex, caudate nucleus, globus pallidus, subthalamic nucleus, substantia nigra, red nucleus, and midbrain tectum from 1020 PSP and 199 CBD cases were assessed by phospho-tau immunohistochemistry. The severity of tau lesions (i.e., neurofibrillary tangle, coiled body, tufted astrocyte or astrocytic plaque, and tau threads) was semi-quantitatively scored in each region. Hierarchical cluster analysis was performed using tau pathology scores. A decision tree classifier was made with tau pathology scores using 914 cases. Cross-validation was done using 305 cases. An additional ten cases were used for a validation study. RESULTS: Cluster analysis displayed two distinct clusters; the first cluster included only CBD, and the other cluster included all PSP and six CBD cases. We built a decision tree, which used only seven decision nodes. The scores of tau threads in the caudate nucleus were the most decisive factor for predicting CBD. In a cross-validation, 302 out of 305 cases were correctly diagnosed. In the pilot validation study, three investigators made a correct diagnosis in all cases using the decision tree. CONCLUSION: Regardless of the morphology of astrocytic tau lesions, semi-quantitative tau pathology scores in select brain regions are sufficient to distinguish PSP and CBD. The decision tree simplifies neuropathologic differential diagnosis of PSP and CBD.

Synthesis and Assessment of Novel Probes for Imaging Tau Pathology in Transgenic Mouse and Rat Models.

Aggregated tau protein is a core pathology present in several neurodegenerative diseases. Therefore, the development and application of positron emission tomography (PET) imaging radiotracers that selectively bind to aggregated tau in fibril form is of importance in furthering the understanding of these disorders. While radiotracers used in human PET studies offer invaluable insight, radiotracers that are also capable of visualizing tau fibrils in animal models are important tools for translational research into these diseases. Herein, we report the synthesis and characterization of a novel library of compounds based on the phenyl/pyridinylbutadienylbenzothiazoles/benzothiazolium (PBB3) backbone developed for this application. From this library, we selected the compound LM229, which binds to recombinant tau fibrils with high affinity (Kd = 3.6 nM) and detects with high specificity (a) pathological 4R tau aggregates in living cultured neurons and mouse brain sections from transgenic human P301S tau mice, (b) truncated human 151-351 3R (SHR24) and 4R (SHR72) tau aggregates in transgenic rat brain sections, and (c) tau neurofibrillary tangles in brain sections from Alzheimer's disease (3R/4R tau) and progressive supranuclear palsy (4R tau). With LM229 also shown to cross the blood-brain barrier in vivo and its effective radiolabeling with the radioisotope carbon-11, we have established a novel platform for PET translational studies using rodent transgenic tau models.

A freeze-and-thaw-induced fragment of the microtubule-associated protein tau in rat brain extracts: implications for the biochemical assessment of neurotoxicity.

Tau is a microtubule-associated protein (MAP) responsible for controlling the stabilization of microtubules in neurons. Tau function is regulated by phosphorylation. However, in some neurological diseases Tau becomes aberrantly hyperphosphorylated, which contributes to the pathogenesis of neurological diseases, known as tauopathies. Western blotting (WB) has been widely employed to determine Tau levels in neurological disease models. However, Tau quantification by WB should be interpreted with care, as this approach has been recognized as prone to produce artifactual results if not properly performed. In the present study, our goal was to evaluate the influence of a freeze-and-thaw cycle, a common procedure preceding WB, to the integrity of Tau in brain homogenates from rats, 3xTg-AD mice and human samples. Homogenates were prepared in ice-cold RIPA buffer supplemented with protease/phosphatase inhibitors. Immediately after centrifugation, an aliquot of the extracts was analyzed via WB to quantify total and phosphorylated Tau levels. The remaining aliquots of the same extracts were stored for at least 2 weeks at either -20 or -80°C and then subjected to WB. Extracts from rodent brains submitted to freeze-and-thaw presented a ∼25 kDa fragment immunoreactive to anti-Tau antibodies. An in-gel digestion followed by mass spectrometry (MS) analysis in excised bands revealed this ∼25 kDa species corresponds to a Tau fragment. Freeze-and-thaw-induced Tau proteolysis was detected even when extracts were stored at -80°C. This phenomenon was not observed in human samples at any storage condition tested. Based on these findings, we strongly recommend the use of fresh extracts of brain samples in molecular analysis of Tau levels in rodents.