Animal Models Of Tauopathies Research Articles

Novel Small‐Molecular Ligands for Prolyl Oligopeptidase to Combat Tauopathies

AbstractBackgroundTauopathies that include e.g. frontotemporal dementia and Alzheimer’s disease, are neurodegenerative diseases characterized by accumulation of hyperphosphorylated Tau in the brain. Tau is a tempting drug target for tauopathies but no Tau‐targeting therapies are in clinical use. We have recently shown that small‐molecular inhibitors of prolyl oligopeptidase (PREP) can activate protein phosphatase 2A (PP2A) by regulating PREP‐PP2A interaction. PP2A is the main phosphatase regulating Tau, and therefore, we decided to test the efficacy of a reference PREP inhibitor, KYP‐2047, and novel PREP ligands on cellular and animal models of tauopathies.MethodThe compounds, KYP‐2047 and HUP‐46, 55 and 59 (1, 5 and 10 μM), were first screened in the GFP‐2N4R‐Tau transfected HEK‐293 cells where the Tau aggregation was induced by small‐dose okadaic acid (OA; 10 nM), a PP2A inhibitor. The results were verified by using 2N4R‐Tau transfected HEK‐293 cells with 10 nM OA. After the treatment the cells were fractioned to RIPA soluble and SDS‐fractions, and total Tau and S262 phosphorylated Tau (pS262 Tau) levels were studied by Western blot (WB). Finally, KYP‐2047 and HUP‐46 were tested in PS19 Tau transgenic mouse. PS19 mice were treated with the ligands for 4 weeks (10 mg/kg/day, Alzet minipump i.p.) starting from 5.5 months old when the behavioral deficits were observed. After the treatment, the mice behavior was tested by Barnes Maze and locomotor activity assay, and total Tau, pS262 Tau and PP2A levels were studied.ResultIn the GFP‐Tau aggregation assay, all compounds decreased the GFP signal as an indicative of Tau aggregation with 10 mM concentration but HUP‐46 was the most effective (Fig. 1). WB that showed that PREP ligands had the most significant effect on total Tau and pS262 Tau in the SDS‐fraction. In the PS19 mice, KYP‐2047 restored normal behavior in Barnes maze and locomotor activity assay. HUP‐46 had similar effect but did not reach significance (Fig. 2). However, both compounds reduced significantly total Tau and pS262 Tau levels in mouse hippocampus and cortex, and activated PP2A (Fig. 3).ConclusionSmall‐molecular PREP ligands, such as KYP‐2047 and HUP‐46 have disease‐modifying effects on cellular and animal models of tauopathies.

New insights into the therapeutic approaches for the treatment of tauopathies.

Tauopathies are a group of neurological disorders, including Alzheimer's disease and frontotemporal dementia, which involve progressive neurodegeneration, cognitive deficits, and aberrant tau protein accumulation. The development of tauopathies cannot currently be stopped or slowed down by treatment measures. Given the significant contribution of tau burden in primary tauopathies and the strong association between pathogenic tau accumulation and cognitive deficits, there has been a lot of interest in creating therapies that can alleviate tau pathology and render neuroprotective effects. Recently, small molecules, immunotherapies, and gene therapy have been used to reduce the pathological tau burden and prevent neurodegeneration in animal models of tauopathies. However, the major pitfall of the current therapeutic approach is the difficulty of drugs and gene-targeting modalities to cross the blood-brain barrier and their unintended side effects. In this review, the current therapeutic strategies used for tauopathies including the use of oligonucleotide-based gene therapy approaches that have shown a promising result for the treatment of tauopathies and Alzheimer's disease in preclinical animal models, have been discussed.

Tau-RNA complexes inhibit microtubule polymerization and drive disease-relevant conformation change.

Alzheimer's disease and related disorders feature neurofibrillary tangles and other neuropathological lesions composed of detergent-insoluble tau protein. In recent structural biology studies of tau proteinopathy, aggregated tau forms a distinct set of conformational variants specific to the different types of tauopathy disorders. However, the constituents driving the formation of distinct pathological tau conformations on pathway to tau-mediated neurodegeneration remain unknown. Previous work demonstrated RNA can serve as a driver of tau aggregation, and RNA associates with tau containing lesions, but tools for evaluating tau/RNA interactions remain limited. Here, we employed molecular interaction studies to measure the impact of tau/RNA binding on tau microtubule binding and aggregation. To investigate the importance of tau/RNA complexes (TRCs) in neurodegenerative disease, we raised a monoclonal antibody (TRC35) against aggregated tau/RNA complexes. We showed that native tau binds RNA with high affinity but low specificity, and tau binding to RNA competes with tau-mediated microtubule assembly functions. Tau/RNA interaction in vitro promotes the formation of higher molecular weight tau/RNA complexes, which represent an oligomeric tau species. Coexpression of tau and poly(A)45 RNA transgenes in Caenorhabditis elegans exacerbates tau-related phenotypes including neuronal dysfunction and pathological tau accumulation. TRC35 exhibits specificity for Alzheimer's disease-derived detergent-insoluble tau relative to soluble recombinant tau. Immunostaining with TRC35 labels a wide variety of pathological tau lesions in animal models of tauopathy, which are reduced in mice lacking the RNA binding protein MSUT2. TRC-positive lesions are evident in many human tauopathies including Alzheimer's disease, progressive supranuclear palsy, corticobasal degeneration and Pick's disease. We also identified ocular pharyngeal muscular dystrophy as a novel tauopathy disorder, where loss of function in the poly(A) RNA binding protein (PABPN1) causes accumulation of pathological tau in tissue from post-mortem human brain. Tau/RNA binding drives tau conformational change and aggregation inhibiting tau-mediated microtubule assembly. Our findings implicate cellular tau/RNA interactions as modulators of both normal tau function and pathological tau toxicity in tauopathy disorders and suggest feasibility for novel therapeutic approaches targeting TRCs.

Multi‐tissue profiling of RNA and DNA‐methylation signatures of progressive tau neuropathology in tau transgenic mice

AbstractBackgroundIntracellular abnormal aggregation of hyperphosphorylated tau protein is a neuropathological hallmark associated with Alzheimer’s Disease (AD). The insoluble tau aggregates called neurofibrillary tangles lead to microtubule dysfunction and neurodegeneration with AD progression. The TG4510 transgenic mice overexpressing the P301L mutant human tau protein exhibit region‐specific tau inclusions, brain atrophy, and behavioral deficits, thereby serving as a widely used animal model of tauopathy in AD. This study aims to characterize the tissue‐specific transcriptomic and epigenomic signatures of progressive tau pathology in TG4510 mice as well as investigate the correlation of brain and blood expression and epigenomic changes.MethodUsing UMI‐based bulk RNA‐Sequencing and Reduced‐representation Bisulfite Sequencing (RRBS), we profiled the gene expression and DNA‐methylation changes respectively, in the cortex, hippocampus, cerebellum and pre‐sacrifice and terminal whole blood between three age groups (2‐3 months, 6‐7 months, 8‐9 months old; N=20/tissue/group) of male and female TG4510 mice.ResultThe cortex and hippocampus show the largest transcriptional differences between mice, recapitulating phenotype differences with loss of neurotransmission and increased inflammation in these tissues. Systems‐level co‐expression analysis reveals common gene modules in immune system, behaviour and cognition and ECM/synaptic pathways between cortex/hippocampus and blood. Furthermore, the genes correlated between brain regions and blood in these mice were prioritized as potential biomarkers of tau pathology, which can be further validated in preclinical animal models and ultimately human studies.ConclusionThis study serves as one of the first evidence of multiomic molecular underpinnings in a tauopathy mouse model providing potential novel targets and biomarkers of progressive tau pathology in AD.DisclosuresAll authors are employees of AbbVie. The design, study conduct, and financial support for this research were provided by AbbVie. AbbVie participated in the interpretation of data, review, and approval of the publication.

Tetrandrine ameliorates cognitive deficits and mitigates tau aggregation in cell and animal models of tauopathies

BackgroundTauopathies are neurodegenerative diseases that are associated with the pathological accumulation of tau-containing tangles in the brain. Tauopathy can impair cognitive and motor functions and has been observed in Alzheimer’s disease (AD) and frontotemporal dementia (FTD). The aetiology of tauopathy remains mysterious; however, recent studies suggest that the autophagic-endolysosomal function plays an essential role in the degradation and transmission of pathological tau. We previously demonstrated that tetrandrine could ameliorate memory functions and clear amyloid plaques in transgenic AD mice by restoring autophagic-endolysosomal function. However, the efficacy of tetrandrine and the associated therapeutic mechanism in tauopathies have not been evaluated and elucidated.MethodsNovel object recognition, fear conditioning and electrophysiology were used to evaluate the effects of tetrandrine on memory functions in transgenic tau mice. Western blotting and immunofluorescence staining were employed to determine the effect of tetrandrine on autophagy and tau clearance in vivo. Calcium (Ca2+) imaging and flow cytometry were used to delineate the role of pathological tau and tetrandrine in lysosomal Ca2+ and pH homeostasis. Biochemical BiFC fluorescence, Western blotting and immunofluorescence staining were used to evaluate degradation of hyperphosphorylated tau in vitro, whereas coculture of brain slices with isolated microglia was used to evaluate tau clearance ex vivo.ResultsWe observed that tetrandrine treatment mitigated tau tangle development and corrected memory impairment in Thy1-hTau.P301S transgenic mice. Mechanistically, we showed that mutant tau expression disrupts lysosome pH by increasing two-pore channel 2 (TPC2)-mediated Ca2+ release, thereby contributing to lysosome alkalinization. Tetrandrine inhibits TPC2, thereby restoring the lysosomal pH, promotes tau degradation via autophagy, and ameliorates tau aggregation. Furthermore, in an ex vivo assay, we demonstrated that tetrandrine treatment promotes pathological tau clearance by microglia.ConclusionsTogether, these findings suggest that pathological tau disturbs endolysosomal homeostasis to impair tau clearance. This impairment results in a vicious cycle that accelerates disease pathogenesis. The success of tetrandrine in reducing tau aggregation suggests first, that tetrandrine could be an effective drug for tauopathies and second, that rescuing lysosomal Ca2+ homeostasis, thereby restoring ALP function, could be an effective general strategy for the development of novel therapies for tauopathies.

Magnetic Resonance Imaging in Tauopathy Animal Models.

The microtubule-associated protein tau plays an important role in tauopathic diseases such as Alzheimer’s disease and primary tauopathies such as progressive supranuclear palsy and corticobasal degeneration. Tauopathy animal models, such as transgenic, knock-in mouse and rat models, recapitulating tauopathy have facilitated the understanding of disease mechanisms. Aberrant accumulation of hyperphosphorylated tau contributes to synaptic deficits, neuroinflammation, and neurodegeneration, leading to cognitive impairment in animal models. Recent advances in molecular imaging using positron emission tomography (PET) and magnetic resonance imaging (MRI) have provided valuable insights into the time course of disease pathophysiology in tauopathy animal models. High-field MRI has been applied for in vivo imaging in animal models of tauopathy, including diffusion tensor imaging for white matter integrity, arterial spin labeling for cerebral blood flow, resting-state functional MRI for functional connectivity, volumetric MRI for neurodegeneration, and MR spectroscopy. In addition, MR contrast agents for non-invasive imaging of tau have been developed recently. Many preclinical MRI indicators offer excellent translational value and provide a blueprint for clinical MRI in the brains of patients with tauopathies. In this review, we summarized the recent advances in using MRI to visualize the pathophysiology of tauopathy in small animals. We discussed the outstanding challenges in brain imaging using MRI in small animals and propose a future outlook for visualizing tau-related alterations in the brains of animal models.

Structure-Based Design and Biological Evaluation of Novel Caspase-2 Inhibitors Based on the Peptide AcVDVAD-CHO and the Caspase-2-Mediated Tau Cleavage Sequence YKPVD314.

Alzheimer's disease (AD) was first described by Alois Alzheimer over 100 years ago, but there is still no overarching theory that can explain its cause in detail. There are also no effective therapies to treat either the cause or the associated symptoms of this devastating disease. A potential approach to better understand the pathogenesis of AD could be the development of selective caspase-2 (Casp2) probes, as we have shown that a Casp2-mediated cleavage product of tau (Δtau314) reversibly impairs cognitive and synaptic function in animal models of tauopathies. In this article, we map out the Casp2 binding site through the preparation and assay of a series of 35 pentapeptide inhibitors with the goal of gaining selectivity against caspase-3 (Casp3). We also employed computational docking methods to understand the key interactions in the binding pocket of Casp2 and the differences predicted for binding at Casp3. Moreover, we crystallographically characterized the binding of selected pentapeptides with Casp3. Furthermore, we engineered and expressed a series of recombinant tau mutants and investigated them in an in vitro cleavage assay. These studies resulted in simple peptidic inhibitors with nanomolar affinity, for example, AcVDV(Dab)D-CHO (24) with up to 27.7-fold selectivity against Casp3. Our findings provide a good basis for the future development of selective Casp2 probes and inhibitors that can serve as pharmacological tools in planned in vivo studies and as lead compounds for the design of bioavailable and more drug-like small molecules.

Cornel iridoid glycoside protects against STAT1‐dependent synapse and memory deficits by increasing NMDA receptor expression in tau transgenic mice

AbstractBackgroundP301S transgenic mice is an animal model of tauopathy and Alzheimer’s disease (AD), exhibiting tau pathology and synaptic dysfunction. Cornel iridoid glycoside (CIG) is an active ingredient extracted from Cornus officinalis, a traditional Chinese herb. The purpose of the present study was to investigate the effects and mechanisms of CIG on tau pathology and synaptic dysfunction using P301S mice.MethodIntragastric administration of CIG for 3.5 months was applied to P301S mice from 9‐month‐old to 12.5‐month‐old of age. Objective recognition test and Y maze task were used to test the cognitive impairments of P301S mice. Electrophysiological recording was applied to record long‐term potential. Transmission electron microscopy was used to investigate the the ultrastructure and number of synapse. Immunohistochemical staining was applied to detect tau phosphorylation. Western blotting was used to detect the expression levels of related proteins.ResultsIntragastric administration of CIG for 3.5 months improved learning and memory abilities, increased the survival rate of P301S mice. Electrophysiological recordings and transmission Electron microscopy study showed that CIG improved synaptic plasticity, increased the ultrastructure and number of synapse. Moreover, CIG increased the expression of N‐methyl‐D‐aspartate receptors (NMDARs) subunits GluN1, GluN2A and GluN2B, α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole propionic acid receptor (AMPAR) subunit GluA1. The major mechanism of CIG involved in the regulation of synaptic dysfunction was to inhibited the activation of Janus kinase‐2 (JAK2) /signal transducer and activator of transcription 1 (STAT1) signaling pathway and STAT1‐induced suppression of NMDAR expression.ConclusionBased on our findings, CIG might be a promising candidate for the prevention of tauopathy such as AD.

HCN channelopathy couples disease‐associated tau to synaptic dysfunction

AbstractBackgroundDeficits in synaptic function and neurite connectivity are early correlates of tauopathies. Altered tau processing and mis‐localization coincide with reductions in synapse density and synaptic function, suggesting a causal role for tau in disease pathogenesis. In parallel, altered activity of hyperpolarization‐activated cyclic nucleotide‐gated (HCN) channels, involved in synaptic integration, is implicated in disease progression in animal models of tauopathy. However, the mechanisms underlying these pathological events are not fully understood.MethodsWe performed histological and biochemical analyses of hippocampal tissues from Alzheimer’s disease (AD), age‐matched controls and Tau35 mice, in which a human tauopathy‐associated tau fragment is minimally expressed. Image analysis and quantification were performed on antibody‐labelled human and mouse hippocampal sections (Visiopharm). Brain tissue from Tau35 and wild‐type (WT) mice was also examined on western blots, by Golgi‐Cox staining and transmission electron microscopy. Primary hippocampal neurons from Tau35 and WT mice were transfected with a plasmid expressing enhanced green fluorescent protein (eGFP) and imaged up to 14 days in vitro. Three‐dimensional digital reconstruction of dendrites and spines enabled morphological analysis of eGFP‐expressing neurons. In parallel, patch‐clamp recordings were performed to assess functional changes and HCN channel activity in Tau35 neurons.ResultsHere we show that HCN channels are functionally linked to tau abnormalities. Alterations in HCN channel expression are detected in both Tau35 mouse and human post‐mortem AD brain. Tau35 neurons exhibit altered synaptic cytoarchitecture, including progressive reductions in dendritic branching, pre‐synaptic vesicles, spine density and synaptic markers, along with the development of tau pathology. These changes are accompanied by functional abnormalities in network activity, including increased HCN‐dependent sag voltage, reduced frequency and slower kinetics of spontaneous excitatory postsynaptic currents.ConclusionsOur findings are consistent with the hypothesis that disease‐associated tau species impact on HCN channels to drive network‐wide structural and functional synaptic deficits, with significant therapeutic relevance for tauopathies.

Animal models of tauopathy

Animal models of tauopathy

Cornel Iridoid Glycoside Protects Against STAT1-Dependent Synapse and Memory Deficits by Increasing N-Methyl-D-aspartate Receptor Expression in a Tau Transgenic Mice.

P301S transgenic mice are an animal model of tauopathy and Alzheimer’s disease (AD), exhibiting tau pathology and synaptic dysfunction. Cornel iridoid glycoside (CIG) is an active ingredient extracted from Cornus officinalis, a traditional Chinese herb. In the present study, the purpose was to investigate the effects and mechanisms of CIG on tau pathology and synaptic dysfunction using P301S transgenic mice. The results showed that intragastric administration of CIG for 3.5 months improved cognitive impairments and the survival rate of P301S mice. Electrophysiological recordings and transmission electron microscopy study showed that CIG improved synaptic plasticity and increased the ultrastructure and number of synapse. Moreover, CIG increased the expression levels of N-methyl-D-aspartate receptors (NMDAR) subunits GluN1, GluN2A, and GluN2B, and α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor (AMPAR) subunit GluA1. We inferred that the major mechanism of CIG involving in the regulation of synaptic dysfunctions was inhibiting the activation of Janus kinase-2 (JAK2)/signal transducer and activator of transcription 1 (STAT1) signaling pathway and alleviating STAT1-induced suppression of NMDAR expressions. Based on our findings, we thought CIG might be a promising candidate for the therapy of tauopathy such as AD.

Reversible Tau Phosphorylation Induced by Synthetic Torpor in the Spinal Cord of the Rat.

Tau is a key protein in neurons, where it affects the dynamics of the microtubule system. The hyperphosphorylation of Tau (PP-Tau) commonly leads to the formation of neurofibrillary tangles, as it occurs in tauopathies, a group of neurodegenerative diseases, including Alzheimer's. Hypothermia-related accumulation of PP-Tau has been described in hibernators and during synthetic torpor (ST), a torpor-like condition that has been induced in rats, a non-hibernating species. Remarkably, in ST PP-Tau is reversible and Tau de-phosphorylates within a few hours following the torpor bout, apparently not evolving into pathology. These observations have been limited to the brain, but in animal models of tauopathies, PP-Tau accumulation also appears to occur in the spinal cord (SpCo). The aim of the present work was to assess whether ST leads to PP-Tau accumulation in the SpCo and whether this process is reversible. Immunofluorescence (IF) for AT8 (to assess PP-Tau) and Tau-1 (non-phosphorylated Tau) was carried out on SpCo coronal sections. AT8-IF was clearly expressed in the dorsal horns (DH) during ST, while in the ventral horns (VH) no staining was observed. The AT8-IF completely disappeared after 6 h from the return to euthermia. Tau-1-IF disappeared in both DH and VH during ST, returning to normal levels during recovery. To shed light on the cellular process underlying the PP-Tau pattern observed, the inhibited form of the glycogen-synthase kinase 3β (the main kinase acting on Tau) was assessed using IF: VH (i.e., in motor neurons) were highly stained mainly during ST, while in DH there was no staining. Since tauopathies are also related to neuroinflammation, microglia activation was also assessed through morphometric analyses, but no ST-induced microglia activation was found in the SpCo. Taken together, the present results show that, in the DH of SpCo, ST induces a reversible accumulation of PP-Tau. Since during ST there is no motor activity, the lack of AT8-IF in VH may result from an activity-related process at a cellular level. Thus, ST demonstrates a newly-described physiological mechanism that is able to resolve the accumulation of PP-Tau and apparently avoid the neurodegenerative outcome.

Characterization and Systemic Delivery of Dibenzoylmethane via the Intranasal Route.

Intranasal (IN) administration is known to be noninvasive with the potential to carry a drug or vaccine directly to the blood, bypassing first-pass metabolism in the liver and the harsh environment of the gastrointestinal system. Orally administered dibenzoylmethane (DBM) has been shown experimentally to be neuroprotective in animal models of tauopathy and prion disease and effective in the treatment of certain forms of cancers. The purpose of this study was to prepare, characterize, and test formulations of DBM designed for IN administration. DBM was formulated in brain homogenate (BH) and hypromellose and as nanoparticles (NPs). These formulations were detected using UPLC and characterized in solid and suspension states; NPs were also characterized by in vitro cell culture-based studies. Particle size for DBM NP was 163.8 ± 3.2nm, and in vitro release studies showed 95.80% of DBM was released from the NPs within 8days. In vitro cell, culture studies suggested no drug uptake until 6h. A histological analysis of nasal cavity (NC) sections and blood detection studies were carried out 30min after inhalation. DBM amounting to 40.77 ± 4.93 and 44.45 ± 5.36ng/mL was detected in the blood of animals administered DBM in polymeric and NP formulation, respectively. Histological studies on NCs confirmed the presence of BH within lymphatic vessels in the lamina propria of each animal; BH was identified traversing the mucosa in 2 animals. Thus, formulations for DBM administered via IN route were successfully designed and characterized and able to cross the nasal mucosa following inhalation.

Neuroprotection of cholinergic neurons with tau aggregation inhibitor and rivastigmine in L1 mice with Alzheimer’s‐like tauopathy

AbstractBackgroundBasal forebrain cholinergic dysfunction, likely linked to tau aggregation pathology, is a characteristic feature of AD. Cholinergic neurons contain choline acetyltransferase (ChAT) and high‐affinity tropomyosin‐related kinase A (TrkA) and send efferents to cortex and hippocampus where they release acetylcholine (ACh). The vesicular acetylcholine transporter (VAChT) is responsible for loading ACh into secretory vesicles and acetylcholine esterase (AChE) hydrolyses ACh in the synaptic cleft. We here aimed to evaluate the cholinergic phenotype in the Line 1 animal model of tauopathy and to determine the effect of the choline esterase inhibitor rivastigmine alone and in conjunction with the tau aggregation inhibitor hydromethylthionine on the cholinergic system.MethodLine 1 (L1) and control NMRI mice, 8‐9 months old, divided into 13 groups (n = 5 each), were treated with rivastigmine (0.1 and 0.5 mg/kg) and hydromethylthionine (5 and 15 mg/kg) and their combinations for 11 weeks. Immunohistochemical staining in brain sections was performed for ChAT, TrkA, VAChT and tau with a repeat domain monoclonal antibody (TauRx Therapeutics Ltd.). AChE activity was measured histochemically. The state of the cholinergic system was determined by stereological counting of ChAT‐ir and TrkA‐ir neurons, while Relative Optical Intensity (ROI) was used to tau immunoreactivity in basal forebrain, and cholinergic projections were evaluated using VAChT and AChE ROI.ResultNumber of ChAT‐ir, TrkA‐ir neurons and ROI value for VAChT and AChE were significantly lower while ROI value for tau was higher in vehicle‐treated L1 mice compared with wild type control. Numbers of ChAT‐ir, TrkA‐ir neurons and ROI value of VAChT and AChE in L1 mice were increased and tau ROI was decreased by hydromethylthionine treatment. Combined treatment decreased numbers of ChAT‐ir, TrkA‐ir neurons and ROI for VAChT and AChE compared to L1 groups treated with hydromethylthionine alone.ConclusionThere was a significant loss of cholinergic basal forebrain neurones, impaired cholinergic projection and increased tau staining in L1 mice. Monotherapy with hydromethylthionine improved the cholinergic neurons phenotype, enhanced their projection and reduced tau pathology. Combination therapy interacted negatively attenuating the effect of hydromethylthionine given alone.

Cellular Biology of Tau Diversity and Pathogenic Conformers

Tau accumulation is a prominent feature in a variety of neurodegenerative disorders and remarkable effort has been expended working out the biochemistry and cell biology of this cytoplasmic protein. Tau's wayward properties may derive from germline mutations in the case of frontotemporal lobar degeneration (FTLD-MAPT) but may also be prompted by less understood cues—perhaps environmental or from molecular damage as a consequence of chronological aging—in the case of idiopathic tauopathies. Tau properties are undoubtedly affected by its covalent structure and in this respect tau protein is not only subject to changes in length produced by alternative splicing and endoproteolysis, but different types of posttranslational modifications that affect different amino acid residues. Another layer of complexity concerns alternate conformations—“conformers”—of the same covalent structures; in vivo conformers can encompass soluble oligomeric species, ramified fibrillar structures evident by light and electron microscopy and other forms of the protein that have undergone liquid-liquid phase separation to make demixed liquid droplets. Biological concepts based upon conformers have been charted previously for templated replication mechanisms for prion proteins built of the PrP polypeptide; these are now providing useful explanations to feature tau pathobiology, including how this protein accumulates within cells and how it can exhibit predictable patterns of spread across different neuroanatomical regions of an affected brain. In sum, the documented, intrinsic heterogeneity of tau forms and conformers now begins to speak to a fundamental basis for diversity in clinical presentation of tauopathy sub-types. In terms of interventions, emphasis upon subclinical events may be worthwhile, noting that irrevocable cell loss and ramified protein assemblies feature at end-stage tauopathy, whereas earlier events may offer better opportunities for diverting pathogenic processes. Nonetheless, the complexity of tau sub-types, which may be present even within intermediate disease stages, likely mitigates against one-size-fits-all therapeutic strategies and may require a suite of interventions. We consider the extent to which animal models of tauopathy can be reasonably enrolled in the campaign to produce such interventions and to slow the otherwise inexorable march of disease progression.

The role of APOE in Microglia regulation in Neurogeneration

Alzheimer’s disease (AD) is the most prevalent senile dementia affecting 4.5 million Americans. Neuroinflammatory changes are prominent and may significantly contribute to the pathologic process. Mononuclear phagocytes (brain resident microglia and recruited peripheral monocytes) accumulate around amyloid plaque in AD brains. However, their exact cellular identity, molecular and functional phenotypes, and their protective or destructive roles in AD are not well understood. This stems in part from the lack of a specific molecular signatures for mononuclear phagocytes, cell type-specific antibodies, and analytic tools for in situ characterization. We recently identified a unique TGFβ-dependent molecular signature of homeostatic (M0)- and APOE-dependent (MGnD)- microglia in mouse models including APP-PS1 mice and human AD. Mechanistically, the TREM2-APOE pathway mediates a switch from M0- to MGnD-microglia phenotype after phagocytosis of apoptotic neurons in a cell-autonomous manner. TREM2 induces APOE signaling which is a negative regulator of the transcription program in M0-microglia.Transcription regulatory network analysis identified direct effect of APOE on suppression of major microglial homeostatic regulators including TGFβ signaling and induction of disease-associated molecules which are essential for pathogenicity in neuroinflammation. Specific genetic ablation of Apoe and/orTrem2 in microglia restored their homeostatic phenotype and genetic ablation of Apoe or Trem2 in TAU (P301S) mice arrested neurodegeneration and brain atrophy. Therefore, APOE plays an important role in microglia phenotype regulation in conditions, and restoration of the homeostatic microglia by targeting the APOE-signaling in microglia represents a novel immunotherapeutic approach. Taken together, our work identifies the TREM2-APOE pathway as a major regulator of microglial functional phenotype in diseases and serves as a novel target to restore homeostatic microglia. These advances have major implications not only for understanding normal CNS function, but have opened up new avenues to understand the role of microglia in disease and most importantly have created the opportunity for consideration of ways in which microglial may be imaged and targeted for the treatment of disease. Since APOE e4 is the major risk factor of the disease, we study the role of APOE e4 in microglia regulation by employing novel tools including new mouse models and techniques to specifically target APOE in order to restore microglia-mediated protein clearance and brain function in animal models of tauopathies and AD. I will present recent advances in understanding the new molecular signature of homeostatic microglia, disease associated microglia and how microglia are regulated in health and disease. This impairment triggers a progressive loss of cognitive skills such as memory and decision making. Neurodegeneration is a key aspect of a large number of diseases that come under the umbrella of neurodegenerative diseases. Of these hundreds of different disorders, attention has so far focused mainly on just a handful, with Parkinson's disease, Huntington disease, being the most notable and Alzheimer's disease. A large proportion of the lesser-advertised diseases were essentially ignored. Such disorders can lead to irreversible damage to the brain and neurodegeneration. While all three of the diseases manifest with different clinical characteristics, the mechanisms of disease tend to be identical at the cellular level. Parkinson's disease, for example , affects the basal ganglia in the Mind, it depletes dopamine. It results in swelling, swelling and tremors in the body's main muscles, which are common features of the disease. There are deposits of tiny protein plaques in Alzheimer's disease which damage various parts of the brain and lead to progressive memory loss. Huntington's disease is a progressive genetic disorder which affects the body’s major muscles Severe restriction on the engine, and ultimately death.

FTDP-17 Mutations Alter the Aggregation and Microtubule Stabilization Propensity of Tau in an Isoform-Specific Fashion.

More than 50 different intronic and exonic autosomal dominant mutations in the tau gene have been linked to the neurodegenerative disorder frontotemporal dementia with Parkinsonism linked to chromosome-17 (FTDP-17). Although the pathological and clinical presentation of this disorder is heterogeneous among patients, the deposition of tau as pathological aggregates is a common feature. Collectively, FTDP-17 mutations have been shown to alter tau's ability to stabilize microtubules, enhance its aggregation, alter mRNA splicing, or induce its hyperphosphorylation, among other effects. Previous in vitro studies from our lab revealed that these mutations differ markedly from each other in the longest 2N4R isoform of tau. However, it is not entirely known whether the effect of a single mutation varies when compared between different isoforms of tau. Differences in the isoelectric points of the N-terminal region of tau isoforms lead to changes in their biochemical properties, raising the possibility that isoforms could also be disproportionately affected by disease-related mechanisms such as mutations. We therefore performed a comparative study of three FTDP-17 mutations present in different regions of tau (R5L, P301L, and R406W) in the three 4R isoforms of tau. We observed significant differences in the effect these mutations exert on the total amount and kinetics of aggregation, aggregate length distributions, and microtubule stabilizing propensity of 4R tau isoforms for all three selected mutants. These results demonstrate that different combinations of FTDP-17 mutations and tau isoforms are functionally distinct and could have important implications for our understanding of disease and animal models of tauopathies.

Regenerating islet-derived 1α (REG-1α) protein increases tau phosphorylation in cell and animal models of tauopathies

REG-1α, a secreted protein containing a C-type lectin domain, is expressed in various organs and plays different roles in digestive system cells in physiological and pathological conditions. Like other members of the Reg family, REG-1α is expressed also in the brain where it has different functions. For instance, we previously reported that REG-1α regulates neurite outgrowth and is overexpressed during the very early stages of Alzheimer's disease (AD). However, REG-1α function in neural cells during neural degeneration remains unknown. First, REG-1α and phosphorylated tau expression were assessed in tissue sections from the hippocampus, representing neurofibrillary tangles (NFTs), from patients with AD, and from basal ganglia, representing subcortical NFTs, from patients with progressive supranuclear palsy (PSP). We found an association between REG-1α expression, tau hyperphosphorylation and NFTs in human brain samples from patients with these neurodegenerative diseases. Then, the effects of REG-1α overexpression on tau phosphorylation and axonal morphology were investigated i) in primary cultures of rat neurons that express human tau P301L and ii) in a transgenic zebrafish model of tauopathy that expresses human tau P301L. In the tau P301L cell model, REG-1α overexpression increased tau phosphorylation at the S202/T205 and S396 residues (early and late stages of abnormal phosphorylation, respectively) through the AKT/GSK3-β pathway. This effect was associated with axonal defects both in tau P301L-expressing rat neurons and zebrafish embryos. Our findings suggest a functional role for REG-1α during tauopathy development and progression and, specifically, its involvement in the modification of tau phosphorylation temporal sequence.

Novel Key Players in the Development of Tau Neuropathology: Focus on the 5-Lipoxygenase.

Tauopathies belong to a large group of neurodegenerative diseases characterized by progressive accumulation of hyperphosphorylated tau. Tau is a microtubule binding protein which is necessary for their assembly and stability. However, tau affinity for microtubules mainly depends on its phosphorylation status, which is the result of a delicate balance between kinases and phosphatases activity. Any significant changes in this equilibrium can promote tau fibrillation, aggregation, neuronal dysfunction, and ultimately neuronal loss. Despite intensive research, the molecular mechanism(s) leading to tau hyperphosphorylation are still unknown and there is no cure for these diseases. Development of an effective strategy that successfully prevents tau excessive phosphorylation and/or tau aggregation may offer a real therapeutic opportunity for these less investigated neurodegenerative conditions. Beside tau, chronic brain inflammation is a common feature of all tauopathies and 5-lipoxygenase, an inflammatory enzyme, is upregulated in brain regions affected by tau pathology. Recently, in vitro studies and preclinical investigations with animal models of tauopathy have implicated 5-lipoxygenase in the regulation of tau phosphorylation through activation of the cyclin-dependent kinase 5 pathway, supporting the novel hypothesis that this protein is a promising therapeutic target for the treatment of tauopathies. In this article, we will discuss the contribution of the 5-lipoxygenase signaling pathway in the development of tau neuropathology, and the promising potential that drugs targeting this enzyme activation hold as a novel disease-modifying therapeutic approach for tauopathies.

Targeting metals rescues the phenotype in an animal model of tauopathy.

Tauopathies are characterized by the pathological accumulation of the microtubule associated protein tau within the brain. We demonstrate here that a copper/zinc chaperone (PBT2, Prana Biotechnology) has rapid and profound effects in the rTg(tauP301L)4510 mouse model of tauopathy. This was evidenced by significantly improved cognition, a preservation of neurons, a decrease in tau aggregates and a decrease in other forms of "pathological" tau (including phosphorylated tau and sarkosyl-insoluble tau). Our data demonstrate that one of the primary mechanisms of action of PBT2 in this model may be driven by an interaction on the pathways responsible for the dephosphorylation of tau. Specifically, PBT2 increased protein levels of both the structural and catalytic subunits of protein phosphatase 2A (PP2A), decreased levels of the methyl esterase (PME1) that dampens PP2A activity, and increased levels of the prolyl isomerase (Pin1) that stimulates the dephosphorylation activity of PP2A. None of these effects were observed when the metal binding site of PBT2 was blocked. This highlights the potential utility of targeting metal ions as a novel therapeutic strategy for diseases in which tau pathology is a feature, which includes conditions such as frontotemporal dementia and Alzheimer's disease.