RESUMO
Tauopathies are age-associated neurodegenerative diseases whose mechanistic underpinnings remain elusive, partially due to a lack of appropriate human models. Here, we engineered human induced pluripotent stem cell (hiPSC)-derived neuronal lines to express 4R Tau and 4R Tau carrying the P301S MAPT mutation when differentiated into neurons. 4R-P301S neurons display progressive Tau inclusions upon seeding with Tau fibrils and recapitulate features of tauopathy phenotypes including shared transcriptomic signatures, autophagic body accumulation, and reduced neuronal activity. A CRISPRi screen of genes associated with Tau pathobiology identified over 500 genetic modifiers of seeding-induced Tau propagation, including retromer VPS29 and genes in the UFMylation cascade. In progressive supranuclear palsy (PSP) and Alzheimer's Disease (AD) brains, the UFMylation cascade is altered in neurofibrillary-tangle-bearing neurons. Inhibiting the UFMylation cascade in vitro and in vivo suppressed seeding-induced Tau propagation. This model provides a robust platform to identify novel therapeutic strategies for 4R tauopathy.
Assuntos
Células-Tronco Pluripotentes Induzidas , Neurônios , Tauopatias , Proteínas tau , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Proteínas tau/metabolismo , Tauopatias/metabolismo , Tauopatias/patologia , Neurônios/metabolismo , Neurônios/patologia , Animais , Camundongos , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Doença de Alzheimer/genética , Encéfalo/metabolismo , Encéfalo/patologia , Paralisia Supranuclear Progressiva/metabolismo , Paralisia Supranuclear Progressiva/patologia , Paralisia Supranuclear Progressiva/genética , Diferenciação Celular , Mutação , AutofagiaRESUMO
Tau (MAPT) drives neuronal dysfunction in Alzheimer disease (AD) and other tauopathies. To dissect the underlying mechanisms, we combined an engineered ascorbic acid peroxidase (APEX) approach with quantitative affinity purification mass spectrometry (AP-MS) followed by proximity ligation assay (PLA) to characterize Tau interactomes modified by neuronal activity and mutations that cause frontotemporal dementia (FTD) in human induced pluripotent stem cell (iPSC)-derived neurons. We established interactions of Tau with presynaptic vesicle proteins during activity-dependent Tau secretion and mapped the Tau-binding sites to the cytosolic domains of integral synaptic vesicle proteins. We showed that FTD mutations impair bioenergetics and markedly diminished Tau's interaction with mitochondria proteins, which were downregulated in AD brains of multiple cohorts and correlated with disease severity. These multimodal and dynamic Tau interactomes with exquisite spatial resolution shed light on Tau's role in neuronal function and disease and highlight potential therapeutic targets to block Tau-mediated pathogenesis.
Assuntos
Mitocôndrias/metabolismo , Degeneração Neural/metabolismo , Mapas de Interação de Proteínas , Sinapses/metabolismo , Proteínas tau/metabolismo , Doença de Alzheimer/genética , Aminoácidos/metabolismo , Biotinilação , Encéfalo/metabolismo , Encéfalo/patologia , Núcleo Celular/metabolismo , Progressão da Doença , Metabolismo Energético , Demência Frontotemporal/genética , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Proteínas Mutantes/metabolismo , Mutação/genética , Degeneração Neural/patologia , Neurônios/metabolismo , Ligação Proteica , Domínios Proteicos , Proteômica , Índice de Gravidade de Doença , Frações Subcelulares/metabolismo , Tauopatias/genética , Proteínas tau/químicaRESUMO
Many neurodegenerative diseases are characterized by the accumulation of insoluble protein aggregates, including neurofibrillary tangles comprised of tau in Alzheimer's disease and Lewy bodies composed of α-synuclein in Parkinson's disease. Moreover, different pathological proteins frequently codeposit in disease brains. To test whether aggregated α-synuclein can directly cross-seed tau fibrillization, we administered preformed α-synuclein fibrils assembled from recombinant protein to primary neurons and transgenic mice. Remarkably, we discovered two distinct strains of synthetic α-synuclein fibrils that demonstrated striking differences in the efficiency of cross-seeding tau aggregation, both in neuron cultures and in vivo. Proteinase K digestion revealed conformational differences between the two synthetic α-synuclein strains and also between sarkosyl-insoluble α-synuclein extracted from two subgroups of Parkinson's disease brains. We speculate that distinct strains of pathological α-synuclein likely exist in neurodegenerative disease brains and may underlie the tremendous heterogeneity of synucleinopathies.
Assuntos
Neurônios/metabolismo , Doença de Parkinson/patologia , alfa-Sinucleína/metabolismo , Proteínas tau/metabolismo , Amiloide/química , Amiloide/metabolismo , Animais , Células Cultivadas , Embrião de Mamíferos/metabolismo , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Doença de Parkinson/metabolismo , Proteínas Recombinantes/metabolismo , alfa-Sinucleína/químicaRESUMO
Amyloid-forming proteins such α-synuclein and tau, which are implicated in Alzheimer's and Parkinson's disease, can form different fibril structures or strains with distinct toxic properties, seeding activities and pathology. Understanding the determinants contributing to the formation of different amyloid features could open new avenues for developing disease-specific diagnostics and therapies. Here we report that O-GlcNAc modification of α-synuclein monomers results in the formation of amyloid fibril with distinct core structure, as revealed by cryogenic electron microscopy, and diminished seeding activity in seeding-based neuronal and rodent models of Parkinson's disease. Although the mechanisms underpinning the seeding neutralization activity of the O-GlcNAc-modified fibrils remain unclear, our in vitro mechanistic studies indicate that heat shock proteins interactions with O-GlcNAc fibril inhibit their seeding activity, suggesting that the O-GlcNAc modification may alter the interactome of the α-synuclein fibrils in ways that lead to reduce seeding activity in vivo. Our results show that posttranslational modifications, such as O-GlcNAc modification, of α-synuclein are key determinants of α-synuclein amyloid strains and pathogenicity.
Assuntos
Amiloide , alfa-Sinucleína , alfa-Sinucleína/metabolismo , alfa-Sinucleína/química , Amiloide/metabolismo , Humanos , Animais , Camundongos , Doença de Parkinson/metabolismo , Doença de Parkinson/patologia , Acetilglucosamina/metabolismo , Acetilglucosamina/química , Processamento de Proteína Pós-Traducional , Microscopia Crioeletrônica , Neurônios/metabolismo , Neurônios/patologiaRESUMO
Cell death in human diseases is often a consequence of disrupted cellular homeostasis. If cell death is prevented without restoring cellular homeostasis, it may lead to a persistent dysfunctional and pathological state. Although mechanisms of cell death have been thoroughly investigated1-3, it remains unclear how homeostasis can be restored after inhibition of cell death. Here we identify TRADD4-6, an adaptor protein, as a direct regulator of both cellular homeostasis and apoptosis. TRADD modulates cellular homeostasis by inhibiting K63-linked ubiquitination of beclin 1 mediated by TRAF2, cIAP1 and cIAP2, thereby reducing autophagy. TRADD deficiency inhibits RIPK1-dependent extrinsic apoptosis and proteasomal stress-induced intrinsic apoptosis. We also show that the small molecules ICCB-19 and Apt-1 bind to a pocket on the N-terminal TRAF2-binding domain of TRADD (TRADD-N), which interacts with the C-terminal domain (TRADD-C) and TRAF2 to modulate the ubiquitination of RIPK1 and beclin 1. Inhibition of TRADD by ICCB-19 or Apt-1 blocks apoptosis and restores cellular homeostasis by activating autophagy in cells with accumulated mutant tau, α-synuclein, or huntingtin. Treatment with Apt-1 restored proteostasis and inhibited cell death in a mouse model of proteinopathy induced by mutant tau(P301S). We conclude that pharmacological targeting of TRADD may represent a promising strategy for inhibiting cell death and restoring homeostasis to treat human diseases.
Assuntos
Apoptose/efeitos dos fármacos , Homeostase/efeitos dos fármacos , Proteína de Domínio de Morte Associada a Receptor de TNF/antagonistas & inibidores , Proteína de Domínio de Morte Associada a Receptor de TNF/metabolismo , Animais , Autofagia/efeitos dos fármacos , Proteína 3 com Repetições IAP de Baculovírus/metabolismo , Proteína Beclina-1/química , Proteína Beclina-1/metabolismo , Bortezomib/antagonistas & inibidores , Bortezomib/farmacologia , Linhagem Celular , Humanos , Proteína Huntingtina/metabolismo , Proteínas Inibidoras de Apoptose/metabolismo , Masculino , Camundongos , Modelos Moleculares , Emaranhados Neurofibrilares/metabolismo , Proteoma/metabolismo , Proteína Serina-Treonina Quinases de Interação com Receptores/química , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Proteína de Domínio de Morte Associada a Receptor de TNF/química , Proteína de Domínio de Morte Associada a Receptor de TNF/deficiência , Fator 2 Associado a Receptor de TNF/metabolismo , Ubiquitinação , alfa-Sinucleína/metabolismo , Proteínas tau/metabolismoRESUMO
Alzheimer's disease (AD) is characterized by the presence of tau protein inclusions and amyloid beta (Aß) plaques in the brain, with Aß peptides generated by cleavage of the amyloid precursor protein (APP) by BACE1 and γ-secretase. We previously described a primary rat neuron assay in which tau inclusions form from endogenous rat tau after seeding cells with insoluble tau isolated from the human AD brain. Here, we used this assay to screen an annotated library of â¼8700 biologically active small molecules for their ability to reduce immuno-stained neuronal tau inclusions. Compounds causing ≥30% inhibition of tau aggregates with <25% loss of DAPI-positive cell nuclei underwent further confirmation testing and assessment of neurotoxicity, and non-neurotoxic hits were subsequently analyzed for inhibitory activity in an orthogonal ELISA that quantified multimeric rat tau species. Of the 173 compounds meeting all criteria, a subset of 55 inhibitors underwent concentration-response testing and 46 elicited a concentration-dependent reduction of neuronal tau inclusions that were distinct from measures of toxicity. Among the confirmed inhibitors of tau pathology were BACE1 inhibitors, several of which, along with γ-secretase inhibitors/modulators, caused a concentration-dependent lowering of neuronal tau inclusions and a reduction of insoluble tau by immunoblotting, although they did not decrease soluble phosphorylated tau species. In conclusion, we have identified a diverse set of small molecules and related targets that reduce neuronal tau inclusions. Notably, these include BACE1 and γ-secretase inhibitors, suggesting that a cleavage product from a shared substrate, such as APP, might affect tau pathology.
Assuntos
Doença de Alzheimer , Secretases da Proteína Precursora do Amiloide , Neurônios , Proteínas tau , Animais , Humanos , Ratos , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Secretases da Proteína Precursora do Amiloide/metabolismo , Ácido Aspártico Endopeptidases/metabolismo , Neurônios/metabolismo , Proteínas tau/genética , Proteínas tau/metabolismoRESUMO
Inclusions comprised of microtubule-associated protein tau (tau) are implicated in a group of neurodegenerative diseases, collectively known as tauopathies, that include Alzheimer's disease (AD). The spreading of misfolded tau "seeds" along neuronal networks is thought to play a crucial role in the progression of tau pathology. Consequently, restricting the release or uptake of tau seeds may inhibit the spread of tau pathology and potentially halt the advancement of the disease. Previous studies have demonstrated that the Mammalian Suppressor of Tauopathy 2 (MSUT2), an RNA binding protein, modulates tau pathogenesis in a transgenic mouse model. In this study, we investigated the impact of MSUT2 on tau pathogenesis using tau seeding models. Our findings indicate that the loss of MSUT2 mitigates human tau seed-induced pathology in neuron cultures and mouse models. In addition, MSUT2 regulates many gene transcripts, including the Adenosine Receptor 1 (A1AR), and we show that down regulation or inhibition of A1AR modulates the activity of the "ArfGAP with SH3 Domain, Ankyrin Repeat, and PH Domain 1 protein" (ASAP1), thereby influencing the internalization of pathogenic tau seeds into neurons resulting in reduction of tau pathology.
Assuntos
Doença de Alzheimer , Tauopatias , Camundongos , Humanos , Animais , Encéfalo/patologia , Proteínas tau/metabolismo , Tauopatias/patologia , Doença de Alzheimer/patologia , Neurônios/patologia , Camundongos Transgênicos , Mamíferos/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismoRESUMO
TAR DNA-binding protein-43 (TDP-43) accumulation is the primary pathology underlying several neurodegenerative diseases. Charting the progression and heterogeneity of TDP-43 accumulation is necessary to better characterize TDP-43 proteinopathies, but current TDP-43 staging systems are heuristic and assume each syndrome is homogeneous. Here, we use data-driven disease progression modelling to derive a fine-grained empirical staging system for the classification and differentiation of frontotemporal lobar degeneration due to TDP-43 (FTLD-TDP, n = 126), amyotrophic lateral sclerosis (ALS, n = 141) and limbic-predominant age-related TDP-43 encephalopathy neuropathologic change (LATE-NC) with and without Alzheimer's disease (n = 304). The data-driven staging of ALS and FTLD-TDP complement and extend previously described human-defined staging schema for ALS and behavioural variant frontotemporal dementia. In LATE-NC individuals, progression along data-driven stages was positively associated with age, but negatively associated with age in individuals with FTLD-TDP. Using only regional TDP-43 severity, our data driven model distinguished individuals diagnosed with ALS, FTLD-TDP or LATE-NC with a cross-validated accuracy of 85.9%, with misclassifications associated with mixed pathological diagnosis, age and genetic mutations. Adding age and SuStaIn stage to this model increased accuracy to 92.3%. Our model differentiates LATE-NC from FTLD-TDP, though some overlap was observed between late-stage LATE-NC and early-stage FTLD-TDP. We further tested for the presence of subtypes with distinct regional TDP-43 progression patterns within each diagnostic group, identifying two distinct cortical-predominant and brainstem-predominant subtypes within FTLD-TDP and a further two subcortical-predominant and corticolimbic-predominant subtypes within ALS. The FTLD-TDP subtypes exhibited differing proportions of TDP-43 type, while there was a trend for age differing between ALS subtypes. Interestingly, a negative relationship between age and SuStaIn stage was seen in the brainstem/subcortical-predominant subtype of each proteinopathy. No subtypes were observed for the LATE-NC group, despite aggregating individuals with and without Alzheimer's disease and a larger sample size for this group. Overall, we provide an empirical pathological TDP-43 staging system for ALS, FTLD-TDP and LATE-NC, which yielded accurate classification. We further demonstrate that there is substantial heterogeneity amongst ALS and FTLD-TDP progression patterns that warrants further investigation in larger cross-cohort studies.
Assuntos
Doença de Alzheimer , Esclerose Lateral Amiotrófica , Demência Frontotemporal , Degeneração Lobar Frontotemporal , Proteinopatias TDP-43 , Humanos , Esclerose Lateral Amiotrófica/genética , Demência Frontotemporal/patologia , Doença de Alzheimer/patologia , Proteinopatias TDP-43/patologia , Degeneração Lobar Frontotemporal/patologia , Proteínas de Ligação a DNA/genéticaRESUMO
Pathologies that are causative for neurodegenerative disease (ND) are also frequently present in unimpaired, older individuals. In this retrospective study of 1647 autopsied individuals, we report the incidence of 10 pathologies across ND and normal ageing in attempt to clarify which pathological combinations are disease-associated and which are ageing-related. Eight clinically defined groups were examined including unimpaired individuals and those with clinical Alzheimer's disease, mixed dementia, amyotrophic lateral sclerosis, frontotemporal degeneration, multiple system atrophy, probable Lewy body disease or probable tauopathies. Up to seven pathologies were observed concurrently resulting in a heterogeneous mix of 161 pathological combinations. The presence of multiple additive pathologies associated with older age, increasing disease duration, APOE e4 allele and presence of dementia across the clinical groups. Fifteen to 67 combinations occurred in each group, with the unimpaired group defined by 35 combinations. Most combinations occurred at a <5% prevalence including 86 that were present in only one or two individuals. To better understand this heterogeneity, we organized the pathological combinations into five broad categories based on their age-related frequency: (i) 'Ageing only' for the unimpaired group combinations; (ii) 'ND only' if only the expected pathology for that individual's clinical phenotype was present; (iii) 'Other ND' if the expected pathology was not present; (iv) 'ND + ageing' if the expected pathology was present together with ageing-related pathologies at a similar prevalence as the unimpaired group; and (v) 'ND + associated' if the expected pathology was present together with other pathologies either not observed in the unimpaired group or observed at a greater frequency. ND only cases comprised a minority of cases (19-45%) except in the amyotrophic lateral sclerosis (56%) and multiple system atrophy (65%) groups. The ND + ageing category represented 9-28% of each group, but was rare in Alzheimer's disease (1%). ND + associated combinations were common in Alzheimer's disease (58%) and Lewy body disease (37%) and were observed in all groups. The Ageing only and Other ND categories accounted for a minority of individuals in each group. This observed heterogeneity indicates that the total pathological burden in ND is frequently more than a primary expected clinicopathological correlation with a high frequency of additional disease- or age-associated pathologies.
Assuntos
Doença de Alzheimer , Esclerose Lateral Amiotrófica , Doença por Corpos de Lewy , Atrofia de Múltiplos Sistemas , Humanos , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Doença por Corpos de Lewy/patologia , Esclerose Lateral Amiotrófica/patologia , Estudos RetrospectivosRESUMO
In Lewy body diseases-including Parkinson's disease, without or with dementia, dementia with Lewy bodies, and Alzheimer's disease with Lewy body co-pathology 1 -α-synuclein (α-Syn) aggregates in neurons as Lewy bodies and Lewy neurites 2 . By contrast, in multiple system atrophy α-Syn accumulates mainly in oligodendrocytes as glial cytoplasmic inclusions (GCIs) 3 . Here we report that pathological α-Syn in GCIs and Lewy bodies (GCI-α-Syn and LB-α-Syn, respectively) is conformationally and biologically distinct. GCI-α-Syn forms structures that are more compact and it is about 1,000-fold more potent than LB-α-Syn in seeding α-Syn aggregation, consistent with the highly aggressive nature of multiple system atrophy. GCI-α-Syn and LB-α-Syn show no cell-type preference in seeding α-Syn pathology, which raises the question of why they demonstrate different cell-type distributions in Lewy body disease versus multiple system atrophy. We found that oligodendrocytes but not neurons transform misfolded α-Syn into a GCI-like strain, highlighting the fact that distinct α-Syn strains are generated by different intracellular milieus. Moreover, GCI-α-Syn maintains its high seeding activity when propagated in neurons. Thus, α-Syn strains are determined by both misfolded seeds and intracellular environments.
Assuntos
Citoplasma/metabolismo , Corpos de Lewy/metabolismo , Corpos de Lewy/patologia , Doença por Corpos de Lewy/metabolismo , Doença por Corpos de Lewy/patologia , Neurônios/metabolismo , alfa-Sinucleína/classificação , alfa-Sinucleína/metabolismo , Animais , Citoplasma/química , Citoplasma/patologia , Feminino , Humanos , Corpos de Lewy/química , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/química , Neurônios/patologia , Oligodendroglia/química , Oligodendroglia/metabolismo , Oligodendroglia/patologia , Especificidade de Órgãos , Dobramento de Proteína , alfa-Sinucleína/químicaRESUMO
INTRODUCTION: Intraneuronal inclusions composed of tau protein are found in Alzheimer's disease (AD) and other tauopathies. Tau normally binds microtubules (MTs), and its disengagement from MTs and misfolding in AD is thought to result in MT abnormalities. We previously identified triazolopyrimidine-containing MT-stabilizing compounds that provided benefit in AD mouse models and herein describe the characterization and efficacy testing of an optimized candidate, CNDR-51997. METHODS: CNDR-51997 underwent pharmacokinetic, pharmacodynamic, safety pharmacology, and mouse tolerability testing. In addition, the compound was examined for efficacy in 5XFAD amyloid beta (Aß) plaque mice and PS19 tauopathy mice. RESULTS: CNDR-51997 significantly reduced Aß plaques in 5XFAD mice and tau pathology in PS19 mice, with the latter also showing attenuated axonal dystrophy and gliosis. CNDR-51997 was well tolerated at doses that exceeded efficacy doses, with a good safety pharmacology profile. DISCUSSION: CNDR-51997 may be a candidate for advancement as a potential therapeutic agent for AD and/or other tauopathies. Highlights There is evidence of microtubule alterations (MT) in Alzheimer's disease (AD) brain and in mouse models of AD pathology. Intermittent dosing with an optimized, brain-penetrant MT-stabilizing small-molecule, CNDR-51997, reduced both Aß plaque and tau inclusion pathology in established mouse models of AD. CNDR-51997 attenuated axonal dystrophy and gliosis in a tauopathy mouse model, with a strong trend toward reduced hippocampal neuron loss. CNDR-51997 is well tolerated in mice at doses that are meaningfully greater than required for efficacy in AD mouse models, and the compound has a good safety pharmacology profile.
Assuntos
Doença de Alzheimer , Modelos Animais de Doenças , Camundongos Transgênicos , Microtúbulos , Placa Amiloide , Proteínas tau , Animais , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/patologia , Camundongos , Placa Amiloide/tratamento farmacológico , Placa Amiloide/patologia , Proteínas tau/metabolismo , Microtúbulos/efeitos dos fármacos , Microtúbulos/metabolismo , Encéfalo/efeitos dos fármacos , Encéfalo/patologia , Encéfalo/metabolismo , Tauopatias/tratamento farmacológico , Tauopatias/patologia , Humanos , Moduladores de Tubulina/farmacologia , Moduladores de Tubulina/uso terapêutico , Peptídeos beta-Amiloides/metabolismoRESUMO
Effective therapies are urgently needed to safely target TDP-43 pathology as it is closely associated with the onset and development of devastating diseases such as frontotemporal lobar degeneration with TDP-43 pathology (FTLD-TDP) and amyotrophic lateral sclerosis (ALS). In addition, TDP-43 pathology is present as a co-pathology in other neurodegenerative diseases such as Alzheimer's disease and Parkinson's disease. Our approach is to develop a TDP-43-specific immunotherapy that exploits Fc gamma-mediated removal mechanisms to limit neuronal damage while maintaining physiological TDP-43 function. Thus, using both in vitro mechanistic studies in conjunction with the rNLS8 and CamKIIa inoculation mouse models of TDP-43 proteinopathy, we identified the key targeting domain in TDP-43 to accomplish these therapeutic objectives. Targeting the C-terminal domain of TDP-43 but not the RNA recognition motifs (RRM) reduces TDP-43 pathology and avoids neuronal loss in vivo. We demonstrate that this rescue is dependent on Fc receptor-mediated immune complex uptake by microglia. Furthermore, monoclonal antibody (mAb) treatment enhances phagocytic capacity of ALS patient-derived microglia, providing a mechanism to restore the compromised phagocytic function in ALS and FTD patients. Importantly, these beneficial effects are achieved while preserving physiological TDP-43 activity. Our findings demonstrate that a mAb targeting the C-terminal domain of TDP-43 limits pathology and neurotoxicity, enabling clearance of misfolded TDP-43 through microglia engagement, and supporting the clinical strategy to target TDP-43 by immunotherapy. SIGNIFICANCE STATEMENT: TDP-43 pathology is associated with various devastating neurodegenerative disorders with high unmet medical needs such as frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS) and Alzheimer's disease. Thus, safely and effectively targeting pathological TDP-43 represents a key paradigm for biotechnical research as currently there is little in clinical development. After years of research, we have determined that targeting the C-terminal domain of TDP-43 rescues multiple patho-mechanisms involved in disease progression in two animal models of FTD/ALS. In parallel, importantly, our studies establish that this approach does not alter the physiological functions of this ubiquitously expressed and indispensable protein. Together, our findings substantially contribute to the understanding of TDP-43 pathobiology and support the prioritization for clinical testing of immunotherapy approaches targeting TDP-43.
Assuntos
Doença de Alzheimer , Esclerose Lateral Amiotrófica , Demência Frontotemporal , Doença de Pick , Camundongos , Animais , Esclerose Lateral Amiotrófica/genética , Demência Frontotemporal/genética , Doença de Alzheimer/genética , Neuroproteção , Proteínas de Ligação a DNA/metabolismo , ImunoterapiaRESUMO
AIMS: Adult polyglucosan body disease (APBD) is a progressive neurogenetic disorder caused by 1,4-alpha-glucan branching enzyme 1 (GBE1) mutation with an accumulation of polyglucosan bodies (PBs) in the central and peripheral nervous systems as a pathological hallmark. Here, we report two siblings in a family with a GBE1 mutation with prominent frontotemporal lobar degeneration with TAR DNA-binding protein 43 (FTLD-TDP) and ageing-related tau astrogliopathy (ARTAG) copathologies with PBs in the central nervous system. METHODS: Whole-genome sequencing (WGS) followed by Sanger sequencing (SS) was performed on three affected and two unaffected siblings in a pedigree diagnosed with familial frontotemporal dementia. Out of the affected siblings, autopsies were conducted on two cases, and brain samples were used for biochemical and histological analyses. Brain sections were stained with haematoxylin and eosin and immunostained with antibodies against ubiquitin, tau, amyloid ß, α-synuclein, TDP-43 and fused in sarcoma (FUS). RESULTS: A novel single nucleotide deletion in GBE1, c.1280delG, was identified, which is predicted to result in a reading frameshift, p.Gly427Glufs*9. This variant segregated with disease in the family, is absent from population databases and is predicted to cause loss of function, a known genetic mechanism for APBD. The affected siblings showed a greater than 50% decrease in GBE protein levels. Immunohistochemical analysis revealed widespread FTLD-TDP (type A) and ARTAG pathologies as well as PBs in the brains of two affected siblings for whom an autopsy was performed. CONCLUSIONS: This is the first report of a family with several individuals with a FTD clinical phenotype and underlying copathologies of APBD, FTLD-TDP and ARTAG with a segregating GBE1 loss-of-function mutation in affected siblings. The finding of copathologies of APBD and FTLD-TDP suggests these processes may share a disease mechanism resulting from this GBE1 mutation.
Assuntos
Enzima Ramificadora de 1,4-alfa-Glucana , Demência Frontotemporal , Degeneração Lobar Frontotemporal , Sistema da Enzima Desramificadora do Glicogênio , Humanos , Demência Frontotemporal/patologia , Enzima Ramificadora de 1,4-alfa-Glucana/genética , Enzima Ramificadora de 1,4-alfa-Glucana/metabolismo , Peptídeos beta-Amiloides/metabolismo , Degeneração Lobar Frontotemporal/patologia , Encéfalo/patologia , Mutação , Proteínas de Ligação a DNA/metabolismo , Proteínas tau/metabolismo , Sistema da Enzima Desramificadora do Glicogênio/genética , Sistema da Enzima Desramificadora do Glicogênio/metabolismoRESUMO
Loss of midbrain dopamine neurons causes the cardinal symptoms of Parkinson's disease. However, not all dopamine neurons are equally vulnerable and a better understanding of the cell-type specific properties relating to selective dopamine neuron degeneration is needed. Most midbrain dopamine neurons express the vesicular glutamate transporter VGLUT2 during development and a subset continue to express low levels of VGLUT2 in adulthood, enabling the co-release of glutamate. Moreover, VGLUT2 expression in dopamine neurons can be neuroprotective since its genetic disruption was shown to sensitize dopamine neurons to neurotoxins. Here, we show that in response to toxic insult, and in two distinct models of alpha-synuclein stress, VGLUT2 dopamine neurons were resilient to degeneration. Dopamine neurons expressing VGLUT2 were enriched whether or not insult induced dopamine neuron loss, suggesting that while VGLUT2 dopamine neurons are more resilient, VGLUT2 expression can also be transcriptionally upregulated by injury. Finally, we observed that VGLUT2 expression was enhanced in surviving dopamine neurons from post-mortem Parkinson's disease individuals. These data indicate that emergence of a glutamatergic identity in dopamine neurons may be part of a neuroprotective response in Parkinson's disease.
Assuntos
Neurônios Dopaminérgicos , Doença de Parkinson , Adulto , Dopamina/metabolismo , Neurônios Dopaminérgicos/metabolismo , Humanos , Mesencéfalo , Degeneração Neural/metabolismo , Doença de Parkinson/metabolismo , Proteína Vesicular 2 de Transporte de Glutamato/genética , Proteína Vesicular 2 de Transporte de Glutamato/metabolismoRESUMO
AIMS: It has long been considered that accumulation of pathological alpha-synuclein (aSyn) leads to synaptic/neuronal loss which then results in behavioural and cognitive dysfunction. To investigate this claim, we investigated effects downstream of aSyn preformed fibrils (PFFs) and 6-hydroxydopamine (6-OHDA), because aSyn PFFs induce spreading/accumulation of aSyn, and 6-OHDA rapidly causes local neuronal loss. METHODS: We injected mouse aSyn PFFs into the medial forebrain bundle (MFB) of Sprague-Dawley rats. We investigated spread of pathological aSyn, phosphorylation of aSyn and tau, oxidative stress, synaptic/neuronal loss and cognitive dysfunction 60, 90 and 120 days after injection. Similarly, we injected 6-OHDA into the MFB and examined the same parameters 1 and 3 weeks after injection. RESULTS: Following aSyn PFF injection, phosphorylated aSyn was found distant from the injection site in the hippocampus and frontal cortex. However, despite neuron loss being evident close to the site of injection in the substantia nigra at 120 days post injection, there were no other neurodegeneration-associated features associated with aSyn including synaptic loss. In contrast, 6-OHDA caused severe neuronal loss in the substantia nigra at 3 weeks post injection that was accompanied by phosphorylation of aSyn and tau, oxidative stress, loss of synaptic proteins, cognitive and motor dysfunction. CONCLUSIONS: Our results demonstrate that spread/replication and slow accumulation of pathological aSyn may not be sufficient to induce neurodegenerative changes. In contrast, oxidative stress responses in addition to aSyn accumulation were associated with other Parkinson's disease (PD)-associated abnormalities and cognitive dysfunction. Our results may be important when considering why only some PD patients develop dementia.
Assuntos
Doença de Parkinson , alfa-Sinucleína , Animais , Camundongos , Oxidopamina/metabolismo , Oxidopamina/farmacologia , Doença de Parkinson/patologia , Ratos , Ratos Sprague-Dawley , Substância Negra/patologia , alfa-Sinucleína/metabolismoRESUMO
Limbic-predominant age-related TDP-43 encephalopathy (LATE) is characterized by the accumulation of TAR-DNA-binding protein 43 (TDP-43) aggregates in older adults. LATE coexists with Lewy body disease (LBD) as well as other neuropathological changes including Alzheimer's disease (AD). We aimed to identify the pathological, clinical, and genetic characteristics of LATE in LBD (LATE-LBD) by comparing it with LATE in AD (LATE-AD), LATE with mixed pathology of LBD and AD (LATE-LBD + AD), and LATE alone (Pure LATE). We analyzed four cohorts of autopsy-confirmed LBD (n = 313), AD (n = 282), LBD + AD (n = 355), and aging (n = 111). We assessed the association of LATE with patient profiles including LBD subtype and AD neuropathologic change (ADNC). We studied the morphological and distributional differences between LATE-LBD and LATE-AD. By frequency analysis, we staged LATE-LBD and examined the association with cognitive impairment and genetic risk factors. Demographic analysis showed LATE associated with age in all four cohorts and the frequency of LATE was the highest in LBD + AD followed by AD, LBD, and Aging. LBD subtype and ADNC associated with LATE in LBD or AD but not in LBD + AD. Pathological analysis revealed that the hippocampal distribution of LATE was different between LATE-LBD and LATE-AD: neuronal cytoplasmic inclusions were more frequent in cornu ammonis 3 (CA3) in LATE-LBD compared to LATE-AD and abundant fine neurites composed of C-terminal truncated TDP-43 were found mainly in CA2 to subiculum in LATE-LBD, which were not as numerous in LATE-AD. Some of these fine neurites colocalized with phosphorylated α-synuclein. LATE-LBD staging showed LATE neuropathological changes spread in the dentate gyrus and brainstem earlier than in LATE-AD. The presence and prevalence of LATE in LBD associated with cognitive impairment independent of either LBD subtype or ADNC; LATE-LBD stage also associated with the genetic risk variants of TMEM106B rs1990622 and GRN rs5848. These data highlight clinicopathological and genetic features of LATE-LBD.
Assuntos
Envelhecimento/patologia , Encéfalo/patologia , Doença por Corpos de Lewy/patologia , Proteinopatias TDP-43/patologia , Idoso , Idoso de 80 Anos ou mais , Doença de Alzheimer/complicações , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Feminino , Humanos , Doença por Corpos de Lewy/complicações , Doença por Corpos de Lewy/genética , Masculino , Pessoa de Meia-Idade , Proteinopatias TDP-43/complicações , Proteinopatias TDP-43/genéticaRESUMO
Cerebral amyloid angiopathy (CAA), limbic-predominant age-related TDP-43 encephalopathy neuropathological change (LATE-NC) and Lewy bodies occur in the absence of clinical and neuropathological Alzheimer's disease, but their prevalence and severity dramatically increase in Alzheimer's disease. To investigate how plaques, tangles, age and apolipoprotein E ε4 (APOE ε4) interact with co-pathologies in Alzheimer's disease, we analysed 522 participants ≥50 years of age with and without dementia from the Center for Neurodegenerative Disease Research (CNDR) autopsy program and 1340 participants in the National Alzheimer's Coordinating Center (NACC) database. Consensus criteria were applied for Alzheimer's disease using amyloid phase and Braak stage. Co-pathology was staged for CAA (neocortical, allocortical, and subcortical), LATE-NC (amygdala, hippocampal, and cortical), and Lewy bodies (brainstem, limbic, neocortical, and amygdala predominant). APOE genotype was determined for all CNDR participants. Ordinal logistic regression was performed to quantify the effect of independent variables on the odds of having a higher stage after checking the proportional odds assumption. We found that without dementia, increasing age associated with all pathologies including CAA (odds ratio 1.63, 95% confidence interval 1.38-1.94, P < 0.01), LATE-NC (1.48, 1.16-1.88, P < 0.01), and Lewy bodies (1.45, 1.15-1.83, P < 0.01), but APOE ε4 only associated with CAA (4.80, 2.16-10.68, P < 0.01). With dementia, increasing age associated with LATE-NC (1.30, 1.15-1.46, P < 0.01), while Lewy bodies associated with younger ages (0.90, 0.81-1.00, P = 0.04), and APOE ε4 only associated with CAA (2.36, 1.52-3.65, P < 0.01). A longer disease course only associated with LATE-NC (1.06, 1.01-1.11, P = 0.01). Dementia in the NACC cohort associated with the second and third stages of CAA (2.23, 1.50-3.30, P < 0.01), LATE-NC (5.24, 3.11-8.83, P < 0.01), and Lewy bodies (2.41, 1.51-3.84, P < 0.01). Pathologically, increased Braak stage associated with CAA (5.07, 2.77-9.28, P < 0.01), LATE-NC (5.54, 2.33-13.15, P < 0.01), and Lewy bodies (4.76, 2.07-10.95, P < 0.01). Increased amyloid phase associated with CAA (2.27, 1.07-4.80, P = 0.03) and Lewy bodies (6.09, 1.66-22.33, P = 0.01). In summary, we describe widespread distributions of CAA, LATE-NC and Lewy bodies that progressively accumulate alongside plaques and tangles in Alzheimer's disease dementia. CAA interacted with plaques and tangles especially in APOE ε4 positive individuals; LATE-NC associated with tangles later in the disease course; most Lewy bodies associated with moderate to severe plaques and tangles.
Assuntos
Doença de Alzheimer/epidemiologia , Doença de Alzheimer/patologia , Angiopatia Amiloide Cerebral/epidemiologia , Doença por Corpos de Lewy/epidemiologia , Proteinopatias TDP-43/epidemiologia , Idoso , Idoso de 80 Anos ou mais , Comorbidade , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos RetrospectivosRESUMO
The hallmark pathological features of Alzheimer's disease (AD) brains are senile plaques, comprising ß-amyloid (Aß) peptides, and neuronal inclusions formed from tau protein. These plaques form 10-20 years before AD symptom onset, whereas robust tau pathology is more closely associated with symptoms and correlates with cognitive status. This temporal sequence of AD pathology development, coupled with repeated clinical failures of Aß-directed drugs, suggests that molecules that reduce tau inclusions have therapeutic potential. Few tau-directed drugs are presently in clinical testing, in part because of the difficulty in identifying molecules that reduce tau inclusions. We describe here two cell-based assays of tau inclusion formation that we employed to screen for compounds that inhibit tau pathology: a HEK293 cell-based tau overexpression assay, and a primary rat cortical neuron assay with physiological tau expression. Screening a collection of â¼3500 pharmaceutical compounds with the HEK293 cell tau aggregation assay, we obtained only a low number of hit compounds. Moreover, these compounds generally failed to inhibit tau inclusion formation in the cortical neuron assay. We then screened the Prestwick library of mostly approved drugs in the cortical neuron assay, leading to the identification of a greater number of tau inclusion inhibitors. These included four dopamine D2 receptor antagonists, with D2 receptors having previously been suggested to regulate tau inclusions in a Caenorhabditis elegans model. These results suggest that neurons, the cells most affected by tau pathology in AD, are very suitable for screening for tau inclusion inhibitors.
Assuntos
Agregados Proteicos/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Proteínas tau/metabolismo , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Animais , Antagonistas dos Receptores de Dopamina D2/química , Antagonistas dos Receptores de Dopamina D2/metabolismo , Antagonistas dos Receptores de Dopamina D2/farmacologia , Células HEK293 , Humanos , Camundongos , Microscopia de Fluorescência , Neurônios/citologia , Neurônios/metabolismo , Ratos , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/metabolismo , Proteínas tau/antagonistas & inibidores , Proteínas tau/genéticaRESUMO
AIM: The heterogeneity in the distribution and morphological features of TAR DNA-binding protein-43 (TDP-43) pathology in the brains of frontotemporal lobar degeneration (FTLD-TDP) patients and their different clinical manifestations suggest that distinct pathological TDP-43 strains could play a role in this heterogeneity between different FTLD-TDP subtypes (A-E). Our aim was to evaluate the existence of distinct TDP-43 strains in the brains of different FTLD-TDP subtypes and characterise their specific seeding properties in vitro and in vivo. METHODS AND RESULTS: We used an inducible stable cell line expressing a mutant cytoplasmic TDP-43 (iGFP-NLSm) to evaluate the seeding properties of distinct pathological TDP-43 strains. Brain-derived TDP-43 protein extracts from FTLD-TDP types A (n = 6) and B (n = 3) cases induced the formation of round/spherical phosphorylated TDP-43 aggregates that morphologically differed from the linear and wavy wisps and bigger heterogeneous filamentous (skein-like) aggregates induced by type E (n = 3) cases. These morphological differences correlated with distinct biochemical banding patterns of sarkosyl-insoluble TDP-43 protein recovered from the transduced cells. Moreover, brain-derived TDP-43 extracts from type E cases showed higher susceptibility to PK digestion of full-length TDP-43 and the most abundant C-terminal fragments that characterise type E extracts. Finally, we showed that intracerebral injections of different TDP-43 strains induced a distinctive morphological and subcellular distribution of TDP-43 pathology and different spreading patterns in the brains of CamKIIa-hTDP-43NLSm Tg mice. CONCLUSIONS: We show the existence of distinct TDP-43 strains in the brain of different FTLD-TDP subtypes with distinctive seeding and spreading properties in the brains of experimental animal models.
Assuntos
Encéfalo/metabolismo , Encéfalo/patologia , Proteínas de Ligação a DNA/metabolismo , Animais , Proteínas de Ligação a DNA/genética , Demência Frontotemporal/metabolismo , Demência Frontotemporal/patologia , Degeneração Lobar Frontotemporal/patologia , Humanos , Camundongos TransgênicosRESUMO
The neurodegenerative diseases amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with TAR DNA-binding protein-43 (TDP-43) inclusions (FTLD-TDP) share the neuropathological hallmark of aggregates of TDP-43. However, factors governing the severity and regional distribution of TDP-43 pathology, which may account for the divergent clinical presentations of ALS and FTLD-TDP, are not well understood. Here, we investigated the influence of genotypes at TMEM106B, a locus associated with risk for FTLD-TDP, and hexanucleotide repeat expansions in C9orf72, a known genetic cause for both ALS and FTLD-TDP, on global TDP-43 pathology and regional distribution of TDP-43 pathology in 899 postmortem cases from a spectrum of neurodegenerative diseases. We found that, among the 110 ALS cases, minor (C)-allele homozygotes at the TMEM106B locus sentinel SNP rs1990622 had more TDP-43 pathology globally, as well as in select brain regions. C9orf72 expansions similarly associated with greater TDP-43 pathology in ALS. However, adjusting for C9orf72 expansion status did not affect the relationship between TMEM106B genotype and TDP-43 pathology. To elucidate the direction of causality for this association, we directly manipulated TMEM106B levels in an inducible cell system that expresses mislocalized TDP-43 protein. We found that partial knockdown of TMEM106B, to levels similar to what would be expected in rs1990622 C allele carriers, led to development of more TDP-43 cytoplasmic aggregates, which were more insoluble, in this system. Taken together, our results support a causal role for TMEM106B in modifying the development of TDP-43 proteinopathy.