Pathogénie des dégénérescences neurofibrillaires de la maladie d'Alzheimer

La maladie d’Alzheimer (MA) est caractérisée par deux lésions neuropathologiques: les plaques séniles (composées essentiellement du peptide amyloïde Ab) et les dégénérescences neurofibrillaires (DNF, composées de formes hyperphosphorylées de protéines tau). Les mécanismes de formation des DNF sont encore mal compris et notre travail expérimental a eu pour objectifs d’étudier certaines hypothèses de formation des DNF.1° Une hypothèse étiopathogénique de la MA est la “cascade amyloïde”, selon laquelle le peptide amyloïde Ab exercerait un effet toxique entraînant la phosphorylation de tau et la formation de DNF. Certaines formes familiales de MA sont dues à des mutations du gène du précurseur du peptide amyloïde (APP) ou des présénilines et nous avons voulu déterminer si la surexpression de ces protéines pouvait entraîner la formation de DNF. Nous avons d’abord étudié une lignée murine double transgénique surexprimant l’isoforme 0N3R de protéine tau humaine “sauvage” et une forme mutée de préseniline 1 (M146L). Nous y avons démontré une co-expression neuronale des deux protéines et une augmentation de la phosphorylation de tau mais nous n’y avons pas observé de formation de DNF, chez des animaux examinés jusqu’à 17 mois. Nous avons ensuite étudié une lignée murine triple transgénique surexprimant l’isoforme 0N3R de protéine tau “sauvage”, une forme mutée de préseniline 1 (M146L) et une forme mutée de l’APP 751 (mutations Swedish K670N, M671L et London V717I). Ces animaux ont développé précocement (2.5 mois) des dépôts extracellulaires de peptide Ab. Nous y avons observé une augmentation de la phosphorylation de tau dans les prolongements neuronaux en contact avec les dépôts amyloïdes et des anomalies de l’organisation du cytosquelette, mais pas de DNF, chez des animaux examinés jusqu’à 18 mois.2° Certaines mutations du gène de tau sont responsables de formes familiales de démence frontotemporales dans lesquelles se développent des DNF. Ces mutations favoriseraient l’agrégation de tau où entraîneraient un déséquilibre de l’expression relative des isoformes de tau. Un tel déséquilibre pourrait également être induit dans les formes sporadiques de MA, en l’absence de mutations de tau. Afin d’investiguer cette hypothèse, nous avons étudié le profil d’expression des ARNm de tau et des isoformes de protéines tau dans plusieurs régions cérébrales de sujets contrôles ou atteints de MA. Un même profil d’expression a été observé dans les deux groupes. Une augmentation relative de l’expression de l’isoforme 0N3R de tau dans le cortex temporal pourrait être liée à la sensibilité de cette région au développement de DNF. Nous avons également étudié des lignées stables de cellules CHO exprimant des formes mutées (P301L, R406W) et non-mutées de protéines tau. Nous n’avons cependant pas observé d’augmentation de l’agrégation de tau dans les lignées exprimant les formes mutées de tau.Nos résultats indiquent que la simple surexpression de formes mutées de l’APP et des présénilines, même en présence d’une protéine tau humaine, ne suffit pas à entraîner la formation de DNF. En outre, l’absence de différence dans le profil d’expression cérébrale des isoformes de tau entre sujets contrôles et atteints de MA suggère que les modifications post-traductionnelles de cette protéine jouent un rôle plus important dans la genèse des DNF.Doctorat en sciences biomédicalesinfo:eu-repo/semantics/nonPublishe

The Innate Immune System in Alzheimer’s Disease

Alzheimer’s disease (AD) is the leading cause for dementia in the world. It is characterized by two biochemically distinct types of protein aggregates: amyloid β (Aβ) peptide in the forms of parenchymal amyloid plaques and congophilic amyloid angiopathy (CAA) and aggregated tau protein in the form of intraneuronal neurofibrillary tangles (NFT). Several risk factors have been discovered that are associated with AD. The most well-known genetic risk factor for late-onset AD is apolipoprotein E4 (ApoE4) (Potter and Wisniewski (2012), and Verghese et al. (2011)). Recently, it has been reported by two groups independently that a rare functional variant (R47H) of TREM2 is associated with the late-onset risk of AD. TREM2 is expressed on myeloid cells including microglia, macrophages, and dendritic cells, as well as osteoclasts. Microglia are a major part of the innate immune system in the CNS and are also involved in stimulating adaptive immunity. Microglia express several Toll-like receptors (TLRs) and are the resident macrophages of the central nervous system (CNS). In this review, we will focus on the recent advances regarding the role of TREM2, as well as the effects of TLRs 4 and 9 on AD

The active form of glycogen synthase kinase-3beta is associated with granulovacuolar degeneration in neurons in Alzheimer's disease.

Glycogen synthase kinase-3beta (GSK-3beta) is a physiological kinase for tau and is a candidate protein kinase involved in the hyperphosphorylation of tau present in paired helical filament (PHF)-tau of neurofibrillary tangles (NFT) in Alzheimer's disease (AD). GSK-3beta is also a key element of several signaling cascades (including cell death cascades). We have investigated the immunocytochemical localization of GSK-3 immunoreactivity in AD. Neurons exhibiting strongly GSK-3-immunoreactive granules were observed in AD, with a much higher frequency than in control subjects. This immunoreactivity was found to co-localize with the granulovacuolar degeneration (GVD) and to be associated with the granules of the granulovacuolar bodies. The GVD granules showed a strong GSK-3alpha and GSK-3beta immunoreactivity, and this immunoreactivity was abolished by preabsorption with recombinant GSK-3. In addition, the GVD immunoreactivity was observed with an antibody against the tyrosine-phosphorylated and active form of GSK-3. Some granules of the granulovacuolar degeneration were also intensely labeled with an antibody specific for tau isoforms containing insert 1 (exon 2) and with antibodies specific for tau phosphorylated on Ser262 and for tau phosphorylated on Thr212/Ser214, two phosphorylation sites generated in vitro by GSK-3alpha and beta. GSK-3beta was expressed in neurons containing NFT but only a small proportion of intracellular NFT were observed to be GSK-3beta immunoreactive. Immunoblotting analysis of fractions enriched in PHF-tau did not reveal any GSK-3beta immunoreactivity in these fractions, indicating that GSK-3beta was only loosely associated to NFT. These results suggest that neurons developing GVD sequester an active, potentially deleterious, form of GSK-3 in this compartment and that increased GSK-3 immunoreactivity in a subset of neurons quantitatively differentiates normal aging from AD.In VitroJournal ArticleResearch Support, Non-U.S. Gov'tinfo:eu-repo/semantics/publishe

Characterisation of cytoskeletal abnormalities in mice transgenic for wild-type human tau and familial Alzheimer's disease mutants of APP and presenilin-1.

To study the role of Abeta amyloid deposits in the generation of cytoskeletal lesions, we have generated a transgenic mouse line coexpressing in the same neurons a wild-type human tau isoform (0N3R), a mutant form of APP (751SL) and a mutant form of PS1 (M146L). These mice developed early cerebral extracellular deposits of Abeta, starting at 2.5 months. A somatodendritic neuronal accumulation of transgenic tau protein was observed in tau only and in tau/PS1/APP transgenic mice, including in neurons adjacent to Abeta deposits. The phosphorylation status of this somatodendritic tau was similar in the two transgenic lines. The Abeta deposits were surrounded by a neuritic reaction composed of axonal dystrophic processes, immunoreactive for many phosphotau epitopes and for the human tau transgenic protein. Ultrastructural observation showed in these dystrophic neurites a disorganisation of the microtubule and the neurofilament network but animals that were observed up to 18 months of age did not develop neurofibrillary tangles. These results indicate that overexpression of mutant PS1, mutant APP and of wild-type human tau were not sufficient per se to drive the formation of neurofibrillary tangles in a transgenic model. The Abeta deposits, however, were associated to marked changes in cytoskeletal organisation and in tau phosphorylation in adjacent dystrophic neurites.Journal ArticleResearch Support, Non-U.S. Gov'tinfo:eu-repo/semantics/publishe

Mutant presenilin 1 proteins induce cell death and reduce tau-dependent processes outgrowth.

The expression of familial Alzheimer's disease mutants of presenilin-1 (PS1) proteins has been observed to induce cell death in cellular systems. To investigate how this phenomenon might be associated to alterations of the microtubule network, we have studied the effect of wild-type and mutant (C263R, P264L and delta9) PS1 proteins expression on the formation of microtubule-dependent processes outgrowth and the association of PS1 to the insoluble cytoskeletal fraction in a cell line expressing the tau microtubule-associated protein. Expression of wild-type and mutant PS1 was associated with increased cell death, most marked for the P264L and delta9 mutants. The three PS1 mutants induced a significant reduction of the length of cell processes. These effects were not associated to a change in tau phosphorylation. However, the mutant PS1 proteins increased the proportion of insoluble tau in the cytoskeletal fraction and they were concentrated in the same fraction. These results suggest that PS1 proteins interact with the microtubule network, affect its organization and that this phenomenon, more marked for the PS1 mutants, might play a role in the cell dysfunction induced by mutant PS1 proteins.Comparative StudyJournal ArticleResearch Support, Non-U.S. Gov'tinfo:eu-repo/semantics/publishe

Increased tau phosphorylation but absence of formation of neurofibrillary tangles in mice double transgenic for human tau and Alzheimer mutant (M146L) presenilin-1.

Neurofibrillary tangles, composed of tau proteins, are a key lesion observed in sporadic forms of Alzheimer's disease and in familial forms associated with mutations of presenilin-1 (PS1). We have generated a double transgenic mouse line expressing a human tau isoform and a mutated form of PS1 (M146L) in neurons. Increased expression of the PS1 holoprotein was observed in the tau/PS1 transgenic mice and the proteolytic fragments of PS1 did not appear to be modified. A somatodendritic accumulation of the transgenic tau and an increase in tau phosphorylation were observed in both tau- and tau/PS1 transgenic mice. Neurofibrillary tangles were not observed in animals analyzed up to 17 months. Immunoprecipitation of tau from brain homogenates demonstrated its binding with active glycogen synthase kinase-3beta in control, tau- and tau/PS1 transgenic lines. These results suggest that overexpression of this Alzheimer mutant PS1 in vivo is not by itself sufficient to induce the formation of neurofibrillary tangles, even in neurons co-expressing and accumulating a human tau isoform.Journal ArticleResearch Support, Non-U.S. Gov'tinfo:eu-repo/semantics/publishe

Anti-β-sheet conformation monoclonal antibody reduces tau and Aβ oligomer pathology in an Alzheimer’s disease model

Abstract Background Oligomeric forms of amyloid-β (Aβ) and tau are increasing being recognized as key toxins in the pathogenesis of Alzheimer’s disease (AD). Methods We developed a novel monoclonal antibody (mAb), GW-23B7, that recognizes β-sheet secondary structure on pathological oligomers of neurodegenerative diseases. Results The pentameric immunoglobulin M kappa chain (IgMκp) we developed specifically distinguishes intra- and extracellular pathology in human AD brains. Purified GW-23B7 showed a dissociation constant in the nanomolar range for oligomeric Aβ and did not bind monomeric Aβ. In enzyme-linked immunosorbent assays, it recognized oligomeric forms of both Aβ and hyperphosphorylated tau. Aged triple-transgenic AD mice with both Aβ and tau pathology infused intraperitoneally for 2 months showed IgMκp in the soluble brain homogenate, peaking at 24 h postinoculation. Treated mice exhibited significant cognitive rescue on radial arm maze testing compared with vehicle control-infused mice. Immunohistochemically, treatment resulted in a significant decrease of extracellular pathology. Biochemically, treatment resulted in significant reductions of oligomeric forms of Aβ and tau. Conclusions These results suggest that GW-23B7, an anti-β-sheet conformational mAb humanized for clinical trials, may be an effective therapeutic agent for human AD

Vaccination of Alzheimer's model mice with A? derivative in alum adjuvant reduces A? burden without microhemorrhages

Immunotherapy holds great promise for Alzheimer's disease (AD) and other conformational disorders but certain adverse reactions need to be overcome. The meningoencephalitis observed in the first AD vaccination trial was likely related to excessive cell-mediated immunity caused by the immunogen, amyloid-? (A?) 1–42, and the adjuvant, QS?21. To avoid this toxicity, we have been using A? derivatives in alum adjuvant that promotes humoral immunity. Other potential side effects of immunotherapy are increased vascular amyloid and associated microhemorrhages that may be related to rapid clearance of parenchymal amyloid. Here, we determined if our immunization strategy was associated with this form of toxicity, and if the therapeutic effect was age-dependent. Tg2576 mice and wild-type littermates were immunized from 11 or 19 months and their behaviour evaluated prior to killing at 24 months. Subsequently, plaque- and vascular-A? burden, A? levels and associated pathology was assessed. The therapy started at the cusp of amyloidosis reduced cortical A? deposit burden by 31% and A? levels by 30–37%, which was associated with cognitive improvements. In contrast, treatment from 19 months, when pathology is well established, was not immunogenic and therefore did not reduce A? burden or improve cognition. Significantly, the immunotherapy in the 11–24 months treatment group, that reduced A? burden, did not increase cerebral bleeding or vascular A? deposits in contrast to several A? antibody studies. These findings indicate that our approach age-dependently improves cognition and reduces A? burden when used with an adjuvant suitable for humans, without increasing vascular A? deposits or microhemorrhages