The following dissertation discusses the role of Aβ42 dependent hyperactivation of AMPK mediating synaptic loss through coordinated Mff-dependent mitochondrial fission and Ulk2-dpendent mitophagy in dendrites of PNs. In Chapter 1, I provide a brief background on Alzheimer’s disease and the cellular and molecular mechanisms that have been relevant to the pathogenesis of the disease including disruption on mitochondrial homeostasis and autophagy. In Chapter 2, I discuss the findings of my main project describing the role of Aβ42 induced mitochondrial remodeling leading to synapse loss in vitro and in vivo in part by hyperactivation of CAMKKII-AMPK. Chapter 3 covers a review article that I participated in in examining the role of mitochondria in various ND. In Chapter 4, I discuss about a project I was involved in in examining the mechanism behind maintaining mitochondrial morphology in axon versus dendrite and its functional consequence. In Chapter 5, I end the dissertation by highlighting key findings, potential future studies, and concluding remarks.

No description available.

No description available.

This thesis contributes to recent interest within medical imaging regarding the development and clinical application of magnetic resonance elastography (MRE) to the human brain. MRE is a non-invasive phase-contrast MRI technique for measurement of brain mechanical properties in vivo, shown to reflect the composition and organisation of the complex tissue microstructure. MRE is a promising imaging biomarker for the early characterisation of neurodegeneration due to its exquisite sensitivity to variation among healthy and pathological tissue. Neurodegenerative diseases are debilitating conditions of the human nervous system for which there is currently no cure. Novel biomarkers are required to improve early detection, differential diagnosis and monitoring of disease progression, and could also ultimately improve our understanding of the pathophysiological mechanisms underlying degenerative processes. This thesis begins with a theoretical background of brain MRE and a description of the experimental considerations. A systematic review of the literature is then performed to summarise brain MRE quantitative measurements in healthy participants and to determine the success of MRE to characterise neurological disorders. This review further identified the most promising acquisition and analysis methods within the field. As such, subsequent visits to three brain MRE research centres, within the USA and Germany, enabled the acquisition of exemplar phantom and brain data to assist in discussions to refine an experimental protocol for installation at the Edinburgh Imaging Facility, QMRI (EIF-QMRI). Through collaborations with world-leading brain MRE centres, two high-resolution - yet fundamentally different - MRE pipelines were installed at the EIF-QMRI. Several optimisations were implemented to improve MRE image quality, while the clinical utility of MRE was enhanced by the novel development of a Graphical User Interface (GUI) for the optimised and automatic MRE-toanatomical coregistration and generation of MRE derived output measures. The first experimental study was performed in 6 young and 6 older healthy adults to compare the results from the two MRE pipelines to investigate test-retest agreement of the whole brain and a brain structure of interest: the hippocampal formation. The MRE protocol shown to possess superior reproducibility was subsequently applied in a second experimental study of 12 young and 12 older cognitively healthy adults. Results include finding that the MRE imaging procedure is very well tolerated across the recruited population. Novel findings include significantly softer brains in older adults both across the global cerebrum and in the majority of subcortical grey matter structures including the pallidum, putamen, caudate, and thalamus. Changes in tissue stiffness likely reflect an alteration to the strength in the composition of the tissue network. All MRE effects persist after correcting for brain structure volume suggesting changes in volume alone were not reflective of the detected MRE age differences. Interestingly, no age-related differences to tissue stiffness were found for the amygdala or hippocampus. As for brain viscosity, no group differences were detected for either the brain globally or subcortical structures, suggesting a preservation of the organisation of the tissue network in older age. The third experiment performed in this thesis finds a direct structure-function relationship in older adults between hippocampal viscosity and episodic memory as measured with verbal-paired recall. The source of this association was located to the left hippocampus, thus complementing previous literature suggesting unilateral hippocampal specialisation. Additionally, a more significant relationship was found between left hippocampal viscosity and memory after a new procedure was developed to remove voxels containing cerebrospinal fluid from the MRE analysis. Collectively, these results support the transition of brain MRE into a clinically useful neuroimaging modality that could, in particular, be used in the early characterisation of memory specific disorders such as amnestic Mild Cognitive Impairment and Alzheimer's disease.

Dementia affects around 850,000 people in the UK. Whilst there is no cure, understanding how to support people to live well with this condition is important. There are 'many faces of dementia' and delineating experiences of living with different dementias in daily life is a key step towards understanding how to support people to live well with different symptoms. Supporting eating-related practices in a socio-relational context, referred to in this thesis as 'dining' experiences, may help promote the psychological needs related to living well with dementia, including inclusion, occupation and identity. This study aimed to understand how dining experiences and interactions were affected when living with typical dementia (tAD) and posterior cortical atrophy (PCA) and the processes involved in supporting dining experiences. Data collection included in-depth interviews with 20 family dyads (9 tAD; 11 PCA) and video-based observations of four dining interactions. An 'informed' grounded theory approach was used and Goffman's (1974) Frame Analysis was identified to support understanding dining experiences in this study. Goffman's (1974) theory provides understanding around how people structure and define their social experiences and coordinate these understandings in interaction. Findings revealed how dining could become disrupted when living with tAD and PCA, relating to vulnerabilities in this experience and eating-related practices becoming more about 'management'. Interacting processes of revising frames, management strategies, optimising opportunities for continuity and engaging in supportive environments, were identified to support maintaining-meaning in dining. Video-based observations extended understanding in terms of contextual factors which shaped experiences and how people used the environment to support their dining interactions. Implications include supporting dining by using a balanced system of various processes to continue to create new meanings and experiences and using the environment to make 'dining' central, as opposed to focusing on eating-related practices as a 'care-task'.

La Protéine Prion cellulaire (PrPc) est une protéine ubiquitaire mais majoritairement présente dans le système nerveux central. Elle est plus particulièrement connue pour sa conversion conformationnelle en PrPSc dans les maladies à Prions qui sont des Protéinopathies comme la maladie d’Alzheimer (MA). La MA est en partie associée à des dépôts de peptides beta-amyloïdes (Aβ) agrégés de façon extracellulaire et issus des clivages successifs par la β- puis la γ-sécrétase de la protéine précurseur amyloïde (APP) exprimée dans les neurones. La PrPc et l’APP partagent des fonctions et des voies protéolytiques communes (α- ou β-sécrétase) les impliquant dans la prolifération, la différenciation, la synaptogenèse et la survie cellulaire. La PrPc est impliquée dans la régulation de la prolifération et la différenciation de différentes cellules souches : neurales adultes (NSC), hématopoïétiques (HSC), embryonnaires humaines (hESC). Si la PrP et l’APP partagent des fonctions communes, plusieurs publications montrent que la PrPc régule négativement le clivage de l'APP en Aβ et positivement le clivage de l’APP en sAPPα suggérant ainsi un rôle anti-amyloïdogénique de la PrPc. La PrP agirait également comme récepteur des Aβ à la surface neuronale induisant notamment l’inhibition des LTP et l’altération synaptique.Dans ce contexte, les objectifs spécifiques de la thèse sont :- L’étude de l’expression de la PrP, de l’APP et ses résidus de clivage au cours de l’induction neurale des hESC en NSC et de la différenciation neuronale- L’impact de la modulation de l’expression de la PrP sur le clivage de l’APP ainsi que sur les propriétés des cellules souches (survie, prolifération, différenciation).1. Induction neurale des hESC en NSC Pour ce projet nous avons utilisé des Cellules Souches Embryonnaires Humaines (hESC) pour lesquelles le laboratoire dispose d’une autorisation de l’Agence de la Biomédecine.Pour l’induction neurale, nous avons testé deux protocoles : l’un permet d’obtenir des neurosphères en suspension puis des «rosettes» constituées de NSC, l’autre protocole en monocouche mime quant à lui la corticogenèse. Une optimisation de ces protocoles a été nécessaire (densité de départ, méthodes de fixation des cellules pour améliorer la détection de la PrP) ainsi que la détermination des conditions d’analyse de l’expression de PrP, d’APP et ses résidus clivés (Aβ, sAPPα/β). 2. Différenciations à partir des NSC Les NSC obtenues ont ensuite été amplifiées puis différenciées en neurones et/ou astrocytes. Les cellules ont été caractérisées notamment par immunofluorescence et RT-qPCR pour l’expression des principaux marqueurs astrocytaires (GFAP) et neuronaux (BIII-tubuline, Doublecortine, Synaptophysine) et la disparition progressive des marqueurs de NSC. Là encore nous avons établi des conditions précises de densité cellulaire ainsi que les points des analyses cinétiques de nos différents paramètres.3. Modulation de l’expression de la PrPc Nous avons utilisés des vecteurs lentiviraux permettant l’expression ou l’inhibition de la PrPc humaine pour transduire des hESC au moment d’initier l’induction neurale et des NSC. Pour cela nous avons également dû réaliser des optimisations de différents paramètres : densité cellulaire, taille des supports d’ensemencement ou MOI de lentivirus afin d’avoir une transduction efficace tout en limitant la cytotoxicité. De même, les échantillons récoltés nous ont permis d’évaluer l’impact de la modulation de la PrPc sur le clivage de l’APP ainsi que sur la biologie des cellules souches (survie, prolifération, différenciation). / The cellular Prion Protein (PrPc) is a ubiquitary protein mainly expressed in the central nervous system. It is particularly known for its conformational conversion in PrPSc in Prion diseases, which are proteinopathies such as Alzheimer’s disease (AD). AD is associated with extracellular deposits of aggregated beta-amyloid peptides (Aβ) derived from successive β- and the γ-secretase cleavages of the amyloid precursor protein (APP) expressed by neurons. PrPc and APP share some common functions and proteolytic pathways (α- or β-secretase), involving them in proliferation, differentiation, synaptogenesis and cellular survival. PrPc is involved in the regulation of proliferation and differentiation of many stem cells: adult neural (NSC), hematopoietic (HSC) and human embryonic (hESC). Several publications also show that PrP downregulates the cleavage of APP in Aβ and positively regulates the cleavage of APP in sAPPα suggesting an anti-amyloïdogenic role of PrPc. PrP could also act as a receptor of Aβ at the neuronal surface inducing LTP inhibition and synaptic alteration. In this context, the specific objectives of my thesis were:- Study of the expression of PrP, APP and its cleavage residues during neural induction of hESC in NSC and neuronal differentiation.- Impact of the modulation of PrP expression on APP cleavages as well as on stem cells properties (survival, proliferation, differentiation). 1. Neural induction of hESC in NSCFor this project, we have used Human Embryonic Stem Cells (hESC) for which the laboratory has an authorization from the “Agence de la Biomédecine”.For the neural induction, we have tested two protocols, the first one allows the obtention of neurospheres in suspension and then figures of “rosettes” composed of NSC, and a “monolayer” protocol that mimics the beginning of corticogenesis. An optimization of these protocols has been necessary (starting cell density, cell fixation methods to improve PrP detection). We have also determined the best conditions to analyze the expression of PrP, APP and its derived peptides (Aß, sAPPα/β). 2. Differentiation of NSCNSC derived from hESC were amplified and differentiated into neurons and/or astrocytes. Cells were characterized in particular by immunofluorescence and RT-qPCR for the expression of the major astrocytic (GFAP) and neuronal markers (BIII-tubulin, doublecortin, synaptophysin) and the progressive decrease of NSC markers. Again we have determined the best conditions for cell density and kinetic time points for our analysis.3. Modulation of PrPC expression We have used lentiviral vectors allowing the expression of an anti-PrP shRNA, human PrP and respective controls. To achieve this task, lentiviral transductions of hESC and NSC were optimized: cell density, size of the seeding culture wells or MOI of lentivirus. Finaly, samples collected allowed us to evaluate the impact of PrPc modulation on the APP cleavages as well as on stem cells properties (survival, proliferation, differentiation).

A doença de Alzheimer (AD) é a doença neurodegenerativa relacionada ao envelhecimento mais frequente no mundo. Uma das características moleculares de AD é a produção exacerbada de peptídeos beta-amiloide (Aβ), principalmente dos fragmentos de 40 e 42 aminoácidos (Aβ1-40 e Aβ1-42). Aβ induz respostas neuroinflamatórias e alterações moleculares relacionadas à perda sináptica e morte neuronal. Diversos relatos mostram que pacientes de AD apresentam redução na concentração plasmática de melatonina, hormônio produzido pela glândula pineal e também alteração na expressão dos receptores de melatonina, mas os mecanismos envolvidos ainda não são conhecidos. De acordo com o conceito do eixo imunepineal, agentes inflamatórios são capazes de atuar diretamente sobre a glândula pineal e inibir a síntese de melatonina. No presente estudo investigamos, portanto, se o peptídeo Aβ atua diretamente sobre o sistema melatonérgico, modulando a síntese de melatonina ou a função de seus receptores. Pineais em cultura tratadas com Aβ 1-40 ou Aβ 1-42 apresentaram redução na produção de melatonina. Aβ 1-40 ativou a via do fator de transcrição NF-κB na pineal, resultando em aumento da transcrição de diversos genes inflamatórios, como interleucinas e quimiocinas, e inibição da expressão da enzima arilalquilamina N-acetiltransferase, essencial à síntese de melatonina. Em células HEK293 expressando estavelmente receptores MT1 ou MT2 recombinantes, a ativação da via ERK1/2 pela melatonina foi inibida tanto por Aβ 1-40 quanto por Aβ 1-42. O mesmo efeito inibitório foi observado em células endoteliais primárias que expressam MT1 e MT2 constitutivamente. O presente trabalho mostra que a síntese de melatonina pela pineal e a função dos receptores de melatonina são diretamente regulados por Aβ, o que amplia nossos conhecimentos a respeito dos efeitos prejudiciais de Aβ. Considerando que a melatonina tem propriedades neuroprotetora e antioxidante, a disfunção do sistema melatonérgico pode contribuir para os processos neurodegenerativos que ocorrem na patologia de AD / Alzheimer\'s disease (AD) is the most common age-related neurodegenerative disorder worldwide. Excess of amyloid beta peptides (Aβ), composed mainly by 40 and 42 aminoacids-long fragments (Aβ 1-40 e Aβ 1-42) is a molecular hallmark in AD. Aβ-induced neuroinflammatory responses and molecular changes are related to synapse impairment and neuronal loss. It is well documented that AD patients show impaired melatonin synthesis, the pineal gland-derived hormone, and altered expression of melatonin receptors, but the underlying mechanisms remain unclear. According to the immune-pineal axis concept, inflammatory mediators act on the pineal gland, leading to inhibition of melatonin synthesis. Therefore, in the present study we sought to investigate whether Aβ? directly targets the melatonergic system, modulating melatonin synthesis and/or melatonin receptors function. Pineal glands cultured in the presence of Aβ 1-40 or Aβ 1-42 showed reduced melatonin production. Aβ 1-40 activated the nuclear factor kappa B (NF-κ B) pathway in the pineal gland, leading to up-regulation of several inflammatory genes, as interleukins and chemokines, and inhibition of the arylalkylamine N-acetyltransferase enzyme expression, the key enzyme in melatonin synthesis. In HEK293 cells stably expressing recombinant melatonin MT1 or MT2 receptors melatonin-induced ERK1/2 activation was markedly impaired by Aβ 1-40 and Aβ 1-42. Similar results were obtained in primary culture of endothelial cells expressing melatonin receptors endogenously. The present study shows that melatonin synthesis and melatonin receptors function are directly impaired by Aβ, thus extending our understanding on the detrimental effects of Aβ. Because melatonin shows neuroprotective and antioxidant properties, impairment of the melatonergic system may contribute to the neurodegenerative processes that take place in AD. 1-42) is a molecular hallmark in AD. Aβ-induced neuroinflammatory responses and molecular changes are related to synapse impairment and neuronal loss. It is well documented that AD patients show impaired melatonin synthesis, the pineal gland-derived hormone, and altered expression of melatonin receptors, but the underlying mechanisms remain unclear. According to the immune-pineal axis concept, inflammatory mediators act on the pineal gland, leading to inhibition of melatonin synthesis. Therefore, in the present study we sought to investigate whether Aβ?directly targets the melatonergic system, modulating melatonin synthesis and/or melatonin receptors function. Pineal glands cultured in the presence of Aβ 1-40 or Aβ 1-42 showed reduced melatonin production. Aβ 1-40 activated the nuclear factor kappa B (NF-&kappa B) pathway in the pineal gland, leading to up-regulation of several inflammatory genes, as interleukins and chemokines, and inhibition of the arylalkylamine N-acetyltransferase enzyme expression, the key enzyme in melatonin synthesis. In HEK293 cells stably expressing recombinant melatonin MT1 or MT2 receptors melatonin-induced ERK1/2 activation was markedly impaired by Aβ 1-40 and Aβ 1-42. Similar results were obtained in primary culture of endothelial cells expressing melatonin receptors endogenously. The present study shows that melatonin synthesis and melatonin receptors function are directly impaired by Aβ, thus extending our understanding on the detrimental effects of Aβ. Because melatonin shows neuroprotective and antioxidant properties, impairment of the melatonergic system may contribute to the neurodegenerative processes that take place in AD

Alzheimer's disease (AD) is characterised by memory loss, insidious cognitive decline, profound neurodegeneration, and the extracellular accumulation of amyloid-beta (Aβ) peptide in senile plaques and intracellular accumulation of tau in neurofibrillary tangles. Synaptic dysfunction and loss is the strongest pathological correlate of cognitive decline in AD with increasing evidence implicating neuropathological forms of both amyloid-beta and tau protein in this process. A large amount of evidence suggests that oligomeric forms of Aβ, associated with senile plaques, are toxic to synapses but the precise localisation of Aβ and which forms are synaptotoxic remain unknown. Using the high-resolution technique, array tomography, this thesis characterised the synaptic localisation of different forms of Aβ oligomers in a mouse model of amyloidopathy. These results show that different oligomeric Aβ species are present in both presynapses and postsynapses. This study highlights the potential of array tomography for rapid testing of aggregation state specific Aβ antibodies in brain tissue. Following these results, the presence of tau at synapses was examined. Despite the knowledge that tau spreads through defined synaptic circuits, it is currently unknown whether synapse loss occurs before the accumulation of tau or as a consequence. To address this, array tomography was used to examine a mouse model in which mutant P301L human tau is expressed primarily in the entorhinal cortex (rTgTauEC). It has previously been shown that rTgTauEC mice exhibit neuronal loss in the entorhinal cortex and synapse density loss in the middle molecular layer (MML) of the dentate gyrus at 24 months of age. The density of tau-expressing and total presynapses, and the spread of tau into the postsynapse in the MML of 3-6, 9, and 18 month old mice were examined. No loss of synapse density was observed in the MML up to 18 months of age, even in axons expressing tau. Despite the maintenance of synapse density, we see spread of human tau from presynaptic terminals to postsynaptic compartments in the MML at very early ages. This indicates that the spread of tau through neural circuits is not due to the degeneration of axon terminals and is an early feature of the disease process. Following examination of both synaptic amyloid-beta and tau in separate models, this thesis then examined how these two proteins may be synergistically working together to drive synaptic pathology. To investigate this a novel mouse model was used in which amyloid-beta deposits are present in combination with non-mutated human tau expression (APP/PS1 + hTau). These results suggested that the addition of human tau expression does not increase plaque associated synapse loss, neither does it increase the proportion of synapses colocalising with amyloid-beta. Similarly the presence of human tau at individual postsynapses was not enhanced in the presence of oligomeric Aβ. Surprisingly, intact long-term recognition memory was observed in APP/PS1 + hTau mice. However a hyperactive phenotype was detected in these mice that could be prevented upon tau suppression. This suggests a synergistic relationship may exist in the presentation of this phenotype. Finally in the last part of this thesis, synapses from post-mortem human Alzheimer's disease and age-matched controls were investigated. It has previously been suggested that both amyloid-beta and tau can interfere with mitochondrial transport to the synapse and mitochondrial function. For this reason the presence of synaptic mitochondria at both the presynapse and postsynapse was determined in order to investigate any alteration in the diseased state. A reduction in the proportion of presynapses with multiple mitochondria present was detected in anterior/posterior transverse temporal cortex (BA41/42). This was not observed in dorsolateral prefrontal cortex (BA46), suggesting either a selective vulnerability of the former brain region or a selective resistance of the latter brain region, to mitochondrial depletion at the synapse. The findings presented in this thesis demonstrate that when investigated in isolation, pathological forms of amyloid-beta are present at a subset of synapses where they may contribute to toxicity, whilst the spread of tau protein is an early feature of the disease process and occurs prior to overt synapse loss. This thesis also explores the proposed synergistic relationship between amyloid-beta and tau using a novel mouse model and human post-mortem brain tissue. Since these two proteins both have been implicated in synaptic dysfunction, investigating Aβ and tau in new mouse models and human brain tissue will be instrumental in furthering our understanding of mechanisms and features of synaptotoxicity that could be important therapeutic targets.

No description available.

Dans cette thèse, nous explorons de nouvelles méthodes d’analyse d’images pour la détection précoce des changements métaboliques cérébraux causés par la maladie d’Alzheimer. Nous introduisons deux apports méthodologiques que nous appliquons à un ensemble de données réelles. Le premier est basé sur l’apprentissage automatique afin de créer une carte des informations pertinentes pour la classification d'un ensemble d’images. Pour cela nous échantillonnons des blocs de Voxels selon un algorithme de Monte-Carlo. La mise en œuvre d’une classification basée sur ces patchs 3d a pour conséquence la réduction significative du volume de patchs à traiter et l’extraction de caractéristiques dont l’importance est statistiquement quantifiable. Cette méthode s’applique à différentes caractéristiques et est adaptée à des types d’images variés. La résolution des cartes produites par cette méthode peut être affinée à volonté et leur contenu informatif est cohérent avec des résultats antérieurs obtenus dans la littérature. Le second apport méthodologique porte sur la conception d’un nouvel algorithme de décomposition de tenseur d’ordre important, adapté à notre application. Cet algorithme permet de réduire considérablement la consommation de mémoire et donc en évite la surcharge. Il autorise la décomposition rapide de tenseurs, y compris ceux de dimensions très déséquilibrées. Nous appliquons cet algorithme en tant que méthode d’extraction de caractéristiques dans une situation où le clinicien doit diagnostiquer des stades précoces de la maladie d'Alzheimer en utilisant la TEP-FDG seule. Les taux de classification obtenus sont souvent au-dessus des niveaux de l’état de l’art. / In this thesis we want to explore novel image analysis methods for the early detection of metabolic changes in the human brain caused by Alzheimer's disease (AD). We will present two methodological contributions and present their application to a real life data set. We present a machine learning based method to create a map of local distribution of classification relevant information in an image set. The presented method can be applied using different image characteristics which makes it possible to adapt the method to many kinds of images. The maps generated by this method are very localized and fully consistent with prior findings based on Voxel wise statistics. Further we preset an algorithm to draw a sample of patches according to a distribution presented by means of a map. Implementing a patch based classification procedure using the presented algorithm for data reduction we were able to significantly reduce the amount of patches that has to be analyzed in order to obtain good classification results. We present a novel non-negative tensor factorization (NTF) algorithm for the decomposition of large higher order tensors. This algorithm considerably reduces memory consumption and avoids memory overhead. This allows the fast decomposition even of tensors with very unbalanced dimensions. We apply this algorithm as feature extraction method in a computer-aided diagnosis (CAD) scheme, designed to recognize early-stage ad and mild cognitive impairment (MCI) using fluorodeoxyglucose (FDG) positron emission tomography (PET) scans only. We achieve state of the art classification rates.