Perineuronal nets as a barrier for a-synuclein spreading

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Project Summary:Parkinson's disease (PD) is a progressive neurodegenerative disorder which is characterised by the presence of Lewy bodies, made up of aggregated misfolded a-synuclein (a-syn) amongst other proteins, in neurones causing cell death. a-syn can be transferred between neurones leading to Lewy body spread. a-syn misfolding and deposition also contributes to other synucleinopathies. Perineuronal nets (PNNs) are a type of extracellular matrix (ECM) found on the surface of neurones. PNNs modulate synapse formation, control neuroplasticity, act as a protective layer against toxic proteins and oxidative stress, serve as an ion reservoir and sequester molecules on neurones. PNNs are made up of hyaluronan (a glycosaminoglycan), chondroitin sulphate proteoglycans, HAPLNs (link proteins) and Tenascin-R (a glycoprotein). Current data shows that wildtype a-syn binds to chondroitin sulphates in rat neuronal cultures. Additionally, characterisation in human brains shows that PNN neurones are spared from Lewy pathology, whilst non-PNN neurones are not.The aims of this project are to (1), characterise the different a-syn species and their structures, retrieved from human PD samples. a-syn isolated from tissue samples will be analysed using fluorescent probe labelling, atomic force microscope and/or cryo-electron microscopy. (2), Determine the interactions between chondroitin sulphates, PNNs and different a-syn species identified in (1). This will be determined through expressing and purifying proteins, oligomer/ fibril preparation and using biophysical techniques to characterise these molecular interactions. (3), Determine the impact of the interactions between a-syn, chondroitin sulphates, PNNs on: a-syn internalisation and seeding, in addition to the impact on neuronal morphology and physiology. This will be determined using neuronal culture, immunocytochemistry, confocal microscopy and quantification, and electrophysiology.This research will improve our understanding of the fundamental molecular mechanisms by which a-syn interacts with PNNs and how PNNs may offer protection to neurones against a-syn related pathologies.