Disruption of the fronto-subcortical neuronal circuits of the brain is not only common in the 30% of those who develop dementia after stroke but other ageing-related dementias. We previously demonstrated region- specific pyramidal cell atrophy within the dorsolateral prefrontal cortex (dlPFC) as a strong substrate for dementia after stroke. This occurred irrespective of any neurodegenerative pathology, indicating that vascular changes drive frontal lobe dysfunction across common dementias. Analysis of single neurons is key to identifying major cellular pathways contributing to dementia. The DTP student will identify neuronal proteomic profiles of individual dlPFC neurons in patients who develop dementia after stroke. Using state-of-the-art quantitative morphometry and novel proteomic analysis the project will allow identification of molecular pathways of neurodegeneration. Student will first analyse post-mortem brains from MRC-funded prospectively assessed cohorts of elderly stroke survivors collected by the primary supervisor. Detailed clinical examination, imaging records and neuropsychological screening have been performed for >90 cases of non-demented and demented post-stroke survivors. A particular strength is that subjects had mostly small cortical strokes and were initially recruited free of dementia 3-months post-stroke for longitudinal follow-up. 3D-Stereological analyses in combination with imaging mass cytometry (IMC) in remote infarct-free regions will be undertaken to identify cellular size characteristics of dlPFC and medial PFC pyramidal neurons. Tissues from stroke-free controls plus those who developed Alzheimer type or mixed dementia will enable comparison of disease specific mechanisms. Laser-capture microdissection (LCM) techniques will be used to isolate single neurons and subject them to proteomic analysis achieved by iTRAQ methods in collaboration with Singapore linked to the primary supervisor’s site. Identification of differentially regulated neuronal proteins in combination with bioinformatics at the second supervisor’s site will allow identification of novel cell-molecular pathways associated with cell atrophy in ageing and dementia. Blood samples are also available for analysis to use for diagnostic profiling in tandem with the proteomic analysis.
DTP student will master range of timely techniques for cell-molecular investigations of ageing-related dementia including 1) Histopathological analysis of brain and cardiac tissues. Routine tinctorial staining methods to assess large vessel and microvascular pathology, 2) Immunocytochemistry, immunofluorescence methods for antigen localisation, 3) Light, fluorescent, confocal and dual-photon microscopy techniques, 4) Imaging mass cytometry (ICM) to assess multiple antigens, a technique used in mass spectrometry to visualize the spatial distribution of chemical compositions e.g. compounds, biomarkers, metabolites, peptides or proteins by their molecular masses. Production of metal tagged antibodies and interpretation, 5) Laser-capture microdissection and related protein methods, 6) 2-D gel electrophoresis and related immunochemical methods, 7) Proteomic methods for analysis of LCM samples and microvessel fractions, 8) Bioinformatics plus related protein data mining to evaluate cell signalling pathways and 9) Relevant software use and computer assisted data processing and 10) Developing good record keeping and writing skills. Both the primary and secondary supervisors offer a rich environment which is already provide wide training to other students both at MRes/MSci as well as PhD levels. Prof Newman Sze (Singapore) and Dr Dr Mark Dunning (Sheffield) will provide guidance on proteomic analysis and bioinformatics.
Benefits of being in the DiMeN DTP:
This project is part of the Discovery Medicine North Doctoral Training Partnership (DiMeN DTP), a diverse community of PhD students across the North of England researching the major health problems facing the world today. Our partner institutions (Universities of Leeds, Liverpool, Newcastle, York and Sheffield) are internationally recognised as centres of research excellence and can offer you access to state-of the-art facilities to deliver high impact research.
We are very proud of our student-centred ethos and committed to supporting you throughout your PhD. As part of the DTP, we offer bespoke training in key skills sought after in early career researchers, as well as opportunities to broaden your career horizons in a range of non-academic sectors.
Being funded by the MRC means you can access additional funding for research placements, international training opportunities or internships in science policy, science communication and beyond. See how our current DiMeN students have benefited from this funding here: http://www.dimen.org.uk/overview/student-profiles/flexible-supplement-awards
Further information on the programme and how to apply can be found on our website:
http://www.dimen.org.uk/how-to-apply/application-overview
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