Investigation of molecular and cellular mechanisms underpinning the neurotoxicity of homocysteine and its metabolites in models of neurodegeneration
Year: 2018
Type: dissertation
Abstract: Elevated levels of homocysteine (HCy) are a known risk factor in several disease states (1). \nHCy has several other metabolites, homocysteine thiolactone (HCy-T) and homocysteic acid \n(HCA). Whilst HCy-mediated neurotoxicity has been extensively studied, the underlying \nmechanisms of HCy-T and HCA mediated neuronal damage remain largely unknown. This \nthesis aims to explore the underlying mechanisms, triggered by HCy and metabolites which \nresult in neuronal cell death, and may be appropriate targets for future research on disease-modifying interventions in neurodegenerative disorders. As ageing is the greatest risk factor for neurodegeneration, a novel model of human neuronal ageing was established, permitting \ninvestigation of the pathways triggered by HCy in ageing. \nUsing SH-SY5Y cells, a novel differentiation protocol was established and categorised, once \nfully differentiated, these cells were shown to be fully functional neurons and could be \nmaintained for a month in culture. Using a range of concentrations of HCy and HCy-T, the \nconcentration cell death occurs at was determined using crystal violet and lactate \ndehydrogenase assays. Mechanisms of toxicity were determined using pharmacological \nintervention at the NMDA receptor, nitric oxide scavengers and antioxidants. Using a \ncombination of immunocytochemistry, live cell imaging and ELISA, alterations in markers of \ncell damage could be examined. \nThe results showed HCy and HCy-T have distinct mechanisms of toxicity. Whilst both are \nneurotoxic, HCy directly acts via the NMDA receptor, however HCy-T appears to be less \npotent. Additionally, HCy-T caused a greater increase in reactive oxygen species generation \nthan HCy, and each metabolite also displayed distinct mitochondrial network abnormalities. \nFinally, using the long-term culture methods, the chronic effects of HCy, HCy-T and HCA \nwere examined. However, extensive cell death was apparent at low doses in all metabolites therefore no definitive mechanisms could be determined. This culture method was deemed not \nappropriate for toxicity experiments.
Author: Lisa Strother
Language: English
Subfield: Nutrition and Dietetics
Field: Nursing
Domain: Health Sciences
Open Access status: green
OpenAlex ID: W2937955185