dc.contributor.advisor |
Connor, Bronwen |
|
dc.contributor.advisor |
McCaughey-Chapman, Amy |
|
dc.contributor.author |
McShane, Emma |
|
dc.date.accessioned |
2022-04-11T01:18:11Z |
|
dc.date.available |
2022-04-11T01:18:11Z |
|
dc.date.issued |
2022 |
en |
dc.identifier.uri |
https://hdl.handle.net/2292/58647 |
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dc.description |
Full Text is available to authenticated members of The University of Auckland only. |
en |
dc.description.abstract |
Much of the pathological mechanisms underpinning the neurodevelopmental disorder fragile X syndrome (FXS) remains unknown due to the limited access and availability of developmentally relevant human cell models. The bone morphogenic protein receptor 2 (BMPR2) has been identified as a potential translational target of fragile X mental retardation protein (FMRP) and has been implicated in the altered development of dendritic spines via the BMPR2-LIMK1-Cofilin axis. LIMK1 and cofilin are also essential components regulating neurite outgrowth, a phenotype significantly dysregulated in FXS. We aimed to investigate the effect of LIMK1 inhibition on neurite outgrowth in neurons generated via reprogramming of control and FXS donor fibroblasts into human induced neuronal precursor cells (hiNPs) and differentiated into cortical neurons.
Utilising a novel cell reprogramming protocol developed by the Connor group at the Centre for Brain Research, healthy and FXS donor fibroblasts were reprogrammed via chemically modified mRNA (cmRNA) transfection into hiNPs, and differentiated into cortical neurons. During differentiation, hiNPs were treated with DMSO vehicle, or 84nM of LIMK1 inhibitor, TH 257. Evaluation of the effects of TH 257 were conducted by quantitative real-time PCR analysis, and Western blotting. Finally, neurite outgrowth between disease phenotypes and treatment groups were evaluated by neurite length measurement.
Validation of our FXS neuron model revealed no significant difference in FMR1 or FMRP expression when compared to healthy controls. Treatment of cells with TH 257 did not alter transcription profiles, nor have a significant effect on protein expression or phosphorylation. No significant difference was observed between FXS and control neurons neurite outgrowth. TH 257 showed no significant effect on neurite length.
The characterisation of the disease genotype and phenotype of our FXS neuron model outlined potential considerations on the effect of reprogramming on the methylation status of FXS-affected cells, and the implications of FXS mosaicism on disease modelling. We did not observe any effects of TH 257 on neurite outgrowth in FXS pathology. Further evaluation is needed, with controlling for the effect of FXS mosaic models, greater case numbers, and more accurate assessment mechanisms to determine if the effects of LIMK1 inhibition observed in FXS animal models translate to humans. |
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dc.publisher |
ResearchSpace@Auckland |
en |
dc.relation.ispartof |
Masters Thesis - University of Auckland |
en |
dc.relation.isreferencedby |
UoA |
en |
dc.rights |
Restricted Item. Full Text is available to authenticated members of The University of Auckland only. |
en |
dc.rights |
Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. |
|
dc.rights.uri |
https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htm |
en |
dc.rights.uri |
http://creativecommons.org/licenses/by-nc-sa/3.0/nz/ |
|
dc.title |
The Effect of LIMK1 Inhibition on Neurite Outgrowth in Fragile X Syndrome |
|
dc.type |
Thesis |
en |
thesis.degree.discipline |
Biomedical Science |
|
thesis.degree.grantor |
The University of Auckland |
en |
thesis.degree.level |
Masters |
en |
dc.date.updated |
2022-03-28T03:19:40Z |
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dc.rights.holder |
Copyright: the author |
en |