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Investigating the role of micro RNAs in retinal development and as agents of degeneration

Holder, Daniel; (2021) Investigating the role of micro RNAs in retinal development and as agents of degeneration. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Micro RNAs (miRNAs) are potent post-transcriptional regulators of gene expression, which play a myriad of roles throughout human development and are key regulators of retinal development, as well as being implicated in retinal disease. The exact roles played by miRNAs in these processes are imperfectly understood. The miR-182, 96, 183 Sensory Cluster is a sensory organ-specific miRNA family and the most highly expressed miRNA family in the murine retina, yet the developmental roles it plays remain unclear. Whilst miRNA dysregulation is associated with certain retinopathies, whether dysregulation is a disease marker or plays a causative role in photoreceptor death, is unknown. This thesis investigated miRNA expression and function in human retinal development and the genetic retinopathy Type I Usher syndrome. Human pluripotent stem cell-derived retinal organoids provided a human tissue model for this study. A CRISPR-Cas9 gene editing platform was applied to modify the genome in hPSCs and investigate the effect on organoid development; Sensory Cluster knockout hESC lines and both patient and isogenic control TypeI Usher patient-derived iPSC lines were generated and analysed. Sensory Cluster function was interrogated using a gain and loss-of-function approach; over expression by miRNA mimic treatment was shown to lead to an increase in expression of certain photoreceptor maturation markers; Sensory Cluster knockout organoids were analysed using morphological and transcriptomic analyses. The molecular phenotype of Type I Usher in vitro was also interrogated using RNAseq. Type I Usher patient organoids displayed reduced expression of photoreceptor-associated genes, including the Sensory Cluster, but these findings were not recapitulated in organoids generated from a wider panel of Type I hPSC lines. These studies provide insight into the role of the Sensory Cluster in the human retina and the value of gene edited hPSCs to analyse human gene function. It also highlighted the heterogeneity between organoid differentiations and hPSC lines.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Investigating the role of micro RNAs in retinal development and as agents of degeneration
Event: UCL (University College London)
Language: English
Additional information: Copyright © The Author 2021. Original content in this thesis is licensed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0) Licence (https://creativecommons.org/licenses/by-nc/4.0/). Any third-party copyright material present remains the property of its respective owner(s) and is licensed under its existing terms. Access may initially be restricted at the author’s request.
Keywords: retina, eye, stem cell, organoid, miRNA
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL GOS Institute of Child Health
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL GOS Institute of Child Health > Developmental Biology and Cancer Dept
URI: https://discovery.ucl.ac.uk/id/eprint/10133462
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