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Thesis
In the Injured Retina, Microrna, Mir-18a, Regulates the Microglia Response and Smad2-mediated Pathways
University of West Florida Libraries
Master of Science (MS), University of West Florida
Summer 2022
Abstract
In humans, when photoreceptors are damaged or injured recovery of these neuronal cells is not possible and can lead to vision problems and even blindness. Photoreceptors are the retinal neurons that start the visual process and the first in line to translate a light signal into a chemical signal (Zhang et al 2021). Human photoreceptors are never regenerated or replaced. In contrast, zebrafish are able to restore and regenerate photoreceptors after injury to the retina (Hitchcock et al 2004; Bernardos et al 2007). This research focuses on identifying molecular mechanisms that are involved in this process. Müller Glia, which act as stem cells in the zebrafish retina, are the initial players that have been shown to divide and produce neural progenitors that proliferate and differentiate into those new photoreceptors. Although the specific processes that control photoreceptor regeneration are still being studied, recent studies have shown that different microRNAs (miRNAs) may play important roles. Previous research has shown that a miRNA, miR-18a, regulates photoreceptor differentiation in the embryonic retina and that it also regulates neuronal progenitor proliferation in the injured retina by regulating inflammation (Taylor et al 2019; Magner 2020). The current research focuses on further understanding miR-18a function in the regenerative process of photoreceptors in the retina in mature zebrafish after injury. Taqman RT-qPCR quantitatively showed that miR-18a is expressed higher at 3, and 5 days post-lesion (dpl), the peak time when Müller glia and MG-derived progenitors are dividing. Immunohistochemistry using bromodeoxyuridine (BrdU) labeling of dividing cells showed that, after injury at 3dpl in miR-18a mutant retinas, cell proliferation does not begin earlier or occur faster, but that proliferation of the progenitors is prolonged. TUNEL and BrdU labeling showed that excess progenitors in the miR-18a mutant retina do not die by apoptosis, suggesting that they differentiate into other cells and stay in the INL or migrate to other layers. With these results, ethynyldeoxyuridine (EdU), labeling of dividing cells and immunolabeling of specific differentiated cells with different antibodies were used to determine if the excess progenitor cells differentiate into microglia, amacrine cells, and Müller glia. Results showed that only a small percentage of the progenitor cells differentiate into these cells. With previous data suggesting that miR-18a is involved in regulation of inflammation, immunohistochemistry was used to label microglia at different time points after injury showing that the microglia response is prolonged in miR-18a mutants. Lastly, western blots and RT-qPCR showed that pSmad2 protein is reduced but RNA expression levels remain unaffected in miR-18a mutant retinas, indicating that miR-18a might regulate the TGFB pathway. Together, this evidence shows that in the injured zebrafish retina, cell proliferation is altered by disruption of miR-18a, and miR-18a affects inflammation by prolonging the microglia response.
Details
- Title
- In the Injured Retina, Microrna, Mir-18a, Regulates the Microglia Response and Smad2-mediated Pathways
- Resource Type
- Thesis
- Contributors
- Scott M Taylor (Advisor)Peter Cavnar (Committee Member)Youngil Lee (Committee Member)
- Publisher
- University of West Florida Libraries
- Format
- pdf
- Number of pages
- 54
- Copyright
- Permission granted to the University of West Florida Libraries to digitize and/or display this item for non-profit research and educational purposes. Any reuse of this item in excess of fair use or other copyright exemptions requires the permission of the copyright holder.
- Identifiers
- 99380092796706600
- Academic Unit
- Biology; Hal Marcus College of Science and Engineering
- Language
- English
- Awarding Institution
- University of West Florida; Master of Science (MS)
- Theses and Dissertations
- Master of Science (MS), University of West Florida