Investigating the roles of HuR during wound healing

• Main Author: Quah, Clarissa Bernice
• Other Authors: -
• Format: Final Year Project
• Language: English
• Published: Nanyang Technological University 2020
• Subjects: Science::Biological sciences
• Online Access: https://hdl.handle.net/10356/138628
• Institution: Nanyang Technological University

Chronic wounds represent a universally devastating medical issue, with an estimated 6.7 million Americans afflicted among others worldwide and an annual healthcare expenditure of over $50 billion. Because of their concomitance with one or more prolonged wound healing phases, precise synchronization of gene and protein expression must be prioritized to warrant the concertation of multiple highly regulated factors central to tissue repair. The most apparent indicator of non-healing wounds is a failure in the epithelial-mesenchymal transition (EMT)-correlated re-epithelialization. Despite extensive knowledge pertaining to RNA-binding protein (RBP)-regulated EMT in tumorigenesis, its mechanism during wound healing remains relatively abstruse. Therefore, this thesis aims to analyze the regulation of EMT effectors E-cadherin, Slug and Snail by HuR, a cancer-enhancing RBP, at different time points following wounding. Doxycycline-inducible TRIPZ lentiviral shRNA-mediated RNA interference (RNAi) was conducted prior to keratinocyte scratch assays to understand whether HuR stimulates cell migration in wounds. Real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and immunoblot analyses confirmed HuR knockdown in established stable N/TERT-1 cell lines albeit with certain leakiness. However, similar assays were unable to show any significant relation between HuR and wound re-epithelialization as a consequence of time-limited circumstances. Henceforth, additional biological replicates are required to make robust conclusions.