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This thesis is an investigation into the role played by a variety of small non-coding RNAs, called microRNAs, in regulating the changes in cell properties that occur at the margin of a healing wound. The study relied on the the development of a model of the healing wound edge, then investigating the changes in microRNAs that occurred there, and the downstream effects of those changes on proteins known to contribute to wound healing.

Background: Non-compliance with infection control guidelines has been reported within healthcare settings. Infection control education in undergraduate healthcare education programs forms a critical component in preparing student healthcare workers for vocational roles. Methods: Clinical sciences students (nutrition science, paramedicine, pharmacy, podiatry, optometry studying for qualifications recognised by the Australian Health Practitioner Regulation Agency) self-reported hygiene perceptions and practices and collected microbiological swabs from personal or medical equipment items before and after recommended disinfection procedures. Results: Cultivable microorganisms were isolated from 95% of student medical equipment items. Disinfection significantly reduced microbial growth on student medical equipment items (P < 0.05). Conclusions: Student perceptions of infection control procedures do not always correlate with infection control practice. Infection control education of undergraduate healthcare students requires ongoing assessment to ensure successful translation into clinical practice.

The scratch or wound-healing assay is used ubiquitously for investigating re-epithelialisation and has already revealed the importance of cells comprising the leading edge of healing epithelial wounds. However it is currently limited to studying the effect of known biochemical agents on the tissue of choice. Here we present an adaptation that extends the utility of this model to encompass the collection of cells from the leading edge of migrating epithelial sheets making available explorative biochemical analyses. The method is scalable and does not require expensive apparatus, making it suitable for large and small laboratories alike. We detail the application of our method and exemplify proof of principle data derived from primary human keratinocyte cultures.

Cells at the wound margin during reepithelialization share phenotypic and genotypic similarities with cancer metastasis. In both scenarios, effected cells increase their epithelial characteristics. A sedentary, epithelial phenotype converts to a more motile, mesenchymal state in a process known as epithelial‐to‐mesenchymal transition (EMT). The extensive evidence that microRNAs induce EMT during cancer cell metastasis and migration is lacking in wound healing. Using an in vitro model of re‐epithelialization, primary keratinocytes at the wound edge were visualized migrating and undergoing dynamic morphological changes as they extended and retract lamellipodia at the leading edge of a collective sheet. Furthermore, expression of mesenchymal proteins vimentin and cytokeratin‐14 increased only at the leading edge suggesting a localized and partial EMT had occurred. Next, RNA was extracted from cells at the leading edge and from cells distal to the edge. Next generation sequencing revealed a number of microRNA species are differentially expressed in the cells at the migrating leading edge including miR‐501‐3p, miR‐4972 and miR‐181a‐5p (padj<0.001). MiR‐501‐3p and miR‐181a‐5p are known to be up‐regulated in tumor cells and microRNA‐4792 has been shown to target and repress an important EMT transcription factor, FOXC1. These results suggest that these miRNAs may play a similar role in EMT during wound healing. MiR‐4792 overexpression and knock down experiments are now underway in keratinocytes to investigate functional effects and mechanisms of action.

In recent years, many studies have provided evidence that exosomes secreted by cells contain various components, including microRNAs [1]. It is thought that exosomes have important roles in many biological processes. However, the role of exosomes and their components, especially miRNAs, in wound healing is poorly understood. In order to understand whether or not primary human epidermal keratinocytes and dermal fibroblasts, two important cell types contributing to wound healing process, release exosomes and what species of wound healing-associated miRNAs accumulate in these vesicles, this project will use a combination of methods to isolate and characterize exosomes, to profile exosomal cargo’s, especially miRNAs in exosomes. The results showed that keratinocytes and fibroblasts released exosomes into conditioned media and these exosomes contain some target miRNAs.

Free to read It is becoming increasing clear that microRNAs contribute to the regulation of many biological processes, including wound healing. After injury, keratinocytes need to undergo what is known as an epithelial-to-mesenchymal transition (EMT) to initiate re-epithelialisation. During this process, keratinocytes reduce their attachment to the underlying matrix, extend membrane protrusions, become motile and migrate over the wound bed, affecting wound closure. MicroRNAs that regulate EMT are aberrantly upregulated in keratinocytes at the edge of non-healing wounds and potentially play a role in the chronicity of these wounds. In vitro and in vivo, downregulation of these microRNAs promotes EMT and migration, facilitating re-epithelialisation in wound models. This review will focus on the role of microRNAs that regulate or have potential to regulate EMT and re-epithelialisation during wound healing