Characterisation of fluorescent extracellular vesicles and their distribution in the prostate cancer microenvironment

Konstantinou, Timothea 2021. Characterisation of fluorescent extracellular vesicles and their distribution in the prostate cancer microenvironment. PhD Thesis, Cardiff University. Item availability restricted.

Preview	PDF - Accepted Post-Print Version Download (29MB) | Preview
	PDF (Cardiff University Electronic Publication Form) - Supplemental Material Restricted to Repository staff only Download (189kB)

Abstract

Small extracellular vesicles (sEVs) are nanovesicles secreted from cells, that elicit intercellular communication by transferring their cargo to recipient cells. This form of communication contributes towards disease-critical processes in prostate cancer (PCa), including sEV-mediated microenvironment remodelling, tumour progression, angiogenesis, and formation of the pre-metastatic niche. The fundamental mechanisms of sEV dissemination through the microenvironment, as well as distribution within tissues have not been extensively investigated. This is the topic of the current study. To address these questions, sEVs require fluorescent labelling; we developed two PCa cell lines that generate endogenously fluorescent sEVs by expressing GFP or Tdtomato fluorescent proteins fused to CD63 or CD81, respectively. Fluorescent sEVs were used to microscopically characterise uptake kinetics (dosing), retention, and distribution in recipient PCa cell- and bone marrow-derived mesenchymal stem-cell (BMMSC) - populations. Additionally, a vital consideration was the impact of the tetraspaninfluorescent protein fusions on the phenotype of the parent cells, and the corresponding sEVs. Forced overexpression of tetraspanin-fluorescent protein fusions drove significant phenotypic and proteomic changes at both the cell and sEV level. The differential impact of a two- versus three-dimensional cell culture microenvironment on the phenotype of cells and sEVs was also demonstrated, emphasising the importance of platform definition and characterisation. Nevertheless, the behaviour of fluorescent sEVs revealed conserved principles in sEV uptake kinetics, retention, and distribution in two dimensional dynamic cell systems, highlighting recipient-cell type differences in signal uptake and retention. In addition, functional effects of sEV uptake were identified demonstrating osteogenic differentiation of BM-MSCs as a result of chronic dosing with PCa-derived sEVs. To conclude, our study generated new optical tools allowing fundamental principles of sEV distribution in tissues or cells to be defined. The data provides the basis for understanding the dispersion of sEVs in tissue microenvironments, and new insights into their great potential for diverse clinical applications.

Item Type:	Thesis (PhD)
Date Type:	Completion
Status:	Unpublished
Schools:	Medicine
Date of First Compliant Deposit:	24 February 2022
Last Modified:	06 Jan 2024 03:16
URI:	https://orca.cardiff.ac.uk/id/eprint/147789

Actions (repository staff only)

Edit Item