Jellied vesicles: probing the interactions between vesicles and the extracellular matrix environment
Year: 2024
Type: dissertation
Abstract: Extracellular vesicles (EVs) are membrane nanostructures released by cells into their external environment that function in signalling and cell-to-cell trafficking of molecular cargo. They are a prominent research topic for their roles in both physiological and pathological processes, from wound healing, immune modulation, and morphogenesis, to cancer progression and metastasis. The ability of EVs to traverse diverse tissue microenvironments and cross different biological barriers, as well as their tendency to carry sensitive materials capable of reprogramming other cells make EVs an attractive model system to mimic in the development of nanoscale delivery systems for therapeutics. Despite growing interest in EVs, it is not fully understood how their diffusion through tissues might be affected by the structure and composition of the environments they are found in. In order to better understand how EVs interact with the extracellular matrix (ECM) that makes up tissues and how these interactions influence EV behaviour, I study the diffusion of fluorescently labelled EVs and model nanoparticles in different hydrogel materials that reflect the structural and compositional properties of ECM. I first use synthetic liposomes and agarose hydrogels to study how particle behaviour is affected by steric and electrostatic interactions with a porous matrix environment. Even without biochemically complex molecules that enable specific receptor-ligand interactions, different levels of particle mobility can be achieved through the intersecting effects of charge-based adhesion or repulsion and steric size exclusion of rigid versus deformable particles (Tam et al, Biophys J, 2024). System complexity is increased by using breast cancer cell-derived EVs and studying how they interact with hydrogels composed of fibrillar collagen I. By comparing EV behaviour with that of trypsinized EVs with their surface proteins digested, extruded vesicles composed of whole plasma membrane, and synthetic liposomes, I am able to identify proteins as being primarily responsible for EV interactions with collagen I. By treating EVs with a synthetic argynylglycylaspartic acid (RGD) peptide inhibitor of integrin complexes, I determine that these interactions and the subsequent immobilization of EVs in collagen I hydrogels are due to the specific binding of EV integrins to RGD motifs in colloidal collagen I (Tam et al, under review, ACS Nano). Following the discovery of this interaction between colloidal collagen I and EVs, I investigate whether EVs can be involved in ECM remodelling. Using rheometry and confocal microscopy, I determine that EVs are capable of accelerating fibrillogenesis by recruiting collagen I molecules from solution and nucleating new fibrils. In doing so, EVs become entangled in the fibril structures they help to create, incorporating themselves into the ECM. This behaviour is not observed in trypsinized EVs, synthetic liposomes, or extruded plasma membrane vesicles, suggesting that this phenomenon is due to specific interactions between EV membrane proteins and collagen I molecules in solution. The integration of EVs into matrix structures also cannot be achieved through having EVs diffusing into pre-formed collagen gels, suggesting that the formation of de novo ECM structures may be a specialized biological role of EVs that has not been previously explored and could lead to the generation of EV-primed ECMs. These have been shown to play a role, for example, in the formation of pre-metastatic niches (Tam et al, in preparation). In studying how EVs interact with ECM, I show not only that EV diffusion is governed by the intersecting effects of physical, electrostatic, and specific receptor-ligand EV-matrix interactions, but also that EVs can take part in ECM remodelling processes. In doing so, they impart cell signalling cues into the matrix, both through the act of remodelling and through being embedded within the matrix. The implications of this work have wide impacts, from the development of more effective delivery systems for therapeutic materials, to improved integration of EVs into bioactive scaffolds for tissue engineering, and even to a better understanding of how EVs enable cancer metastasis through remodelling of ECM.
Author: Tam, Nicky W.
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W6964483766
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