Takov, Kaloyan Valentin;
(2020)
Cardioprotective and proangiogenic activities of small extracellular vesicles released by amniotic fluid stem cells.
Doctoral thesis (Ph.D), UCL (University College London).
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Abstract
Protection against myocardial ischaemia/reperfusion injury and regeneration of the damaged myocardium are long-sought goals. The use of small extracellular vesicles (sEVs) released by mesenchymal stem cells (MSCs) was shown to be of benefit in the myocardial infarction setting. However, MSCs are frequently harvested from aged or diseased patients and suboptimal sEV isolation methods are used. A subtype of young, foetal MSCs, namely spindle-shaped amniotic fluid stem cells (SS-AFSCs), is known to possess better expansion and functional capacity than its adult counterparts. Here, sEVs released by SS-AFSCs were isolated using size-exclusion chromatography (SEC) – an isolation technique that yields vesicles of superior purity – and their cardioprotective and proangiogenic activities were studied. Firstly, using rat blood plasma, it was demonstrated that SEC isolates higher sEV yields with significantly compromised purity, mostly due to the presence of lipoproteins. To overcome this, a serum-free environment was used for sEVs isolation from SS-AFSC-conditioned medium. Comprehensive characterisation experiments showed that the harvested SS-AFSC sEVs are of high purity. Functionally, SS-AFSC sEVs protected the rat myocardium from ischaemia/reperfusion injury in vivo, but not isolated cardiomyocytes in vitro, indicative of indirect cardioprotective effects. Additionally, SS-AFSC sEVs promoted migration of endothelial cells in vitro and recapitulated the promigratory effects of the SS-AFSC-conditioned medium. Using pharmacological inhibition, it was shown that PI3K pathway, a known player in cell migration, mediates the sEV effects, while a series of potential candidates in the sEV cargo were excluded. Finally, cellular sEV uptake was studied by use of lipophilic dye-labelling experiments. Surprisingly, this commonly used approach was found to be unsuitable for sEV tracking due to non-specific dye retention by non-sEV contaminants. Overall, SEC-isolated SS-AFSC sEVs possess cardioprotective potential manifested only in vivo, and promigratory activity which requires PI3K signalling. These data indicate that SS-AFSC sEVs have multifactorial beneficial effects in a myocardial infarction setting.
| Type: | Thesis (Doctoral) |
|---|---|
| Qualification: | Ph.D |
| Title: | Cardioprotective and proangiogenic activities of small extracellular vesicles released by amniotic fluid stem cells |
| Event: | UCL (University College London) |
| Open access status: | An open access version is available from UCL Discovery |
| Language: | English |
| Additional information: | © The Author 2020. Original content in this thesis is licensed under the terms of the Creative Commons Attribution 4.0 International (CC BY 4.0) Licence (https://creativecommons.org/licenses/by/4.0/). Any third-party copyright material present remains the property of its respective owner(s) and is licensed under its existing terms. Access may initially be restricted at the author’s request. |
| UCL classification: | UCL |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10088992 |
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