Awan, Maooz Afzal;
(2022)
Cryopreservation of Hydrogel Encapsulated Spheroids.
Doctoral thesis (Ph.D), UCL (University College London).
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Abstract
The clinical delivery of tissue engineered (TE) therapies is limited by an inability to effectively store them, but responsive manufacture is unsuitable for most TE therapies. It is important that TE therapies can be effectively stored, so they can be delivered to patients in a clinically relevant time frame. Hence, cryopreservation of TE therapies has become increasingly important to improve their availability. One such TE therapy is the BioArtificial Liver (BAL), an extra-corporeal organ support device designed for the treatment of acute liver failure (ALF). The work presented herein aimed to improve the cryopreservation outcomes of the BAL biomass in order to provide an off-the-shelf treatment for ALF. Upon investigation of the physical aspects of cryopreservation, slow cooling was found to produce a less crystalline frozen matrix than rapid cooling. Improved post-thaw recovery of viable cell number was demonstrated with very slow cooling (−0.3°C/min) and slow warming (12 °C/min). At large volumes the biomass was shown to nucleate uniformly at the same temperature without the need for controlled nucleation, thus eliminating the need for a nucleating agent. Post-thaw washing of the biomass with tapering concentration of osmotic buffer effectively removed the cryoprotectant to undetectable levels, thereby minimising toxicity to the patient. Reactive oxygen species (ROS) were found not to be the primary mediators of cryopreservation induced cell death. ROS formation was successfully inhibited, either through the use of antioxidants or by metabolic suppression, however it did not improve post-thaw recovery. A less cell-dense biomass was shown to produce a more rapid post-thaw recovery, which appeared to be linked to improved preservation of microtissue structure, specifically through focal adhesion induced survival. Post-thaw preservation of focal adhesions at lower cell densities was linked to increased ERK and Akt survival signalling. Together these findings indicated that a cryopreserved BAL biomass can recover to a performance competent cell dose within 72 hours post-thaw, which will aid clinical translation of the BAL.
| Type: | Thesis (Doctoral) |
|---|---|
| Qualification: | Ph.D |
| Title: | Cryopreservation of Hydrogel Encapsulated Spheroids |
| Event: | UCL (University College London) |
| Open access status: | An open access version is available from UCL Discovery |
| Language: | English |
| Additional information: | Copyright © The Author 2022. Original content in this thesis is licensed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International (CC BY-NC 4.0) Licence (https://creativecommons.org/licenses/by-nc/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 > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Div of Medicine UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10142669 |
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