Dale, Tina Patricia (2016) Investigating the chondrogenic phenotype in clinically relevant cells: the effect of hTERT expression. Doctoral thesis, Keele University.

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Damaged or diseased mature articular cartilage cannot undergo effective tissue repair and due to its avascular, hypocellular nature defects become widespread and painful. No ‘gold standard’ treatment exists for this indication and the ultimate recourse is prosthetic joint replacement. Cartilage is therefore an ideal target for regenerative medicine therapies aiming to recapitulate native cartilage.

Despite over fifty years of research and encouraging outcomes, re-creation of the hyaline tissue has yet to be consistently achieved, possibly as a result of the application of a sub-optimal cell type. Chondrocytes and bone marrow mesenchymal stem cells (MSCs) have been used clinically, with future prospects for other alternative MSC sources and human embryonic stem cell (hESC)-derived cells. Further in vitro study of cellular chondrogenic capacity is desirable but hampered by cell changes and senescence. This work examines the hypothesis that the re-introduction of the catalytic sub-unit human telomerase reverse transcriptase (hTERT) can extend the proliferative cell capacity of cells whilst concomitantly bypassing changes associated with cell aging and senescence. The utility of umbilical cord blood (UCB) as a possible alternative source of more naive MSCs was also investigated.

Human bone marrow MSCs, chondrocytes, and hESC-derived cells were transduced with hTERT and their resulting chondrogenic capacity, assessed principally by extracellular matrix (ECM) production and gene expression, examined and compared to that of the three non-transduced, parental cell sources.

UCB was not found to be a viable alternative MSC source due to a very low cell number and colony recovery; however, foetal bovine serum (FBS) batch and atmospheric oxygen tension were identified as key to influencing recovery outcomes. Of the three parental cell types examined for chondrogenic potential MSCs and chondrocytes produced similar amounts of sGAG but chondrocytes produced a more homogeneous ECM with persistent chondrogenesis, whereas MSCs became hypertrophic. hESC derived cells had a more muted chondrogenic response with similarities to both chondrocytes and MSCs. TERT extended the proliferative capacity of all three cell types, two extensively but was also associated with changes in cell phenotype and a reduction, although not complete ablation, in the subsequent chondrogenic capacity.

Taken together the results demonstrate that with current differentiation techniques primary articular chondrocytes provide the most optimal result, supporting their continued use for clinical therapies, and this capacity may not be preserved by the application of hTERT transduction strategies.

Item Type: Thesis (Doctoral)
Uncontrolled Keywords: articular cartilage, chondrogenesis, cell therapy, regenerative medicine, extracellular matrix, telomerase, hTERT, senescence, immortal, chondrocyte, mesenchymal, embryonic, stem cell, hypertrophy, umbilical cord blood
Subjects: R Medicine > R Medicine (General)
Divisions: Faculty of Medicine and Health Sciences > Institute for Science and Technology in Medicine
Contributors: Forsyth, NR (Thesis advisor)
Depositing User: Lisa Bailey
Date Deposited: 07 Nov 2016 10:30
Last Modified: 10 Aug 2022 15:56
URI: https://eprints.keele.ac.uk/id/eprint/2440

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