A thesis submitted to the Faculty of Science in fulfilment of the requirement for the degree of Master of Science, University of the Witwatersrand, Johannesburg, 2019 / The 37/67kDa Laminin Receptor (LRP/LR) LR is a multifunctional cell surface receptor that
maintains several survival processes. It has recently been found that there is a direct relationship
between LRP/LR and the stem cell marker telomerase. Studies have shown that overexpression of
FLAG tagged LRP increases hTERT levels and telomerase activity. Telomerase is a reverse
transcriptase enzyme found in actively dividing cells whose core function involves telomere
maintenance and elongation. The rate limiting component of telomerase, hTERT, is also often upregulated
in rapidly dividing cells to aid stem cell renewal and cell survival and its ectopic
expression can induce epithelial to mesenchymal transition (EMT) resulting in stem cell like
characteristics. In adults, the primary role of stem cells is to repair and regenerate tissue. Hence, this
study aims to determine whether overexpressing LRP::FLAG and the subsequent increase in
hTERT levels induces stem-cell-like characteristics and promotes repair and regeneration in MRC5
lung fibroblasts and HEK293 embryonic kidney cells. Cells were stably transfected with the
pCIneo-moLRP-FLAG plasmid in order to induce LRP::FLAG overexpression. Post-transfection,
an increase in hTERT and phospho-TERT protein levels was observed in both cell lines which is
crucial in maintaining the self-renewal capacity of stem cells. Additionally, an increase in the levels
of pluripotency stem cell markers involved in cell reprogramming and alkaline phosphatase activity
was also observed for HEK293 after transfection. However, in MRC5 cells there was an insufficient
expression of reprogramming factors but, a Cadherin switch indicative of EMT. Moreover,
HEK293 cells overexpressing LRP::FLAG showed no significant changes in the protein levels of
the pro-inflammatory cytokine NF-κB1 and an increase in the anti-inflammatory cytokine TGFβ1
which modulates wound healing. In turn, it led to an increase in the adhesion and migratory
capacity of HEK293 cells. This data suggests that overexpressing LRP::FLAG induces EMT similar
to that observed during induced cell reprogramming and could possibly promote wound healing by
upregulating TERT and TGFβ1 protein levels resulting in stem-cell-like characteristics. / PH2021
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:wits/oai:wiredspace.wits.ac.za:10539/31395 |
| Date | January 2019 |
| Creators | Chigumba, Stephanie |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | Online resource (various pagings), application/pdf |
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