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Title: Differentiated mesenchymal stem cells enhance nerve regeneration by trophic influences
Author: Mahay, Daljeet
ISNI:       0000 0001 3616 5585
Awarding Body: University of Manchester
Current Institution: University of Manchester
Date of Award: 2008
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Schwann cells (SC) are essential facilitators for peripheral nerves following injury by releasing supporting neurotrophic factors that provide physical support and guidance for regeneration. In vitro these cells are slow growing, hence not well suited to a tissue engineering approach to nerve repair. To consider alternatives, adult rat bone marrow mesenchymal stem cells (MSC) were differentiated (dMSC) into SC-like cells using an established cocktail of growth factors. Qualitative reverse transcriptase polymerase chain reaction (RT-PCR), western blotting and immunocytochemistry approaches showed that dMSC expressed glial cell markers and SC associated neurotrophic factors at a transcriptional and translational level. To further assess the properties of the dMSC, the cells were co-cultured with dorsal root ganglia (DRG) neurons and the secretion of the neurotrophic factors was evaluated. The neurite outgrowth of the DRG neurons was enhanced in co-culture with dMSC. Like SC, dMSC were responsible for the stimulation of longer and branched neurites in the co-culture model. The neurite outgrowth of DRG neurons was enhanced when co-cultured with dMSC due to trophic influences. Studies with ELISA methodology and blocking antibodies showed that this effect was due to the predominant release of brain derived neurotrophic factor and nerve growth factor which were up-regulated in dMSC following co-culture. This finding was confirmed by the addition of preconditioned medium from the dMSC and the expression of tropomyosin-related kinase receptors in the DRG neuron co-cultures. The functional properties of the dMSC were further elucidated in vivo by transplanting green fluorescent protein (GFP) transfected cells using a poly-3-hydroxybutyrate conduit in a rat sciatic nerve injury model. The regenerative capacity was evaluated after I and 2 months at the distal nerve stump by transmission electron microscopy. The number of myelinated regenerated nerve fibres was greater in the 2 months post-transplantation compared to the 1 month groups. The survival of the transplanted cells was confirmed by the detection of the GFP plasmid by quantitative PCR and the GFP expression in the cells by quantitative RT-PCR. For a further extension of the study, human-derived MSC from three patients were differentiated along a SC lineage and displayed SC characteristics. The results of this thesis further support the notion that MSC differentiated into SC-like cells display cellular, molecular and functional characteristics of SC to provide therapeutical substitutes for nerve regeneration following injury.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available