Regeneration of Ablated Motor Nerves Using Olfactory Neuroepithelial Derived Stem Cells In a Pre-Clinical Model
Peripheral nerve injury is a life-changing problem, which presents significant therapeutic challenges to clinicians and patients. A number of different approaches to dealing with the issue exist but to date there is no consensus on the most effective method of treatment. With large gap nerve transection or central to peripheral nervous system intersection injury, spontaneous meaningful axonal recovery seldom occurs. Despite surgical intervention with autografts, the functional outcomes are often unfavorable. Tissue engineered strategies utilising scaffolds which mimic the properties of nerve grafts, but which prevent the need for the secondary injury associated with harvesting autografts, are currently being investigated. Considerable interest has been given to the combined use of biologically active scaffolds with progenitor cells which function synergistically to enable functional tissue growth. Combining the purely surgical approach with contemporary developments in tissue engineering offers a challenging but potentially rewarding way forward. In this thesis, the possibilities and the challenges of a fusion between surgery and tissue engineering is examined. While much of the analysis is determined by the surgical background and training of the author, the outcome of the research points to potentially very exciting prospects for further joint research.
This dual purpose, prospective in vitro and in vivo research project has critically evaluated the capacity of stem cells derived from the olfactory neuroepithelium (ONS cells) to act with a purpose designed collagen-hyaluronic acid based biological nerve guidance conduit, as an efficient graft material in the repair of surgically ablated peripheral nerve tissue. The approach incorporated (i) an evaluation of the effect of biomaterials and growth factors on the neurogenic and gliogenic capacity of ONS cells, (ii) an evaluation of the in vitro behavior of ONS cells cultured within a purpose designed collagen-hyaluronic acid biological nerve guidance conduit, and (iii) an in vivo evaluation of the capacity of ONS cells combined with the nerve guidance conduit to mediate functional healing following critical defect injury in a rat sciatic nerve model.
The two-dimensional in vitro analysis described in Chapter 2, demonstrated that multipotent highly plastic stem cells with the potential to differentiate along neural and glial lineage could be harvested from explants taken from the rat nasal cavity. Interestingly, ONS cells derived from young donor animals were found to have increased multipotentiality in comparison to ONS cells derived from older animals. In addition, the work demonstrated that ONS cells proliferation and differentiation potential is dependent on biomaterials to varying degrees. Nerve growth factor was also found to have similar effects to biomaterials on the differentiation capacity of ONS cells.
The three-dimensional in vitro analysis described in Chapter 3 demonstrated that an ergonomic and robust biphasic nerve guidance conduit, produced by the combination of a tubular collagen conduit with a biologically active, laminin functionalised hyaluronic acid hydrogel luminal filler, supported ONS cell viability and differentiation whilst the addition of nerve growth factor promoted both differentiation and morphological extension of ONS cells.
The in vivo study described in Chapter 4 demonstrated that the biphasic nerve guidance conduit promoted anisotropic peripheral nerve regeneration following critical sciatic nerve injury, and that functional and morphological repair was enhanced with the addition of ONS cells. Moreover the addition of nerve growth factor was found to mediate functional healing.
Collectively, this thesis points to the development of a novel biomimetic biphasic nerve guidance conduit with an optimised composition and suitable mechanical properties with which to deliver multipotent ONS cells for use in peripheral nerve defects. This research work highlights the importance of biomaterials and growth factor selection in the design of cell specific delivery systems for peripheral nerve regeneration with potential for use in future clinical trials.