Growth Factor Regulation of Olfactory Neurospheres

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DESCRIPTION (provided by applicant): The olfactory mucosa (OM) of both humans and rodents is an accessible tissue that exhibits ongoing neurogenesis and epithelial reconstitution following injury with demonstrated potential for cell-based therapies. To realize the OM's therapeutic potential requires a culture system that can be used to expand, understand the regulation of, and serve as a biomarker for engraftment potential of progenitor populations. The hypothesis will be tested that olfactory neurosphere formation serves as a biomarker for engraftment potential by varying the "state" of the stem and progenitor cells and also the growth factor influences that act on them. A subsidiary goal is to establish the use of neurospheres as a means of evaluating the network of interactions acting on stem and multipotent progenitor cells in vivo. OM-derived neurospheres will be tested as biomarkers for multipotency and stemness using the sphere-forming activity of cells isolated from in vivo conditions that involve different endogenous stem and progenitor cell activation: normal adult, selective neuronal regeneration, full reconstitution of the epithelium after methyl bromide (MeBr)-induced toxic injury, and early postnatal expansion. Resulting spheres will be quantified and their immunophenotype determined. Olfactory neurospheres will then cultured from constitutively expressing GFP mice and transplanted into MeBr lesioned host animals to determine whether the engrafted cells produce OM cell types. To test whether factors that act on globose basal cells (GBCs) or horizontal basal cells (HBCs) specifically enhance sphere formation and engraftment potential, olfactory neurospheres will be treated with a variety of growth factors with both known tropism as well as unknown function in the OM. Reporter mice that allow for live tracking of GBCs and their progeny will be used to obtain readouts of growth factor activity on olfactory neurospheres. Growth factors that produce a significant change in the size distribution and fluorescence intensity of spheres will be tested via the aforementioned transplantation assay. the stem and progenitor cells ofthe OE are accessible in adult humans and undergo neurogenesis throughout adulthood. As in other systems that have translated stem cell biology to clinical practice, such as the hematopoietic system, it is apparent that the ability to expand the progenitor populations will be required for clinical applications. This work will expand on initial studies showing the clinical potential of olfactory mucosal-derived cells for treatment of spinal cord injury and neurodegenerative diseases by studying what factors can be used to enhance the proliferation and propagation of these stem cell populations.