Neural stem cell proliferation and differentiation on functional electrospun fiber matrices.

摘要

Recent progress shows that neurons and oligodendrocytes can be generated from various stem cell sources in culture and when transplanted survive and provide a level of functional integration, fueling efforts to develop stem cell-based transplantation therapies for treating neurodegenerative disorders. A major challenge remains the efficient expansion of neural stem/progenitor cells (NSCs) and strategies to direct their differentiation.;In vivo, NSCs are regulated by a complex array of biochemical and physical cues emanating from their microenvironment, the stem cell niche. Examination of the niche reveals a preponderance of fibrous topography, and increasing experimental evidence suggests active roles of nanoscale features in regulating stem cell behavior.;The objective of this thesis research is to investigate the effect of a fiber topographical cue, i.e. diameter, on the proliferation and differentiation of NSCs. First, we devised a method for preparing polyethersulfone electrospun fiber matrices with controlled diameter and narrow variability representative of the diameter range found in basal lamina in vivo. We then examined the proliferation and differentiation of rat adult NSCs (rNSCs) on laminin-coated fiber matrices with average fiber diameters of 283 +/- 45 nm, 749 +/- 153 nm and 1452 +/- 312 nm in proliferation and differentiation media.;Under expansion condition, rNSCs cultured on fiber substrates showed lower proliferation and more rounded morphology compared to 2D surfaces. Under the differentiation influence of retinoic acid and fetal bovine serum, small diameter fiber substrates yielded an increase in oligodendrocyte differentiation, while larger diameter fibers directed cells towards neuronal differentiation. Combining 228-nm fiber topography with an oligodendrocyte-differentiation cue synergistically promoted oligodendroglial differentiation and produced nearly 100% of oligodendrocyte precursors.;Initial cultures of fiber substrates with human NSCs revealed that topographical regulation is cell-type dependent, and that effective configurations need to be optimized for each cell type. We developed a convenient fiber bonding and fusion technique as a method for manufacturing fiber substrates suitable for future human NSC culture.;Collectively, these studies demonstrated that fiber topography plays an important role in regulating differentiation and proliferation of NSCs in culture, and underscores the importance of properly examining the cellular mechanisms triggered in response to topographical cues.