Proliferation and Potential of Neural and Retinal Stem Cells

The term “stem cell” is often broadly applied to a range of cell types that are relatively undifferentiated and have some capacity for proliferation. In this thesis, I employ a strict definition of stem cells as cells that are capable of both self-renewal and multilineage differentiation. These properties are tested in single precursor cells from the forebrain and its derivative, the retina, using clonal assays. Poor survival is a common problem in single cell cultures, and I show that low oxygen dramatically improves viability in neural stem cells clonally derived from mouse embryonic stem cells, as well as in cultured forebrain neural stem cells. Caspase-dependent and apoptosis-inducing factor-dependent cell death pathways were found to be differentially influenced in low oxygen culture of early, primitive and later, definitive neural stem cells. I isolate precursors from 2 separate regions of the adult mouse forebrain, the lateral ventricle and the hippocampus and argue that only cells resident in the lateral ventricle can be classified as stem cells while the hippocampus contains restricted progenitor cells. Unlike neural stem cells, the very existence of retinal precursors in the adult mammal is controversial. I investigate methods to prospectively identify a rare stem cell population in the pigmented ciliary epithelium of the adult mouse eye and show that, although this population intrinsically gives rise to all retinal cell types, cells can be directed specifically towards a photoreceptor fate by the addition of exogenous factors to the culture media. Pigmentation of retinal stem cells is used as a convenient marker to isolate a retinal stem cell from human embryonic stem cells differentiating under conditions known to promote neural differentiation. Retinal stem cells derived from human embryonic stem cells have highly similar properties to those directly isolated from the eye, and their progeny can similarly be driven to differentiate into photoreceptors. The findings presented in this thesis help to define intrinsic properties of adult neural and retinal precursors and provide a basis for manipulating these cells, potentially for future use in clinical applications.Ph