Generating exogenic neural cells through blastocyst complementation for neurodegenerative diseases

Abstract

Neurodegenerative diseases burden individuals, their families, and the economy. The progressive symptoms and neuropathology affect cognitive skills and daily activities. The only available treatments alleviate symptoms, but do not stop or change the course of disease progression. Therapeutics are rather limited for neurodegenerative diseases. Cellular transplantation to treat neurodegenerative diseases has shown promise as a therapeutic, but a novel source of cells is needed. Fetal/embryonic tissue is fraught with ethical concerns and many stem cell derived cells behave differently than primary cell types. This dissertation seeks to bridge some of these gaps by characterizing and quantifying cell types found within chimeric brains. Insights from this research have the potential to inform neurodegenerative therapeutics, deliver new information of cell survival and differentiation, and elucidate some interspecies chimeric differences. We reviewed the current state of cellular transplantation, relevant therapeutic neuronal and non-neuronal types, interspecies chimeric barriers, intraspecies and interspecies chimeras, and generation of chimeric brains. Our study of E12.5 intraspecies mouse-mouse chimeric brains identified, characterized, and quantified serotonergic, dopaminergic, and cholinergic precursors derived from donor cells. Microglia-like and macrophage-like cells derived from donor cells were also observed and quantified. To better understand development of these cells, E16.5 intraspecies mouse-mouse chimeric brains were generated. To support interspecies chimera characterization, we generated E12.5 interspecies rat-mouse chimeric brains, and began to characterize and quantify cell types. Dopaminergic precursors, microglia-like and macrophage-like cells were identified cells in these interspecies chimeras. To demonstrate that a targeted developmental niche could be made, we developed and tested single guide RNA to target a PU.1 gene that is important for microglia development. This research seeks to improve our understanding of cell types and proportions of those cell types generated in wild type intraspecies and interspecies chimeras. It also highlights new methods, like automated imaging, for quantifying cell types within these chimeric animals. This research also provides the foundational steps for making a new PU.1 knockout model to be used with blastocyst complementation.