IMMORTALIZED AND GRAFTING OF NEURAL PROGENITORS

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In the field of neural transplantation, alternatives to primary fetal tissue are being sought as graft material. Previously, through retrovirus- mediated gene transfer, we generated immortalized, clonal, multipotent neural progenitor lines from mouse. Examination of their differentiation potential in vitro suggested enormous plasticity at the level of the individual progenitor. Upon transplantation, these immortalized progenitors engrafted in a cytoarchitecturally and functionally appropriate manner, recapitulating their multipotency in vivo & expressing retrovirally-transduced exogenous genes in a robust, stable fashion throughout brain parenchyma for prolonged periods. These progenitors could engraft & participate normally in the development of multiple structures along the neuraxis & at multiple stages spanning from embryonic to the adult. They differentiated into multiple cell types, presumably responding to signals of the respective region at the particular developmental stage. The progenitors, though most engraftable when initially mitotic, migratory, & plastic, quickly differentiated, down-regulated their immortalizing gene product, & never formed tumors. CNS cytoarchitecture was never disrupted, the blood-brain barrier remained intact, & recipient animals behaved normally. When transplanted into various animal models of neural degeneration, the progenitors integrated into cytoarchitecture often assuming the phenotype of the missing cell type. Furthermore, progenitors expressing gene products missing in the host successfully engrafted into the CNS of various animal models in which defects in those genes had been defined (e.g., certain neurovisceral storage diseases). These data suggest the feasibility of using immortalized progenitors to provide exogenous factors of therapeutic or developmental interest (constitutiveiy or through genetic engineering), or to effect repair as integral members of CNS cytoarchitecture, may be feasible. The generation of progenitor lines with similar potential from human fetal neural tissue is the next step. Starting with primary human fetal neural tissue, the proposed studies will attempt to establish a paradigm of neural progenitor transplantation as a therapy for developmental, degenerative, & acquired injury to CNS through the following 4 aims: (1) Determine conditions for maintaining human fetal neural tissue in vitro which would optimize the presence of proper progenitors for immortalization. (2) Generate & characterize in vitro clonal, immortalized, multipotent human neural progenitor lines. (3) Transplant candidate lines into mouse hosts (cyclosporin-treated or SClD mice) to assess ability to engraft, differentiate in vivo, & express a reporter transgene. (4) Attempt transplantation of engraftable progenitor lines into mouse models of neural degeneration.