The over all objectives of this project is to establish a hemopoietic stem cell model which can be tested to study and understand the mechanism of self renewal and/or commitment to differentiated cell types by a primitive cell. A high incidence of lineage, undifferentiated cells was identified in the spleens of female New Zealand Black Mice. Phenotypic, morphological, histochemical, genomic and functional characterization of the purified population suggested that these may be very primitive cells of the hemopoietic system. During the present reporting period (October 1990 through September 1991) comparative and quantitative restriction fragment mapping of the genomic DNA was carried out to confirm the germ line configuration of the genome of these cells. In Vitro, these cells were found to differentiate when exposed to a few early-acting lymphokines and a bone marrow stromal cell conditioned medium. In that, both myeloid (adherent mac-1+), and lymphoid (CD3+, surface Ig+) cell types were detectable. In vivo, these cells were found to differentiate and constitute immunedefficient SCID mice (lack functional T and B cells). Circulating immunoglobulin s of both IgM and Igg class were detectable by ELISA in the reconstituted mice 6, 8 and 12 week post transfer. Mac-1+, CD3+, and sIg+ cells were detectable in the spleens of these reconstituted mice. These results further define the NZB splenic primitive cells as true multipotential stem cells. Using this model system, experiments are underway to study the clonal analysis using this model system, experiments are underway to study the clonal analysis of stem cells in vivo, to define the in vitro culture conditions to support long term maintenance and differentiation of these cells, to investigate the early transcriptional events occurring in these cells in response to the inducing stimuli.
