This projects long term objectives are to control production and differentiation of primitive hemopoietic stem cells (PHSC) for engraftment PHSC are the precursors from which all lymphoid and myeloid cells are continuously derived. This project focuses on the precursors that produce most differentiated cells over many months. Success of clinical marrow transplantation depends on their function. Currently PHSC can be assayed only in vivo using rodent models. PHSC populations will be analyzed using competitive repopulation techniques that directly assay repopulating and differentiating ability in vivo over 12 months. A math model win estimate numbers of the precursors from which most differentiated cells are descended. These measures will be complemented by unique retroviral insertions to define duration and extent of function of individual PHSC clones.
Specific aims are: (1) To assay PHSC from alternative sources: 14-18 day fetal liver, and 19 day fetal blood (model for umbilical cord blood). Numbers and functional abilities will be defined. (2) To measure immunogenicity of PHSC antigens. Antigenically disparate cells will repopulate immune competent W-anemic mice, in comparison with immune deficient irradiated mice. (3) To identify the multilineage precursors that produce significant numbers of different cell types in engraftment, by correlations between cell types in recipients of genetically marked marrow mixtures at time periods before cells are all descended from PHSC. (4) To optimize stimuli for PHSC proliferation in vitro. Tissue preparations will be used from W-anemic recipients with maximal PHSC proliferation, and growth factors will be identified in successful preparations. (5) To develop models for assessing human PHSC number and function. Recipients will be triple mutant scid/scid, W-anemic, and bg/bg mice that cannot reject xenografts and have defective PHSC and natural resistance. 'Me models will be proven on PHSC from Peromyscus, to show that they assay PHSC accurately despite species differences.
