description) The investigators employed targeted gene disruption of two fucosytransferase (FT) genes, FTIV and FTVII to eliminate the alpha(1,3) fucosylation required for the synthesis of sialyl Lewis X. As alpha(1,3) fucosylated oligosaccharides represent components of leukocyte counter-receptors for E- and P-selectin, and L-selectin ligand, their absence results in a leukocyte adhesion deficiency characterized by blood leukocytosis, impaired leukocyte extravasation, and faulty lymphocyte homing (see 1 and Preliminary Results), and these characteristics are exacerbated in FTIV/VII double-null animals (Preliminary Results). Importantly, FTVII-null and FT-IV/VII-null animals lack detectable infections; but show a profound alteration of bone marrow hematopoietic homeostasis (Preliminary Results). This group of investigators hypothesize that selectin/selectin-ligand counter-receptor pairs are responsible for maintaining bone marrow homeostasis, and that the homeostasis control mechanism involves selectin-mediated interactions between hematopoietic stem (HSC) and/or progenitor (HPC) cells and hematopoietic stromal cells. They further postulate that the general defect in leukocyte extravasation apparent in FT-null animals also extends to the ability of HSC to home to, or engraft within, the hematopoietic microenvironment. The first specific aim of this proposal addresses the mechanism of altered hematopoietic homeostasis in these animals. Multiparameter flow cytometry is used to identify the display of specific selectin-ligands on Lin- Sca+ HSC. As well immune and affinity-adhesion studies will evaluate selectin-ligand expression on HPC of different lineages. Functional studies will determine the role of selectin and their ligands in cell adhesion, cell:cell interactions, and in the ability to initiate and sustain long-term in vitro hematopoiesis. The second specific aim investigates the role of selectins in early events of HSC homing and/or engraftment. The rolling of Lin- Sca+ cells on selectins at physiological shear rates will be studied by Video microscopy. Likewise, the rolling of Lin- Sca+ on endothelial cells will be evaluated using human umbilical vein endothelial cells, and endothelioma cell line, and purified populations of murine bone marrow endothelial cells. Finally, the role of selectins in bone marrow transplantation will be evaluated in 3 systems. Gross phenotypic differences in homing/engraftment are evaluated by transplanting Lin- Sca+ cells from each animal genotype (WT, FTIV-null, FTVIII-null and FTIV/VII-null) into wild-type recipients. Subtle deficiencies in FT-null cell transplantation potential are evaluated in competitive repopulation assays using FT-null cells as the donor cells and wild-type (WT) as the competitor cell population. Finally, they will use parabiotic animals to evaluate HSC bone marrow trafficking in a more physiological setting (i.e., without myeloablation/irradiation). In all these transplantation studies the contribution of WT vs FT-null cells to recipient hematopoiesis is monitored by determining the relative contribution of each to blood leukocyte and
| Cui, J; Wahl, R L; Shen, T et al. (1999) Bone marrow cell trafficking following intravenous administration. Br J Haematol 107:895-902 |
