Author:

Keywords:

Adult, Bone Marrow Cells, Cell Division, Cell Survival, Cells, Cultured, Chemokine CCL3, Chemokine CCL4, Disaccharides, Endothelial Growth Factors, Extracellular Matrix Proteins, Fibronectins, Hematopoietic Stem Cells, Heparitin Sulfate, Humans, Interleukin-3, Lymphokines, Macrophage Inflammatory Proteins, Platelet Factor 4, Stromal Cells, Structure-Activity Relationship, Thrombospondins, Time Factors, Vascular Endothelial Growth Factor A, Vascular Endothelial Growth Factors, Science & Technology, Life Sciences & Biomedicine, Hematology, ENDOTHELIAL GROWTH-FACTOR, HEMATOPOIETIC STEM-CELLS, BONE-MARROW STROMA, COLONY-STIMULATING FACTOR, PROGENITOR CELLS, IN-VITRO, EXTRACELLULAR-MATRIX, MAMMARY CELLS, GM-CSF, GLYCOSAMINOGLYCANS, 1102 Cardiorespiratory Medicine and Haematology, 1103 Clinical Sciences, 1114 Paediatrics and Reproductive Medicine, Immunology, 3101 Biochemistry and cell biology, 3201 Cardiovascular medicine and haematology, 3213 Paediatrics

Abstract:

We have shown that stromal O-sulfated heparan sulfate glycosaminoglycans (O-S-GAGs) regulate primitive human hematopoietic progenitor cell (HPC) growth and differentiation by colocalizing heparin-binding cytokines and matrix proteins with HPC in stem cell "niches" in the marrow microenvironment. We now show that long-term culture-initiating cells (LTC-IC) are maintained for 5 weeks in the absence of stroma when O-S-GAGs are added to IL-3 and either MIP-1alpha or PF4 (LTC-IC maintenance without GAGs, 32 +/- 2%; with GAGs, 95 +/- 7%; P <.001). When cultured with 5 additional cytokines, O-S-GAGs, IL-3, and MIP-1alpha, LTC-IC expanded 2- to 4-fold at 2 weeks, and 92 +/- 8% LTC-IC were maintained at 5 weeks. Similar results were seen when PF4 replaced MIP-1alpha. Although O-S-GAG omission did not affect 2-week expansion, only 20% LTC-IC were maintained for 5 weeks. When O-S-heparin was replaced by completely desulfated-, N-sulfated (O-desulfated), or unmodified heparins, LTC-IC maintenance at week 5 was not better than with cytokines alone. Unmodified- and O-S-heparin, but not desulfated- or N-sulfated heparin, bound to MIP-1alpha, IL-3, PF4, VEGF, thrombospondin, and fibronectin. However, the affinity of heparin for thrombospondin and PF4, and the association and dissociation rates of heparin for PF4, were higher than those of O-S-heparin. We conclude that (i) although cytokines may suffice to induce early expansion, adult human LTC-IC maintenance for longer than 1 month requires O-S-GAGs, and (ii) HPC support may depend not only on the ability of GAGs to bind proteins, but also on optimal affinity and kinetics of interactions that affect presentation of proteins in a biologically active manner to progenitors. (Blood. 2000;95:147-155)