Maintenance of CD34 Expression During Proliferation of CD34 + Cord Blood Cells on Glycosaminoglycan Surfaces

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Abstract
Recent studies have indicated that glycosaminoglycan (GAG) interactions with hematopoietic progenitors play a significant role in the regulation of hematopoiesis. However, the details of these interactions are not clear. In this study, we examined the role of soluble and immobilized GAGs in the proliferation of CD34+ cells. Chitosan, a cationic polysaccharide, was used to immobilize GAGs in ionic complex membranes. The GAGs studied were heparin, hyaluronate, and chondroitin sulfates A, B, and C. CD34-enriched umbilical cord blood cells were seeded onto tissue culture plates coated with the GAG-chitosan complex membranes. Cultures were maintained in medium supplemented with stem cell factor and interleukin 3 for up to six weeks, during which total and CD34+ cell numbers were determined by flow cytometry. Total cell number expansion ranged from 25-fold to 40-fold after six weeks. However, only heparin and chondroitin sulfate B (CSB) surfaces retained a significant CD34+ fraction. All other surfaces exhibited declines in CD34 expression, with negligible CD34+ percentages remaining after four weeks. In contrast, heparin and CSB surfaces exhibited CD34+ fractions as high as 90% after four weeks. GAG desorption studies indicated that the observed effects were partly mediated by desorbed GAGs in a concentration dependent manner. Subsequent studies showed that sustained high (160 μg/ml) heparin levels had toxic effects, while the same concentration of CSB exhibited more rapid early proliferation of CD34+ cells. In conclusion, this culture system has demonstrated the ability to produce simultaneous proliferation and CD34+ cell enrichment of a partially purified cord blood population by controlling the nature and levels of GAG moieties to which the cells are exposed. The results indicate that specific GAGs can significantly influence the growth and differentiation characteristics of cultured CD34+ cells.