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

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 g/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.

Glycosaminoglycans enhance osteoblast differentiation of bone marrow derived human mesenchymal stem cells

“…Although this result clearly indicates the stringent binding specificity of these GAGs to growth factors or other regulatory molecules, we were not able to establish the relation between the position of the sulphate group and osteogenic potential or elucidate the exact mechanism of regulation of osteogenesis by these ECM components. Similar studies have reported the difference in the functional properties of these ECM components in a hematopoietic cell culture experiment (Madihally et al ., ). They observed a difference in the response to C4S and C6S by hematopoietic cell populations.…”

Hematopoietic stem and progenitor cells in adhesive microcavities

“…We propose that the greater engagement of adhesion receptors in the small microcavities is the main mechanism responsible for the HSC behavior described. However, given that spatial constriction can have a pronounced effect on cell proliferation and differentiation, 32,33 we cannot rule out additional contributions from either mechanical constraints affecting cell morphology or paracrine signaling activated by cell-cell contacts. In technical terms, our results demonstrate that microstructured adhesive culture carriers can support HSC quiescence in vitro if the size of the constraining cavity is appropriate to host individual cells.…”

Expansion of hematopoietic stem/progenitor cells

“…Li et al reported that murine neonatal neural cell line C17.2 significantly increased the expansion of total nucleated cells, CD341 cells, and multilineage colony forming cells at a noncontact condition [8]. Moreover, porcine microvascular endothelial cell [9], human yolk sac-derived endothelial cells (hYSECs) [10], umbilical vein endothelial cells (HUVEC) [11], osteoblasts [12], and extracellular matrix [13] have also been reported to play a significant role in HSPCs expansion. To improve the hematopoietic supporting ability, there is a recent trend toward using MSC that have been immortalized or engineered to over-express various cytokines or signaling molecules, and significantly enhanced expansion have been achieved (Table I).…”