Covalently immobilized glycosaminoglycans enhance megakaryocyte progenitor expansion and platelet release

Application of umbilical cord blood (UCB) transplantation in adults as a treatment post-chemotherapy is hampered due to delayed platelet recovery. A potential solution suggested is the transfusion of ex vivo expanded megakaryocytes (Mks) from hematopoietic stem cells (HSCs). Alternatively, large-scale production of platelets in vitro has also been attempted with the goal of transfusing them into patients with thrombocytopenia. Glycosaminoglycans (GAGs) have been shown to influence the proliferation and differentiation of HSCs. This study sought to examine the effects of immobilized GAGs on the expansion, apoptosis, and platelet release activity of CD41a+ Mk progenitors in vitro. Freshly isolated HSCs from UCB were cultured in serum-free media supplemented with thrombopoietin on GAG-derivatized chitosan membranes for 17 days. Controls consisted of uncoated and chitosan-coated wells. Wells were demidepopulated at periodic intervals and analyzed by flow cytometry. Heparin and dermatan sulfate surfaces significantly enhanced total cell and Mk cell expansion (p < 0.05) compared to both the controls. The apoptotic Mk fraction was significantly lower on GAG surfaces (p < 0.05) compared to the polystyrene control during the early stages of the culture (days 7 and 11). However, by day 17, the apoptotic Mk fraction was comparable on all surfaces. The cumulative number of platelets generated on dermatan sulfate and heparan sulfate surfaces was significantly higher (p < 0.05) than on both the controls. These results suggest that immobilized GAGs delay Mk apoptosis and thereby enhance Mk expansion and platelet production.