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Ultrastructural and phenotypic analysis of in vitro erythropoiesis from human cord blood CD34+ cells

Title
Ultrastructural and phenotypic analysis of in vitro erythropoiesis from human cord blood CD34+ cells
Authors
Kie J.-H.Jung Y.-J.Woo S.-Y.Ryu K.-H.Park H.-Y.Chung W.-S.Seoh J.-Y.
Ewha Authors
서주영정화순유경하우소연
SCOPUS Author ID
서주영scopus; 정화순scopus; 유경하scopus; 우소연scopus
Issue Date
2003
Journal Title
Annals of Hematology
ISSN
0939-5555JCR Link
Citation
vol. 82, no. 5, pp. 278 - 283
Indexed
SCI; SCIE; SCOPUS WOS scopus
Abstract
Erythropoietin (EPO) induces erythropoiesis in vitro as well as in vivo, and the process of erythroid differentiation has been explored phenotypically and morphologically. However, morphological analysis of in vitro erythropoiesis of human hematopoietic progenitor cells at the ultrastructural level has not been reported before. In the present study, we have traced the ultrastructural changes of erythroid differentiation during ex vivo expansion of human cord blood (CB) CD34+ cells in the presence of EPO by electron microscopy (EM), along with concurrent phenotypic analysis. CD34+ cells purified from ten CBs by immunomagnetic selection were cultured in serum-free essential media in the presence of a combination of the several cytokines including EPO, thrombopoietin, flt3-ligand (FL), stem cell factor (SCF), granulocyte colony-stimulating factor, interleukin (IL)-3 and/or IL-11. Phenotypic analysis was performed by flow cytometric analysis for erythroid markers, including glycophorin C (GPC), Kell-related, glycophorin A (GPA , band 3, Lub , and RhD. Ultrastructural analysis was performed by electron-microscopic examination of the cultured cells stained with uranyl acetate and lead citrate. Phenotypic analysis revealed that in the absence of EPO, genuine erythroid fraction expressing the typical pattern of erythroid markers did not appear. The order of the above markers expressed in the cultured cells in the presence of EPO was GPC, Kell-related, GPA, band 3, Lub, and RhD, irrespective of the type of cytokine added. Of the cytokines used in combination with EPO, FL + IL-3 was the most efficient in inducing erythroid differentiation, which was followed by SCF + IL-3. EM examination demonstrated complete process of erythroid development from pronormoblasts to reticulocytes with nuclei having been extruded and mature erythrocytes. These results suggest that morphologically intact erythrocytes could be produced by ex vivo expansion of CB CD34+ cells using EPO.
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의학전문대학원 > 의학과 > Journal papers
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