Functional, phenotypic and molecular characterization of cytokine low‐responding circulating CD34+ haemopoietic progenitors
Open Access
- 1 September 1998
- journal article
- research article
- Published by Wiley in British Journal of Haematology
- Vol. 102 (5) , 1139-1150
- https://doi.org/10.1046/j.1365-2141.1998.00885.x
Abstract
Circulating CD34+ cell populations characterized by a low rate (up to five) or high rate (more than five) of cell divisions were isolated from 8 d cultures in the presence of stem cell factor (SCF), interleukin‐3 (IL‐3), granulocyte‐macrophage colony stimulating factor (GM‐CSF), granulocyte colony stimulating factor (G‐CSF), erythropoietin (EPO), Flt3 ligand and Peg‐rHu megakaryocyte growth and development factor (Peg‐rHuMGDF) using the fluorescent dye 5,6‐carboxyfluorescein diacetate succinimidyl ester (CFDA‐SE) and flow cytometric cell sorting. Phenotypic characterization of cells which had experienced up to five divisions (CFDA‐SEbright) showed a similar surface antigen expression to starting, freshly isolated CD34+ cells. Conversely, cells which experienced more than five divisions (CFDA‐SEdim) showed a differentiating behaviour, down‐regulating CD34 antigen and acquiring differentiation markers. CFDA‐SEbright cells were significantly enriched in CD105 (endoglin) positive precursors as compared to both freshly isolated CD34+ and CFDA‐SEdim cells. Functional analysis indicated that CFDA‐SEbright had a 3‐fold and 10‐fold greater cumulative cloning efficiency as compared to freshly isolated CD34+ cells and CFDA‐SEdim cells, respectively. CFDA‐SEbright cells retained the vast majority of LTC‐IC and showed a LTC‐IC frequency 2.8‐fold higher than that found in freshly isolated CD34+ cells. RT‐PCR and Western blot analyses showed significantly higher bcl‐2 RNA and protein levels in CFDA‐SEbright cells as compared to freshly isolated CD34+ and CFDA‐SEdim cells. This study indicates that cytokine low‐responding circulating CD34+ cells (CFDA‐SEbright cells) represent a functionally, phenotypically and molecularly distinct multipotent progenitor population with biological properties associated with primitive precursors.Keywords
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