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Functional, phenotypic and molecular characterization of cytokine low‐responding circulating CD34 + haemopoietic progenitors
Author(s) -
Pierelli Luca,
Scambia Giovanni,
Fattorossi Andrea,
Bonanno Giuseppina,
Battaglia Alessandra,
Rumi Carlo,
Marone Maria,
Mozzetti Simona,
Rutella Sergio,
Menichella Giacomo,
Romeo Virgilio,
Mancuso Salvatore,
Leone Giuseppe
Publication year - 1998
Publication title -
british journal of haematology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.907
H-Index - 186
eISSN - 1365-2141
pISSN - 0007-1048
DOI - 10.1046/j.1365-2141.1998.00885.x
Subject(s) - stem cell factor , microbiology and biotechnology , cd34 , biology , cytokine , progenitor cell , cell sorting , stem cell , flow cytometry , immunology
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‐SE bright ) showed a similar surface antigen expression to starting, freshly isolated CD34 + cells. Conversely, cells which experienced more than five divisions (CFDA‐SE dim ) showed a differentiating behaviour, down‐regulating CD34 antigen and acquiring differentiation markers. CFDA‐SE bright cells were significantly enriched in CD105 (endoglin) positive precursors as compared to both freshly isolated CD34 + and CFDA‐SE dim cells. Functional analysis indicated that CFDA‐SE bright had a 3‐fold and 10‐fold greater cumulative cloning efficiency as compared to freshly isolated CD34 + cells and CFDA‐SE dim cells, respectively. CFDA‐SE bright 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‐SE bright cells as compared to freshly isolated CD34 + and CFDA‐SE dim cells. This study indicates that cytokine low‐responding circulating CD34 + cells (CFDA‐SE bright cells) represent a functionally, phenotypically and molecularly distinct multipotent progenitor population with biological properties associated with primitive precursors.