Pegylated granulocyte colony-stimulating factor mobilizes CD34+ cells with different stem and progenitor subsets and distinct functional properties in comparison with unconjugated granulocyte colony-stimulating factor.
BACKGROUND
Pegylated granulocyte colony-stimulating factor (G-CSF) has recently been introduced as a new compound for mobilization of CD34(+) hematopoietic stem and progenitor cells. In this study, we compared the molecular and functional characteristics of CD34(+) cells mobilized by pegylated G-CSF with those mobilized by unconjugated G-CSF.


METHODS
Gene expression of immunomagnetically enriched CD34(+) cells from leukapheresis products of patients who were given pegylated-G-CSF or unconjugated G-CSF was analyzed using Affymetrix HG Focus microarrays and quantitative reverse transcriptase polymerase chain reaction. Flow cytometry and fluorescence activated cell sorting was conducted to assess the CD34(+) subset composition and to obtain Lin(-), CD34(+), CD38(-) hematopoietic stem cells. Cell cycle assays and clonogenic assays were performed for functional corroboration.


RESULTS
Pegylated G-CSF and unconjugated G-CSF mobilized CD34(+) and hematopoietic stem cells with different molecular phenotypes and functional properties. The CD34(+) cells mobilized by pegylated G-CSF had higher expression levels of genes indicative of early hematopoiesis, including HOXA9, MEIS1 and GATA3. We found lower expression of genes characteristic of erythroid and later stages of myeloid differentiation and a lower functional burst-forming unit erythroid/colony-forming unit-granulocyte-macrophage ratio. Consistently, greater numbers of hematopoietic stem cells and common myeloid progenitors and fewer megakaryocyte-erthrocyte progenitors were found in the pegylated-G-CSF-mobilized CD34(+) cells. Additionally, sorted pegylated-G-CSF-mobilized hematopoietic stem cells displayed higher expression of HOXA9 in comparison to G-CSF-mobilized hematopoietic stem cells. In line with the gene expression data, CD34(+) cells mobilized by pegylated G-CSF, as well as sorted hematopoietic stem cells, showed a significantly greater cell cycle activity.


CONCLUSIONS
Stimulation with pegylated-G-CSF or G-CSF results in different expression of key regulatory genes and different functional properties of mobilized hematopoietic stem cells as well as their progeny, a finding that might be relevant for the application of these cells in blood stem cell transplantation.
Bruns
Ingmar
I
Dept. of Hematology, Oncology and Clinical Immunology, Heinrich-Heine-University. Moorenstr. 5, 40225 Düsseldorf, Germany. brunsin@med.uni-duessel-dorf.de
Steidl
Ulrich
U
Fischer
Johannes C
JC
Czibere
Akos
A
Kobbe
Guido
G
Raschke
Sascha
S
Singh
Raminder
R
Fenk
Roland
R
Rosskopf
Michael
M
Pechtel
Sabrina
S
von Haeseler
Arndt
A
Wernet
Peter
P
Tenen
Daniel G
DG
Haas
Rainer
R
Kronenwett
Ralf
R
D000328:Adult; D000368:Aged; D018952:Antigens, CD34 / Q000032:analysis; D002453:Cell Cycle; D002455:Cell Division / Q000235:genetics; D003114:Colony-Forming Units Assay; D005260:Female; D000069585:Filgrastim; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D016179:Granulocyte Colony-Stimulating Factor / Q000008:administration & dosage / Q000494:pharmacology*; D006410:Hematopoiesis / Q000235:genetics; D019650:Hematopoietic Stem Cell Mobilization / Q000379:methods*; D006412:Hematopoietic Stem Cells / Q000145:classification / Q000166:cytology / Q000187:drug effects*; D006801:Humans; D018189:Immunomagnetic Separation; D007937:Leukapheresis; D008297:Male; D008875:Middle Aged; D009101:Multiple Myeloma / Q000097:blood / Q000628:therapy; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D011092:Polyethylene Glycols; D011994:Recombinant Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction
D018952:Antigens, CD34; D009363:Neoplasm Proteins; D011994:Recombinant Proteins; D016179:Granulocyte Colony-Stimulating Factor; C455861:pegfilgrastim; D011092:Polyethylene Glycols; D000069585:Filgrastim