Growth factors G-CSF and GM-CSF differentially preserve chemotaxis of neutrophils aging in vitro

Baruch Wolach*, Luc J.W. van der Laan, Nikolai A. Maianski, Anton T.J. Tool, Robin van Bruggen, Dirk Roos, Taco W. Kuijpers

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

21 Citations (Scopus)


Objective: The ability of human neutrophils to migrate was studied during culture in vitro. Methods: Neutrophils were isolated from human blood and cultured at 37°C. Apoptosis was determined by Annexin-V fluorescein isothiocyanate binding. Receptor expression was measured by fluorescence in situ hybridization analysis with monoclonal antibodies. Migration was assessed with Transwell Fluoroblock inserts and calcein-stained neutrophils. Extracellular signal-regulated kinase 1/2 (ERK-1/2) activation was determined with monoclonal antibody against phosphorylated ERK-1/2. Results: Upon culture, untreated neutrophils downregulated the chemotaxin receptors FPR, CXC chemokine receptor 1, and CXC chemokine receptor 2 and lost the ability to migrate to formyl-methionyl-leucyl-phenylalanin, interleukin 8 (IL-8), and C5a. In contrast, expression of CXCR4 was induced; this receptor was able to signal (increase in intracellular free calcium ions [Ca2+]i, ERK-1/2 activation) but was nonfunctional (no chemotaxis to stromal cell-derived factor-1α). The myeloid growth factors granulocyte colony-stimulating factor (G-CSF) and granulocyte-macrophage colony-stimulating factor (GM-CSF) retarded the process of functional decay during cell culture. However, while preserving chemotaxis of neutrophils toward formyl-methionyl-leucyl-phenylalanin or C5a, GM-CSF-in contrast to G-CSF-did not preserve chemotaxis toward IL-8, with a corresponding downregulation of the IL-8 receptors. The decay in neutrophil chemotaxis occurred prior to detectable phosphatidylserine (PS)-exposure. In contrast, the induction of [Ca2+]i rises and ERK-1/2 activation correlated with chemotaxin receptor expression unless the cells were truly apoptotic. Conclusion: Neutrophils aging in vitro lose their chemotactic capacity. Functional decay starts prior to PS exposure and can be partially prevented by G-CSF and GM-CSF, in a differential fashion. These growth factors act by increasing the number of viable neutrophils, by altering the levels of chemotaxin receptor expression, and-independently-by affecting signaling cascades.

Original languageEnglish
Pages (from-to)541-550
Number of pages10
JournalExperimental Hematology
Issue number4
Publication statusPublished - 1 Apr 2007
Externally publishedYes

Bibliographical note

Funding Information:
This study was supported by NWO, the Netherlands Organization for Scientific Research (to B.W. with a visiting-scientist grant and to L.v.d.L. with a fellowship).


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