The in vitro production and characterization of neutrophils from embryonic stem cells.
An embryonic stem (ES) cell/OP9 coculture system for the effective production of functional neutrophils is described. A 3-step differentiation strategy was developed that uses liquid culture, enabling reliable and abundant production of neutrophils at high purity without the need of sorting for isolation of mature neutrophils. Use of the OP9 stromal cell line significantly enhances the number, percentage, and duration of differentiated neutrophils produced from embryonic stem cells. Effective and sustained differentiation of ES cells into neutrophils provides a useful model system for studying neutrophil differentiation and function and the factors that regulate them. Morphologic and functional evaluation of these ES-derived neutrophils indicates that large numbers of mature neutrophils can be produced from pluripotent ES cells in vitro. Specifically, their morphology, ability to produce superoxides, flux calcium, undergo chemotaxis in response to macrophage inflammatory protein 2 (MIP-2), stain for the granulocyte-specific marker-specific chloroacetate esterase, and contain the neutrophil-specific markers Gr-1 and the mouse neutrophil-specific antigen indicates that they are comparable with purified mouse bone marrow neutrophils. They also express gelatinase and lactoferrin granule proteins. During the differentiation of these ES-derived neutrophils, regional areas of neutrophil production can be identified that have been designated as neutrophil generating regions (NGRs).
Lieber
Jonathan G
JG
National Jewish Medical and Research Center, 1400 Jackson St, Denver, CO 80206, USA. lieberjg@njc.org
Webb
Saiphone
S
Suratt
Benjamin T
BT
Young
Scott K
SK
Johnson
Gary L
GL
Keller
Gordon M
GM
Worthen
G Scott
GS
D000818:Animals; D002454:Cell Differentiation; D002460:Cell Line; D002469:Cell Separation; D002634:Chemotaxis, Leukocyte; D018920:Coculture Techniques; D048728:MAP Kinase Kinase Kinase 1; D020930:MAP Kinase Kinase Kinases / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D009154:Mutation; D009504:Neutrophils / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D013234:Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D017154:Stromal Cells; D013481:Superoxides / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D013481:Superoxides; D048728:MAP Kinase Kinase Kinase 1; D020930:MAP Kinase Kinase Kinases; C482578:Map3k1 protein, mouse; D013755:Tetradecanoylphorbol Acetate