scholarly journals Granulocyte colony-stimulating factor administration to healthy volunteers: analysis of the immediate activating effects on circulating neutrophils

Blood ◽  
1994 ◽  
Vol 84 (11) ◽  
pp. 3885-3894 ◽  
Author(s):  
M de Haas ◽  
JM Kerst ◽  
CE van der Schoot ◽  
J Calafat ◽  
CE Hack ◽  
...  
Keyword(s):  
Healthy Volunteers ◽  
Plasma Levels ◽  
Subcutaneous Administration ◽  
Secretory Vesicles ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Specific Granules ◽  
Stimulating Factor

In four healthy volunteers, we analyzed in detail the immediate in vivo effects on circulating neutrophils of subcutaneous administration of 300 micrograms of granulocyte colony-stimulating factor (G-CSF). Neutrophil activation was assessed by measurement of degranulation. Mobilization of secretory vesicles was shown by a decrease in leukocyte alkaline phosphatase content of the circulating neutrophils. Furthermore, shortly postinjection, Fc gamma RIII was found to be upregulated from an intracellular pool that we identified by immunoelectron microscopy as secretory vesicles. Intravascular release of specific granules was shown by increased plasma levels of lactoferrin and by upregulation of the expression of CD66b and CD11b on circulating neutrophils. Moreover, measurement of fourfold elevated plasma levels of elastase, bound to its physiologic inhibitor alpha 1- antitrypsin, indicated mobilization of azurophil granules. However, no expression of CD63, a marker of azurophil granules, was observed on circulating neutrophils. G-CSF--induced mobilization of secretory vesicles and specific granules could be mimicked in whole blood cultures in vitro, in contrast to release of azurophil granules. Therefore, we postulate that the most activated neutrophils leave the circulation, as observed shortly postinjection, and undergo subsequent stimulation in the endothelial microenvironment, resulting in mobilization of azurophil granules. Our data demonstrate that G-CSF should be regarded as a potent immediate activator of neutrophils in vivo.

scholarly journals Effective stimulation of donors for granulocyte transfusions with recombinant methionyl granulocyte colony-stimulating factor [see comments]

Blood ◽  
1993 ◽  
Vol 81 (11) ◽  
pp. 2866-2871 ◽  
Author(s):  
CB Caspar ◽  
RA Seger ◽  
J Burger ◽  
J Gmur
Keyword(s):  
Subcutaneous Administration ◽  
Neutrophil Granulocytes ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Superoxide Anion Production ◽  
Consistent Increase ◽  
Pmn Functions ◽  
Stimulating Factor

Abstract Effective granulocyte transfusion (GT) therapy has been hampered by the low yield of neutrophil granulocytes (PMN) obtainable from normal donors even by use of corticosteroid prestimulation, hydroxyethyl starch (HES), and modern leukapheresis (LA) techniques. To increase the PMN yield we performed LA in 22 healthy volunteer donors after a single subcutaneous administration of 300 micrograms of granulocyte colony- stimulating factor (G-CSF) 12 to 16 hours before LA. Five to 7 L of blood was processed within 1.9 to 3 hours using the standard CS- 3000Plus (Baxter, Deerfield, IL) LA protocol including HES. The mean number of PMN harvested was 44.32 +/- 15.5 x 10(9), corresponding to 6.88 +/- 2.1 x 10(9)/L of blood processed. In the final product PMN functions (in vitro: chemotaxis, phagocytosis, chemiluminescence, superoxide anion production; in vivo: chemiluminescence, half-life) were at least normal. In all donors G-CSF induced a consistent increase of white blood cell (mean 16.46 +/- 3.8 x 10(9)/L) and PMN counts (15.94 +/- 3.6 x 10(9)/L). No G-CSF-related side effects were observed and LA was well tolerated. G-CSF prestimulation allows to harvest three to five times higher numbers of functionally normal PMN by LA compared with corticosteroid pretreatment. This may help to overcome one of the major limitations of an effective PMN support.

scholarly journals Effective stimulation of donors for granulocyte transfusions with recombinant methionyl granulocyte colony-stimulating factor [see comments]

Blood ◽  
1993 ◽  
Vol 81 (11) ◽  
pp. 2866-2871 ◽  
Author(s):  
CB Caspar ◽  
RA Seger ◽  
J Burger ◽  
J Gmur
Keyword(s):  
Subcutaneous Administration ◽  
Neutrophil Granulocytes ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Superoxide Anion Production ◽  
Consistent Increase ◽  
Pmn Functions ◽  
Stimulating Factor

Effective granulocyte transfusion (GT) therapy has been hampered by the low yield of neutrophil granulocytes (PMN) obtainable from normal donors even by use of corticosteroid prestimulation, hydroxyethyl starch (HES), and modern leukapheresis (LA) techniques. To increase the PMN yield we performed LA in 22 healthy volunteer donors after a single subcutaneous administration of 300 micrograms of granulocyte colony- stimulating factor (G-CSF) 12 to 16 hours before LA. Five to 7 L of blood was processed within 1.9 to 3 hours using the standard CS- 3000Plus (Baxter, Deerfield, IL) LA protocol including HES. The mean number of PMN harvested was 44.32 +/- 15.5 x 10(9), corresponding to 6.88 +/- 2.1 x 10(9)/L of blood processed. In the final product PMN functions (in vitro: chemotaxis, phagocytosis, chemiluminescence, superoxide anion production; in vivo: chemiluminescence, half-life) were at least normal. In all donors G-CSF induced a consistent increase of white blood cell (mean 16.46 +/- 3.8 x 10(9)/L) and PMN counts (15.94 +/- 3.6 x 10(9)/L). No G-CSF-related side effects were observed and LA was well tolerated. G-CSF prestimulation allows to harvest three to five times higher numbers of functionally normal PMN by LA compared with corticosteroid pretreatment. This may help to overcome one of the major limitations of an effective PMN support.

scholarly journals In vitro and in vivo effects of recombinant human granulocyte colony-stimulating factor on neutrophil functions.

1989 ◽  
Vol 35 (6) ◽  
pp. 647-652
Author(s):  
Akimichi Ohsaka ◽  
Seiichi Kitagawa ◽  
Akira Yuo ◽  
Takashi Obata ◽  
Youichi Amemiya ◽  
...  
Keyword(s):  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
In Vivo Effects ◽  
Stimulating Factor

scholarly journals Neutrophils express the high affinity receptor for IgG (Fc gamma RI, CD64) after in vivo application of recombinant human granulocyte colony- stimulating factor

Blood ◽  
1991 ◽  
Vol 78 (4) ◽  
pp. 885-889 ◽  
Author(s):  
R Repp ◽  
T Valerius ◽  
A Sendler ◽  
M Gramatzki ◽  
H Iro ◽  
...  
Keyword(s):  
Fc Receptors ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
High Affinity ◽  
Healthy Donors ◽  
High Affinity Receptor ◽  
Affinity Receptor ◽  
Stimulating Factor

Abstract Fc receptors are important effector molecules of neutrophilic granulocytes (polymorphonuclear neutrophils [PMN]), connecting phagocytic cells and the specific immune response. Neutrophils from healthy donors express the low-affinity receptors for IgG Fc gamma RII (CD32) and Fc gamma RIII (CD16), but not the high-affinity receptor Fc gamma RI (CD64). The latter has been found on neutrophils from patients with certain bacterial infections and can be induced in vitro after incubation with interferon-gamma. We show here that neutrophils strongly express Fc gamma RI after in vivo application of recombinant human granulocyte colony-stimulating factor (rhG-CSF). PMN from patients receiving rhG-CSF displayed higher cytotoxicity against Daudi lymphoma cells in vitro compared with control patients and with healthy donors. Fab fragments against Fc gamma RII (monoclonal antibody [MoAb] IV.3) inhibited neutrophil-mediated cytotoxicity of healthy donors but not of patients during rhG-CSF therapy. Therefore, expression of Fc receptors by PMN was investigated by flow cytometry and the mean fluorescence intensity (MFI) was compared. After staining with MoAb 32.2 against Fc gamma RL, the median MFI of neutrophils from G-CSF patients (median, 4.78; range, 2.40 to 8.50; n = 5) was significantly higher (P = .002 and P = .001, respectively) than the median MFI of patients not receiving G-CSF (median, 1.23; range, 1.01 to 1.58; n = 6) and the median MFI of healthy donors (median, 1.04; range, 0.67 to 1.12; n = 6). Fc gamma RI disappeared after the discontinuing of the G- CSF injections, but was reinduced during the next treatment cycle with rhG-CSF. The high expression of Fc gamma RI during rhG-CSF therapy correlated with enhanced cytotoxicity. In vitro incubation with rhG-CSF also enhances cytotoxicity, but only minor increments in Fc gamma RI expression were observed. Thus, during in vivo application of rhG-CSF neutrophils acquire an additional potent receptor for mediating tumor cell killing in vitro by induction of the high-affinity receptor for IgG (Fc gamma RI, CD64).

scholarly journals In vitro killing of neuroblastoma cells by neutrophils derived from granulocyte colony-stimulating factor-treated cancer patients using an anti-disialoganglioside/anti-Fc gamma RI bispecific antibody

Blood ◽  
1995 ◽  
Vol 86 (3) ◽  
pp. 1124-1130 ◽  
Author(s):  
J Michon ◽  
S Moutel ◽  
J Barbet ◽  
JL Romet-Lemonne ◽  
YM Deo ◽  
...  
Keyword(s):  
Binding Site ◽  
Cancer Patients ◽  
Bispecific Antibody ◽  
Neuroblastoma Cells ◽  
Bispecific Antibodies ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Stimulating Factor

Abstract Neutrophils isolated from cancer patients treated with granulocyte colony-stimulating factor (G-CSF) express high levels of Fc gamma RI. They exhibited an efficient killing of GD2+ neuroblastoma cells in the presence of an antidisialoganglioside (GD2) mouse monoclonal antibody (MoAb; 7A4, IgG3 kappa). However, this cytotoxicity was totally blocked by human monomeric IgG. In contrast, a bispecific antibody (7A4 bis 22/MDX-260), prepared by chemically linking an F(ab') fragment of 7A4 with an F(ab') fragment of an anti-Fc gamma RI MoAb, 22, which binds outside the Fc binding domain, triggered antibody-dependent cell cytotoxicity, even when neutrophils were preincubated with human monomeric IgG. F(ab')2 22 MoAb abrogated the MDX-260 killing without affecting that of 7A4. The 3G8 MoAb, directed against the Fc gamma RIII binding site, did not inhibit the cytotoxicity induced by either antibody. Thus, these results indicate that G-CSF-activated neutrophils exert their cytotoxic effect against neuroblastoma cells through Fc gamma RI and not Fc gamma RIII, and that the saturation of the high affinity Fc gamma RI by monomeric IgG can be overcome by the use of bispecific antibodies binding epitopes outside the IgG Fc gamma RI binding site. A combined administration of such bispecific antibodies and G-CSF may be, therefore, an efficient therapeutic approach to trigger tumor lysis by cytotoxic neutrophils in vivo.

In vivo administration of granulocyte colony-stimulating factor promotes neutrophil survival in vitro

2009 ◽  
Vol 53 (3) ◽  
pp. 129-134 ◽  
Author(s):  
Souichi Adachi ◽  
Masaru Kubota ◽  
Ying Wei Lin ◽  
Akiro Okuda ◽  
Kousaku Matsubara ◽  
...  
Keyword(s):  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Neutrophil Survival ◽  
In Vivo Administration ◽  
Stimulating Factor

Signaling by the Granulocyte-Colony Stimulating Factor Receptor Involves Jak2-Dependent Phosphorylation of Gab2.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2173-2173
Author(s):  
Lin Wang ◽  
Jia Xue ◽  
Seth J. Corey ◽  
Lisa J. Robinson
Keyword(s):  
Fusion Protein ◽  
Kinase Inhibitor ◽  
Dominant Negative ◽  
Site Directed Mutagenesis ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Erk Phosphorylation ◽  
Stimulating Factor

Abstract Granulocyte colony stimulating factor (G-CSF) is the major cytokine involved in neutrophil production. G-CSF has pleiotropic effects on myeloid cells, initially stimulating proliferation but later promoting differentiation. The specific signaling pathways that mediate the diverse effects of G-CSF remain incompletely understood. Recently, the scaffolding molecule Grb2-associated binder protein 2 (Gab2) was shown to play an important role in G-CSF induced myeloid differentiation (Zhu et al. Blood 2004). Ligand stimulation of the G-CSF receptor results in the rapid phosphorylation of Gab2, but the identity of the responsible kinases and the molecular events dependent on Gab2 phosphorylation remain unclear. Because Janus kinases (Jaks) play a central role in G-CSF signaling, we investigated the involvement of Jaks in G-CSF-stimulated Gab2 phosphorylation using the hematologic DT40 cell line stably transduced with the human G-CSF receptor (DT40GR). Antisense Jak1 and Jak2 constructs expressed in DT40GR cells each produced a marked reduction in their target Jak protein, but only antisense Jak2 reduced G-CSF-stimulated Gab2 phosphorylation. To determine whether Gab2 phosphorylation required Jak2 kinase activity, dominant negative Jak2 mutants lacking catalytic activity were expressed in the DT40GR cells. Expression of dominant negative Jak2 inhibited Gab2 phosphorylation in response to G-CSF. Similarly, treatment with the Jak2-selective kinase inhibitor AG490 markedly reduced G-CSF-dependent Gab2 phosphorylation. Co-immunoprecipitation studies further demonstrated a G-CSF- and Gab2 phosphorylation-dependent association of Jak2 with Gab2 in vivo, which was detectable by 30 seconds after G-CSF stimulation. To determine whether Gab2 was a direct substrate of Jak2, we performed in vitro phosphorylation studies using Gab2-GST fusion protein substrates. Jak2 immunoprecipitated from G-CSF-stimulated cells, but not from control cells, phosphorylated the Gab2 fusion protein. To identify potential Jak2 tyrosine phosphorylation sites in Gab2, we used site-directed mutagenesis to produce three Gab2 tyrosine mutants. Tyrosines 409, 452, and 476 were each replaced by phenylalanine (Y409F, Y452F, and Y476F). The Y452F and Y476F mutations of Gab2 each inhibited G-CSF-stimulated Jak2-dependent phosphorylation of Gab2, both in stably-transfected DT40GR cells and in transiently-transfected 293 cells also transduced with the G-CSF receptor. In contrast, G-CSF-stimulated Gab2 phosphorylation appeared unaffected by the Y409F mutation. We also evaluated downstream events in G-CSF signaling in cells expressing these Gab2 tyrosine- mutants. Akt and Erk phosphorylation following G-CSF stimulation was inhibited by both the Y452F and Y476F Gab2 mutations, but was unaffected by the Y409F mutation. These results suggest that Jak2 may mediate G-CSF differentiation signals through Stat-independent mechanisms.

scholarly journals In vivo and in vitro suppression of primary B lymphocytopoiesis by tumor-derived and recombinant granulocyte colony-stimulating factor

Blood ◽  
1993 ◽  
Vol 82 (7) ◽  
pp. 2062-2068 ◽  
Author(s):  
MY Lee ◽  
KL Fevold ◽  
K Dorshkind ◽  
R Fukunaga ◽  
S Nagata ◽  
...  
Keyword(s):  
B Cell ◽  
Stromal Cell ◽  
In Vivo Studies ◽  
Granulocyte Colony Stimulating Factor ◽  
B Lymphopoiesis ◽  
Colony Stimulating Factor ◽  
Tumor Bearing ◽  
Stimulating Factor

Transplantation of a granulocytosis-inducing murine CE mammary carcinoma into mice suppresses primary B lymphopoiesis in the marrow. The mechanisms of this tumor-induced B-cell suppression were investigated using Whitlock-Witte-type lymphoid cultures. When seeded with normal marrow progenitors, stromal cells of tumor-bearing mice supported the production of B220+ cells as well as did either stomal cells derived from control mice or the stromal cell line S17. Cultured over normal stroma, marrow cells of tumor-bearing mice depleted of adherent cells and B220+ cells generated B220+ cells as effectively as a similar cell population from control mice. However, interleukin-7- responsive progenitors, were completely depleted from the marrow of tumor-bearing mice. When conditioned medium (CM) of cloned CE tumor cells known to produce granulocyte colony-stimulating factor (G-CSF) and macrophage-CSF, or recombinant murine G-CSF was added to the cultures established with S17 cells, B220+ cell production was significantly diminished. Antiserum to murine G-CSF blocked these effects. These in vitro observations were corroborated by the elimination of marrow B220+ cells in mice injected with G-CSF. These in vitro and in vivo studies suggest that G-CSF plays an inhibitory role in primary B lymphopoiesis by blocking stromal cell-mediated differentiation of early B-cell progenitors into phenotypically recognizable B220+ pre-B cells.

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