Up-regulation of neutral endopeptidase (CALLA) in human neutrophils by granulocyte-macrophage colony-stimulating factor

1993 ◽  
Vol 53 (6) ◽  
pp. 685-690 ◽  
Author(s):  
Jon C. Connelly ◽  
Robin Chambless ◽  
David Holiday ◽  
Kim Chittenden ◽  
Alice R. Johnson
Keyword(s):  
Neutral Endopeptidase ◽  
Human Neutrophils ◽  
Colony Stimulating Factor ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

scholarly journals Biologic properties in vitro of a recombinant human granulocyte- macrophage colony-stimulating factor

Blood ◽  
1986 ◽  
Vol 67 (1) ◽  
pp. 37-45 ◽  
Author(s):  
D Metcalf ◽  
CG Begley ◽  
GR Johnson ◽  
NA Nicola ◽  
MA Vadas ◽  
...  
Keyword(s):  
Colony Formation ◽  
Specific Activity ◽  
Human Neutrophils ◽  
Colony Stimulating Factor ◽  
Direct Stimulation ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor ◽  
Marrow Cultures

Recombinant human granulocyte-macrophage colony-stimulating factor (rH GM-CSF) was purified to homogeneity from medium conditioned by COS cells transfected with a cloned human GM-CSF cDNA and shown to be an effective proliferative stimulus in human marrow cultures for GM and eosinophil colony formation. The specific activity of purified rH GM- CSF in human marrow cultures was calculated to be at least 4 X 10(7) U/mg protein. Clone transfer experiments showed that this proliferation was due to direct stimulation of responding clonogenic cells. Acting alone, rH GM-CSF did not stimulate erythroid colony formation, but in combination with erythropoietin, increased erythroid and multipotential colony formation in cultures of peripheral blood cells. rH GM-CSF had no proliferative effects on adult or fetal murine hematopoietic cells, did not induce differentiation in murine myelomonocytic WEHI-3B cells, and was unable to stimulate the survival or proliferation of murine hematopoietic cell lines dependent on murine multi-CSF (IL 3). rH GM- CSF stimulated antibody-dependent cytolysis of tumor cells by both mature human neutrophils and eosinophils and increased eosinophil autofluorescence and phagocytosis by neutrophils. From a comparison of these effects with those of semipurified preparations of human CSF alpha and -beta, it was concluded that rH GM-CSF exhibited all the biologic activities previously noted for CSF alpha.

Receptor expression and oxidase activity in human neutrophils: Regulation by granulocyte-macrophage colony-stimulating factor and dependence upon protein biosynthesis

1990 ◽  
Vol 10 (4) ◽  
pp. 393-401 ◽  
Author(s):  
Steven W. Edwards ◽  
Fiona Watson ◽  
Ronald MacLeod ◽  
John Davies
Keyword(s):  
De Novo ◽  
Protein Biosynthesis ◽  
Neutrophil Function ◽  
Membrane Receptors ◽  
Receptor Expression ◽  
Human Neutrophils ◽  
Colony Stimulating Factor ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Incubation of human bloodstream neutrophils with 50 u/ml recombinant granulocyte-macrophage colony-stimulating factor (rGM-CSF) “primed” the respiratory burst (as assessed by fMet-Leu-Phe stimulated luminol-dependent chemiluminescence) and resulted in a rapid (within 15 min) upregulation of expression of CD11b and CD18 (as measured by FACS analysis). This rapid “priming” and modulation of receptor expression was not inhibited by cycloheximide and hence appeared to be independent of de novo protein biosynthesis. When neutrophils were incubated for up to 5 h in culture, the fluorescence distributions of CD11b and CD18 declined indicating the loss of expression of these receptors as the neutrophils aged, but in rGM-CSF treated suspensions receptor expression was maintained. When neutrophils were incubated in the presence of cycloheximide, they progressively lost their ability to generate reactive oxidants in response to fMet-Leu-Phe so that by 5 h incubation with this inhibitor they could only generate about 25% of the oxidative response stimulated in untreated cells, and the expression of CD16 and CD18 was grossly impaired. Similar effects were observed in rGM-CSF treated suspensions except that cycloheximide required longer incubation times (typically 4–5 h) before impairment of function or receptor expression occurred. These data show that de novo protein biosynthesis is required for both the maintenance of neutrophil function and also for the continued expression of some plasma membrane receptors.

scholarly journals β2-Integrins and Acquired Glycoprotein IIb/IIIa (GPIIb/IIIa) Receptors Cooperate in NF-κB Activation of Human Neutrophils

2007 ◽  
Vol 282 (38) ◽  
pp. 27960-27969 ◽  
Author(s):  
Birgit Salanova ◽  
Mira Choi ◽  
Susanne Rolle ◽  
Maren Wellner ◽  
Friedrich C. Luft ◽  
...  
Keyword(s):  
Tyrosine Kinases ◽  
Surface Expression ◽  
Human Neutrophils ◽  
Colony Stimulating Factor ◽  
Mobility Shift ◽  
Reverse Transcription Pcr ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor ◽  
Β2 Integrins

Microparticles from various cells are generated during inflammation. Platelet-derived microparticles (PMPs) harbor receptors that are not genuinely expressed by neutrophils. We tested whether or not functional glycoprotein IIb/IIIa (GPIIb/IIIa) receptors can be acquired by neutrophils via PMPs and whether these receptors participate in pro-inflammatory signaling. Surface expression was analyzed by flow cytometry and confocal microscopy. NF-κB activation was analyzed by Western blot experiments, electrophoretic mobility shift assays, and reverse transcription-PCR. Cell adhesion and spreading were estimated by myeloperoxidase assay and light microscopy. We found that PMPs transfer GPIIb/IIIa receptors to isolated and whole blood neutrophils via PMPs. We used specific antibodies in granulocyte macrophage colony-stimulating factor-treated neutrophils and observed that acquired GPIIb/IIIa receptors co-localized with β2-integrins and cooperated in NF-κB activation. We show that Src and Syk non-receptor tyrosine kinases, as well as the actin cytoskeleton, control NF-κB activation. In contrast to NF-κB, acquisition of GPIIb/IIIa receptors was not necessary to induce adhesion to fibronectin or phosphatidylinositol 3-kinase/Akt signaling. When granulocyte macrophage colony-stimulating factor-stimulated neutrophils were incubated on fibronectin, strong NF-κB activation was observed, but only after loading with PMPs. Blocking either β2-integrins or GPIIb/IIIa receptors abrogated this effect. Therapeutic GPIIb/IIIa inhibitors were similarly effective. The compounds also inhibited NF-κB-dependent tumor necrosis factor-α mRNA up-regulation. The data implicate GPIIb/IIIa receptors as new therapeutic targets in neutrophil-induced inflammation.

scholarly journals Granulocyte-macrophage colony-stimulating factor (GM-CSF) promotes phosphorylation and an increase in the activity of cytosolic phospholipase A2 in human neutrophils

1996 ◽  
Vol 313 (2) ◽  
pp. 503-508 ◽  
Author(s):  
Nabeel NAHAS ◽  
Waltraut H. WATERMAN ◽  
Ramadan I. SHA'AFI
Keyword(s):  
Phospholipase A2 ◽  
Cytosolic Phospholipase A2 ◽  
Time Dependent ◽  
Human Neutrophils ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Cytosolic Phospholipase ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Incubation of human neutrophils with 500 pM granulocyte-macrophage colony-stimulating factor (GM-CSF) results in a rapid and time-dependent increase in the phosphorylation of cytosolic phospholipase A2 (cPLA2), which was reflected in a slower electrophoretic mobility of the enzyme. The GM-CSF-induced phosphorylation of cPLA2 was accompanied by a parallel and time-dependent increase in the enzyme activity. Preincubation of neutrophils with the tyrosine kinase inhibitor genistein caused inhibition of the GM-CSF-stimulated phosphorylation and activity of cPLA2. Immunoprecipitation of the enzyme following incubation of neutrophils with [32P]Pi shows that cPLA2 is phosphorylated by GM-CSF. Potato acid phosphatase caused dephosphorylation of the enzyme, indicating that cPLA2 is indeed phosphorylated by GM-CSF. The subcellular distribution of cPLA2 in GM-CSF-stimulated neutrophils revealed that the enzyme resides almost completely in the cytosolic fraction. Addition of Ca2+ to the lysis buffer before homogenization results in the translocation of the phosphorylated and the dephosphorylated forms of the enzyme to the membranes. Translocation of cPLA2 was also achieved after incubation with 0.1 μM N-formylmethionyl-leucyl-phenylalanine (fMLP) after GM-CSF stimulation and when neutrophils were challenged with the Ca2+ ionophore A23187. EDTA and EGTA were unable to solubilize the translocated enzyme from the neutrophil membranes, indicating that cPLA2 is attached to the membranes by strong bonds and not merely due to ionic forces exerted by Ca2+. The inability of GM-CSF to promote arachidonic acid mobilization is probably due to the fact that GM-CSF does not cause an increase in intracellular Ca2+, which is necessary for the translocation of the enzyme to the membranes where its substrate(s) reside.

scholarly journals Direct stimulation by tyrosine phosphorylation of microtubule-associated protein (MAP) kinase activity by granulocyte-macrophage colony-stimulating factor in human neutrophils

1993 ◽  
Vol 291 (1) ◽  
pp. 211-217 ◽  
Author(s):  
J Gomez-Cambronero ◽  
J M Colasanto ◽  
C K Huang ◽  
R I Sha'afi
Keyword(s):  
Okadaic Acid ◽  
Kinase Activity ◽  
Human Neutrophils ◽  
Colony Stimulating Factor ◽  
Tyrosine Residues ◽  
Gm Csf ◽  
Mapk Activity ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Human polymorphonuclear neutrophils exhibit a low level of the microtubule-associated protein kinase (MAPK) activity. This enzymic activity is enhanced up to 3-fold upon cell stimulation with the human haematopoietic hormone granulocyte-macrophage colony-stimulating factor (GM-CSF). This is demonstrated both in whole-cell lysates and in DEAE-anion-exchange semi-purified fractions prepared from GM-CSF-stimulated neutrophils, by assaying the kinase activity against either myelin basic protein or a phosphoacceptor peptide that bears the specific phosphorylation site of the MAPK natural substrate. Similarly, phosphorylation of MAPK in tyrosine residues, as found in immunoblots using anti-phosphotyrosine antibodies, follows similar time- and dose-response curves as the kinase activation. Pretreatment of the cells with the tyrosine kinase inhibitor genistein abrogates the above-mentioned effect, whereas the phosphatase inhibitor okadaic acid enhances both the basal and the GM-CSF-stimulated kinase activities. Likewise, MAPK tyrosine phosphorylation is diminished in genistein-treated neutrophils, and enhanced in okadaic acid-treated cells. We conclude that MAPK activity is present in human neutrophils, and that it is stimulated by GM-CSF. This stimulation of the activity is most likely due to the phosphorylation of MAPK in tyrosine residues triggered upon binding of GM-CSF to its receptors.

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