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Biomedical subjects

M A Vadas

Publications and source records attributed to M A Vadas.

At least 145 records · Page 8Linked to original sources
PubMed1987

Localization of the G-CSF gene on chromosome 17 proximal to the breakpoint in the t(15;17) in acute promyelocytic leukemia.

The human granulocyte-colony stimulating factor gene (G-CSF) is localized at 17q11.2-q21, the region of one of the breakpoints in the 15;17 chromosome translocation specific for acute promyelocytic leukemia (APL). As G-CSF induces differentiation and loss of tumorigenicity in myeloid leukemic cells or cell lines, it was possible that the translocation in APL involved the DNA of the G-CSF coding region or its regulatory region. In situ hybridization to chromosomes with the t(15;17) from patients with the APL translocation using a G-CSF cDNA clone revealed that the coding region of this gene is proximal to the t(15;17) breakpoint on chromosome 17. Southern analysis of DNA from patients with the APL translocation showed no differences in hybridization between normal and leukemic cells. These results indicate that the G-CSF coding sequence is not disrupted by the chromosomal rearrangement characteristic of APL.

Acute Disease↗
PubMed1987

Potentiation of delayed-type hypersensitivity by pertussigen or cyclophosphamide with release of different lymphokines.

The effects of lymphokine production of two agents known to potentiate delayed-type hypersensitivity (DTH), pertussigen (pertussis toxin) (PT) and cyclophosphamide (CY) have been investigated. These two agents were administered to immunized mice. Subsequently, lymph nodes and spleen cells were exposed to specific antigen in vitro. The resulting culture supernatants were assayed for the presence of lymphokines. Only supernatants of cells from the mice given PT contained appreciable quantities of interferon-gamma (IFN-gamma) and stimulated cells of the monocyte-like WEHI-265 cell line to produce procoagulant activator and plasminogen activator. On the other hand, CY was more effective than PT on the production of interleukin-3 (IL-3). Both adjuvants had small enhancing effects on the production of interleukin-2 (IL-2). With either adjuvant, the cell populations induced had a similarly enhanced capacity to transfer DTH. These results demonstrate that the capacity of cells to transfer DTH does not necessarily correlate with their release of particular lymphokines. The potentiation of DTH by cyclophosphamide did not depend on significantly enhanced generation of IFN-gamma, procoagulant activator, or plasminogen activator. The amount of IFN-gamma in the culture supernatants correlated with their capacity to produce procoagulant activator and plasminogen activator, whereas the amount of IL-2 and IL-3 did not.

Adjuvants, Immunologic↗
PubMed1986

Pertussigen in vivo enhances antigen-specific production in vitro of lymphokine that stimulates macrophage procoagulant activity and plasminogen activator.

Pertussigen is a protein toxin of Bordetella pertussis that acts as a powerful stimulator of the intensity and duration of delayed-type hypersensitivity (DTH) in mice. This study describes the potent in vivo effect of pertussigen on the levels of antigen-specific macrophage-activating lymphokine(s); lymphokine(s) was measured by the stimulation of macrophage procoagulant activity (mPCA), or plasminogen activator (PA) activity. Lymphoid cells were removed from immunized animals and cultured with specific antigen, keyhole limpet hemocyanin, ovalbumin, or human gamma-globulin. The culture supernatants were then incubated with the monocyte-like cell line WEHI-265 to measure mPCA or with WEHI-265 or resident peritoneal macrophages to measure PA activity. Mice were given pertussigen at the time of immunization, and the subsequent generation by lymphocyte supernatants of both of these macrophage activities proved to be greatly enhanced; the effect of pertussigen was antigen specific. Pertussigen thus induces an increase in lymphokine(s) production responsible for the in vitro increase in macrophage mPCA and PA activity and which may be responsible for some of the potent immune effects of this agent in vivo.

Animals↗
PubMed1986

Stimulation of neutrophils by tumor necrosis factor.

Human recombinant tumor necrosis factor (TNF) was shown to be a weak direct stimulus of the neutrophil respiratory burst and degranulation. The stimulation, as measured by iodination, H2O2 production, and lysozyme release, was considerably increased by the presence of unopsonized zymosan in the reaction mixture, an effect which was associated with the increased ingestion of the zymosan. TNF does not act as an opsonin but, rather, reacts with the neutrophil to increase its phagocytic activity. TNF-dependent phagocytosis, as measured indirectly by iodination, is inhibited by monoclonal antibodies (Mab) 60.1 and 60.3, which recognize different epitopes on the C3bi receptor/adherence-promoting surface glycoprotein of neutrophils. Other neutrophil stimulants, namely N-formyl-methionyl-leucyl-phenylalanine, the Ca2+ ionophore A23187, and phorbol myristic acetate, also increase iodination in the presence of zymosan; as with TNF, the effect of these stimulants is inhibited by Mab 60.1 and 60.3, whereas, in contrast to that of TNF, their stimulation of iodination is unaffected by an Mab directed against TNF. TNF may be a natural stimulant of neutrophils which promotes adherence to endothelial cells and to particles, leading to increased phagocytosis, respiratory burst activity, and degranulation.

Cytoplasmic Granules↗
PubMed1986

Murine eosinophil differentiation factor. An eosinophil-specific colony-stimulating factor with activity for human cells.

A purified murine lymphokine, eosinophil differentiation factor (EDF), was found to be a selective stimulus for the clonal proliferation and differentiation of murine eosinophil progenitor cells, establishing it as the murine eosinophil colony-stimulating factor (Eo-CSF). EDF was also active on human eosinophil progenitors and mature blood eosinophils, but had no effect on neutrophil or macrophage precursor cells, nor on blood neutrophils. In culture of human bone marrow cells, EDF stimulated equal numbers and equal sizes of eosinophil colonies to develop when compared with human placental conditioned medium, a source of human CSFs, suggesting that all responsive progenitor cells were stimulated. Clone transfer experiments and the linear relationship between number of bone marrow cells plated and colonies produced confirmed that the action of EDF was directly on eosinophil progenitor cells. EDF increased the capacity of human blood eosinophils, but not neutrophils, to kill antibody-coated tumor cells and to phagocytose serum-opsonized yeast cells. This functional activation was associated with the enhanced expression of functional antigens (GFA-1, GFA-2, and the receptor for C3bi) on eosinophils. The possession by EDF (Eo-CSF) of all the properties expected of a human eosinophil CSF raises the possibility that a human analog of this molecule exists, and is involved in the regulation of production and function of human eosinophils in vivo.

Animals↗
PubMed1986

The IIb-IIIa glycoprotein complex that mediates platelet aggregation is directly implicated in leukocyte adhesion.

Evidence is presented that the IIb-IIIa glycoprotein complex, which functions as the receptor for fibrinogen on platelets and is central to platelet aggregation, is expressed on the surface of leukocytes where it may function as a receptor for fibronectin. F(ab')2 fragments of a monoclonal antibody, 25E11, raised against activated large granular lymphocytes, inhibited killing by natural killer cells, blocked the binding of fibronectin-coated particles by monocytes, and stimulated neutrophils to exhibit increased antibody-dependent killing. Immunoprecipitation studies of leukocytes and platelets, and the ability of 25E11 to inhibit platelet aggregation, identified the antigen as an epitope on the IIb-IIIa complex. This glycoprotein thus constitutes the first example of a receptor mediating both platelet aggregation and leukocyte adhesion.

Adult↗
PubMed1986

A comparison of eosinophil-activating factor (EAF) with other monokines and lymphokines.

Monocytes from moderately eosinophilic individuals secrete material that enhances the cytotoxic activity of eosinophils against antibody-coated schistosomula of Schistosoma mansoni. This material is not a single substance, but can be fractionated into several active components of different size and different charge. Gel filtration of mononuclear cell supernatants separated the eosinophil-activating activity into a major component of molecular mass of 40 kDa and a minor component of molecular mass of less than 10 kDa. The major component exhibited further heterogeneity on fractionation by high performance liquid chromatography. The bulk of the eosinophil-activating activity could be separated from both colony-stimulating factor (CSF) alpha activity and from tumor necrosis factor (TNF) activity. However, human recombinant CSF alpha (GM-CSF), human recombinant TNF and rabbit tumor necrosis serum all had eosinophil-activating activity when tested against schistosomula. Eosinophils were not activated by interleukin 1, interleukin 2, interferon-alpha, lipopolysaccharide or phorbol myristate acetate.

Colony-Stimulating Factors↗
PubMed1986

Down-modulation of receptors for granulocyte colony-stimulating factor on human neutrophils by granulocyte-activating agents.

Purified human blood neutrophils were able to bind radioiodinated murine granulocyte-colony-stimulating factor (G-CSF) in a specific manner. This factor has previously been shown to stimulate functional activities of human and murine neutrophilic granulocytes and to be functionally analogous to human-derived CSF beta. The binding of 125I G-CSF to human neutrophils was competed for equally by unlabeled G-CSF and CSF beta but not by other CSF's. Saturation analysis indicated that human neutrophils displayed about 700-1,500 receptors for G-CSF/CSF beta per cell. Three other agents (N-formyl-methionine-leucine phenylalanine, bacterial lipopolysaccharide, and human CSF alpha) known to activate neutrophils did not compete directly for G-CSF binding sites but, in preincubation experiments at 37 degrees C, were able to down-modulate the expression of G-CSF receptors on human neutrophils in a dose- and time-dependent manner. This effect was specific since the same agents have been shown elsewhere to up-regulate the expression of other granulocyte surface antigens and other agents were much less effective at down-modulating G-CSF receptors. Since the granulocyte-activating agents increase the sensitivity of human neutrophils to G-CSF/CSF beta and mimic some of the actions of G-CSF on neutrophils, it is suggested that G-CSF receptor down-modulation might be a mechanism whereby these agents activate G-CSF receptors and thereby exert some of their effects.

Animals↗
PubMed1986

Pertussigen enhances antigen-driven interferon-gamma production by sensitized lymphoid cells.

We have studied the effects on interferon-gamma (IFN-gamma) production of pertussigen, a protein toxin from Bordetella pertussis that augments and prolongs delayed-type hypersensitivity (DTH) reactions. Lymphoid cell suspensions from immunized mice were incubated with antigen or mitogen, and the culture supernatants were assayed for IFN-gamma. The production of IFN-gamma on exposure to specific antigen or concanavalin A was greatly enhanced if mice were given pertussigen at the time of immunization. There was no detectable IFN-gamma production when cells were exposed to saline or to an irrelevant antigen. The effect of pertussigen on antigen-driven IFN-gamma production correlated with its effect on the capacity of the same cell populations to transfer DTH. The enhanced IFN-gamma production by cells from mice given pertussigen could not be attributed to an increased antigen-presenting capacity of this cell population. Production of IFN-gamma was abolished if the cells were pretreated with emetine, but not with mitomycin C, and the release of IFN-gamma was not detected in the first 8 hr of culture. After immunization with pertussigen, IFN-gamma was produced by lymph node and spleen cells from 7 days onward and both cell types produced IFN-gamma until at least 30 days after immunization. It is suggested that the augmentation of antigen-specific IFN-gamma production may contribute to the prolonged DTH reactions induced by pertussigen in vivo.

Animals↗
PubMed1986

Role of cell surface receptors in the regulation of intracellular killing of bacteria by murine peritoneal exudate neutrophils.

The role of the Fc and third component of complement (C3) receptors on mouse neutrophils in the control of killing of Proteus mirabilis, opsonized in normal mouse serum (NMS) or heated immune mouse serum (HIMS), was studied. The events following incubation of neutrophils with P. mirabilis and the events associated with bacterial killing were assayed. The respiratory burst was quantified by chemiluminescence (CL). Levels of leukocyte-associated bacteria were determined after a 20-min ingestion period as a measure of phagocytosis. Bacterial killing was measured while ingestion was allowed to continue or as a discrete process when extracellular, noningested bacteria had been removed and neutrophils with intracellular bacteria were incubated in the presence of serum. Modification of these responses in the presence of three monoclonal antibodies (MAb), NIMP-R10 and M1/70, which bind to different epitopes of the mouse C3 receptor, and 2.4G2, which binds to the mouse Fc receptor, was investigated. MAb to the C3, but not to the Fc, receptors reduced CL, ingestion, and intracellular killing of NMS-opsonized P. mirabilis. MAb to the Fc receptor diminished CL to and reduced the rate of ingestion of HIMS-opsonized bacteria. The two MAb to the C3 receptor each produced a similar inhibition of ingestion and intracellular killing of HIMS-opsonized bacteria, but they only partially blocked CL. A range of MAb preparations reactive with other murine antigens did not inhibit these events, either with NMS- or HIMS-opsonized P. mirabilis. The results suggest that C3 receptors on mouse neutrophils played a predominant role in regulation of the killing of P. mirabilis. Similar results were found for Staphylococcus aureus. C3 receptors were necessary for maximal expression of all functions culminating in bacterial kill. That MAb to the C3 receptor inhibited phagocytosis of HIMS-opsonized bacteria in similar fashion to the effect of MAb to the Fc receptor and in contrast to the lack of effect of control MAb may reflect steric hindrance of the Fc receptor by MAb binding to the C3 receptor, or it may reflect that the receptors are linked in murine neutrophils as they are in human neutrophils.

Animals↗
PubMed1986

Recombinant human granulocyte-macrophage colony-stimulating factor stimulates in vitro mature human neutrophil and eosinophil function, surface receptor expression, and survival.

A purified recombinant human granulocyte-macrophage colony stimulating factor (rH GM-CSF) was a powerful stimulator of mature human eosinophils and neutrophils. The purified rH GM-CSF enhanced the cytotoxic activity of neutrophils and eosinophils against antibody-coated targets, stimulated phagocytosis of serum-opsonized yeast by both cell types in a dose-dependent manner, and stimulated neutrophil-mediated iodination in the presence of zymosan. In addition, rH GM-CSF enhanced N-formylmethionylleucylphenylalanine(FMLP)-stimulated degranulation of Cytochalasin B pretreated neutrophils and FMLP-stimulated superoxide production. In contrast, rH GM-CSF did not promote adherence of granulocytes to endothelial cells or plastic surfaces. rH GM-CSF selectively enhanced the surface expression of granulocyte functional antigens 1 and 2, and the Mo1 antigen. rH GM-CSF induced morphological changes and enhanced the survival of both neutrophils and eosinophils by 6 and 9 h, respectively. These experiments show that granulocyte-macrophage colony stimulating factor can selectively stimulate mature granulocyte function.

Antibody-Dependent Cell Cytotoxicity↗
PubMed1986

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

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.

Animals↗
PubMed1986

Purified colony-stimulating factors enhance the survival of human neutrophils and eosinophils in vitro: a rapid and sensitive microassay for colony-stimulating factors.

The survival of purified human blood neutrophils and eosinophils was monitored using microwell cultures. Survival was enhanced in cultures containing human or murine colony-stimulating factors (CSFs). The survival of both cell types was enhanced by purified recombinant human granulocyte-macrophage CSF and partially purified preparations of the native molecule, CSF alpha. Neutrophil but not eosinophil survival was enhanced by murine granulocyte-CSF and its human analogue CSF beta. Eosinophil but not neutrophil survival was enhanced by murine eosinophil differentiation factor (eosinophil CSF). The mature cell survival provided an assay system for CSF that was 10(2) to 10(3) more sensitive than the standard assay of stimulation of colony formation in agar cultures and could be completed within 48 hours. These results demonstrated that CSF induced and enhanced survival of mature human neutrophils and eosinophils in vitro. Furthermore, the lineage specificity of purified murine CSFs was retained in their action on human cells.

Animals↗
PubMed1985

Stimulation of the adherence of neutrophils to umbilical vein endothelium by human recombinant tumor necrosis factor.

Recombinant human tumor necrosis factor (TNF) was found to enhance the adherence of human peripheral blood neutrophils to human umbilical vein endothelial (HUVE) cell monolayers in vitro. The enhancement was due to the effects both on neutrophils and HUVE cells. The effect on neutrophils was maximally induced within 5 min and did not require protein or RNA synthesis. By contrast, maximal effects on HUVE cells took 4 hr to develop and required de novo protein and RNA synthesis; however, exposure of HUVE cells to TNF for as little as 5 min was sufficient to initiate changes leading to maximal adherence of neutrophils at 4 hr. Both the effect on neutrophils and that on HUVE cells were blocked by a monoclonal antibody against TNF. TNF also rapidly induced an increased surface expression of neutrophil antigens recognized by monoclonal antibodies directed against epitopes of a glycoprotein required for optimum adherence and for complement component C3bi receptor (CR3) function. Thus, the mechanism of action of TNF may involve the regulation of expression of cell surface molecules. Our observations show that TNF induces a process central to the development of all inflammatory reactions and that both blood neutrophils and endothelial cells are targets of TNF action. The regulation of inflammatory reactions by TNF or antagonists of TNF has wide-ranging clinical implications.

Antibodies, Monoclonal↗
PubMed1985

Selective enhancement of the expression of granulocyte functional antigens 1 and 2 on human neutrophils.

The regulation of expression of two human granulocyte functional antigens (GFA-1 and GFA-2) was examined. N-formylmethionylleucylphenylalanine (fMet-Leu-Phe) caused a rapid, dose-dependent enhancement of the expression of these antigens, 2- to 4-fold within 30 min, but not of another surface structure, beta 2-microglobulin. Pretreatment of the cells with cytochalasin B at 5 micrograms/ml further enhanced the effect of fMet-Leu-Phe on the expression of GFA-2, raising its surface expression 11-fold. Lipopolysaccharide also stimulated the expression of GFA-1 and GFA-2. The effect of lipopolysaccharide was less than that of fMet-Leu-Phe and was more marked on GFA-1 than on GFA-2. Pretreatment of neutrophils with fMet-Leu-Phe not only stimulated their cytotoxic activity against antibody-coated target cells but also increased their capacity to be stimulated by monoclonal antibodies to GFA-1 and GFA-2. These findings show that the expression of functional surface structures on human neutrophils is subject to rapid and selective regulation.

Antibodies, Monoclonal↗
PubMed1985

Fc receptors on mouse neutrophils and eosinophils: antigenic characteristics, isotype specificity and relative cell membrane density measured by flow cytometry.

The antigenic characteristics, isotype specificity and density of Fc receptors (FcR) on mouse neutrophils and eosinophils were studied with the aid of the rat monoclonal antibody 2.4 G2 to the mouse macrophage FcR (Unkeless, 1979). This MAb was tested for its reactivity with mouse neutrophil and eosinophil FcR, and for its ability to block the binding of sheep erythrocytes (E) coated with mouse antibodies of different isotypes to granulocytes. The use of E conjugated with fluorescein isothiocyanate (FITC) allowed an objective read-out by flow cytometry. The MAb 2.4.G2 reacted with both neutrophil and eosinophil FcR, blocking the binding of E coated with mouse IgG1, IgG2a and IgG2b in a dose-dependent manner. Blocking was specific, since it did not occur with any of several control MAb of the same rat isotype (IgG2b) as 2.4.G2. Furthermore, the binding to E through the granulocyte receptor for complement (C) was unaffected. IgG3 was unable to promote binding of E to either neutrophils or eosinophils, although it induced high levels of binding to macrophages. These results show that: (i) neutrophil, eosinophil and macrophage FcR have antigenic similarities; (ii) neutrophils and eosinophils, in contrast to macrophages, either have a common FcR for IgG1, IgG2a and IgG2b, or have different FcR for these isotypes which share the antigenic determinant recognized by 2.4.G2; (iii) in contrast to macrophages, neutrophils and eosinophils lack the FcR for IgG3. The MAb 2.4.G2 was used in an indirect immunofluorescence assay monitored by flow cytometry to measure the relative FcR density on neutrophils and eosinophils. This assay showed that neutrophils possess about 65% more FcR than eosinophils on a cell-for-cell basis, providing an explanation for the higher binding of neutrophils to IgG-coated particles at suboptimal antibody concentrations.

Animals↗
PubMed1985

Regulation of human granulocyte function by products derived from murine tumors.

Murine EL-4 thymoma was found to produce a factor that activated antibody-dependent cell-mediated cytotoxicity by human eosinophils and neutrophils. The eosinophil-activating factor had some properties similar to human eosinophil colony-stimulating factor in being heat stable, inactivated by 2-mercaptoethanol, and cochromatographing by gel filtration with a factor that promoted the growth of pure eosinophil colonies from human bone marrow. The neutrophil-activating factor from EL-4 was also heat stable but was not inactivated by 2-mercaptoethanol and was found in two peaks on chromatography, neither having human neutrophil-CSF activity but one cochromatographing with mouse granulocyte-macrophage CSF. EL-4 therefore secretes factors that activate two types of human granulocytes. Several other murine thymomas and macrophage cell lines produced a neutrophil-activating factor, the production of which could not be correlated with that of several other known lymphokines. Murine tumors produce factors that activate human granulocytes.

Animals↗