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Biomedical subjects

A Celada

Publications and source records attributed to A Celada.

At least 19 recordsLinked to original sources
PubMed1992

Transforming growth factor-beta enhances the M-CSF and GM-CSF-stimulated proliferation of macrophages.

Transforming growth factor-beta (TGF-beta) has been shown to regulate the proliferation and function of several different cell types in the immune system. We have examined the effect of TGF-beta on the proliferation of murine macrophages in liquid culture. TGF-beta by itself did not induce proliferation of differentiated (7 days in culture) bone marrow-derived macrophages (BMM). In the presence of M-CSF, TGF-beta enhanced the proliferation of differentiated BMM and elicited peritoneal macrophages but had an inhibitory effect on the proliferation of nonadherent BMM (3 days in culture). The effect of TGF-beta was not restricted to M-CSF-dependent proliferation but was also observed for GM-CSF-dependent proliferation. The autocrine production of TGF-beta appeared to contribute to the proliferation of BMM. The addition of antibody against TGF-beta inhibited M-CSF- and GM-CSF-dependent proliferation 32% and 28%, respectively. In bone marrow, TGF-beta may be an important negative regulator of macrophage proliferation; whereas, in the tissues, TGF-beta may enhance macrophage proliferation.

Animals↗
PubMed1991

IFN-gamma induces the expression of the genes for MHC class II I-A beta and tumor necrosis factor through a protein kinase C-independent pathway.

The mechanism of gene expression for the MHC I-A beta and TNF genes was studied in murine bone marrow macrophages. The treatment of macrophages with PMA stimulated the expression of TNF, but not I-A beta, suggesting that the TNF gene is responsive to activators of protein kinase C whereas the I-A beta gene is not. The treatment of macrophages with IFN-gamma led to an increase in the level of RNA for both TNF and I-A beta. The increase in expression of I-A beta and TNF, induced by IFN-gamma, was blocked by naphthalenesulfonamide or phenothiazine (trifluoperazine) but was not affected by the addition of isoquinolinesulfonamide or sphingosine. These results suggest that the induced expression of I-A beta and TNF by IFN-gamma is mediated by a pathway that is protein kinase C independent. This was supported by the finding that calcium ionophores were also able to induce the gene expression of both TNF and I-A beta. We observed that when both IFN-gamma and PMA were added to the macrophages, the level of RNA for TNF increased to a higher level than the level seen when either agent alone was added to the cells. In contrast, the addition of both IFN-gamma and PMA to macrophages had an inhibitory effect on the expression of the I-A beta gene. These results further emphasize the complex nature of gene regulation during the activation of macrophages.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1990

The macrophage and B cell-specific transcription factor PU.1 is related to the ets oncogene.

We have isolated a cDNA clone, PU.1, that codes for a new tissue-specific DNA binding protein. Analysis of the binding site by methylation interference and DNAase 1 protection revealed that the PU.1 protein recognized a purine-rich sequence, 5'-GAGGAA-3' (PU box). The PU.1 protein was shown to be a transcriptional activator that is expressed in macrophages and B cells. cDNA constructions used to generate proteins lacking portions of either the amino- or carboxy-terminal ends of the PU.1 protein placed the DNA binding domain in the highly basic carboxy-terminal domain of the protein. The amino acid sequence in the binding domain of PU.1 has considerable identity with proteins belonging to the ets oncogene family.

Amino Acid Sequence↗
PubMed1989

The expression of I-A correlates with the uptake of interferon-gamma by macrophages.

The current studies were designed to examine some of the requirements for I-A expression when macrophages (M phi) were treated with interferon-gamma (IFN-gamma). In order to define the minimum time required for IFN-gamma to induce surface expression of I-A antigen on bone marrow-derived M phi, cells were incubated with IFN-gamma for varying lengths of time, washed and thereafter incubated for 72 h before assaying I-A surface expression. Using saturating amounts of IFN-gamma (300 IRU/ml), we found that between 0 and 30 min of IFN-gamma treatment there is a direct correlation between the length of treatment and the level of I-A surface expression. When the steady state level of RNA for the I-A beta gene was assayed, a low level of I-A beta RNA was seen in cells treated for 10 min with saturating amounts of IFN-gamma (300 IRU/ml) while a 30-min or 60-min exposure of cells to the same concentration of IFN-gamma resulted in a steady increase in the level of I-A beta RNA. Similar results were found when we measured the levels of RNA for the tumor necrosis factor and C3 complement genes, both of which are induced by IFN-gamma in M phi. M phi treated with low amounts of IFN-gamma (3 IRU/ml) for 30 min do not express cell surface I-A. Cells incubated continuously for 72 h with 3 IRU/ml of IFN-gamma expressed a level of I-A on the surface equivalent to the level of I-A expressed on cells treated for only 30 min with 300 IRU/ml of IFN-gamma. Based on the observed correlation between either the IFN-gamma concentration or the length of time the cells were exposed to IFN-gamma, or the level of I-A expression on M phi, we conclude that the expression of I-A correlates with the uptake of IFN-gamma.

Animals↗
PubMed1989

Interferon-gamma activates multiple pathways to regulate the expression of the genes for major histocompatibility class II I-A beta, tumor necrosis factor and complement component C3 in mouse macrophages.

The purpose of this study was to obtain additional information on the mechanism by which interferon-gamma (IFN-gamma) is able to regulate gene expression in macrophages. The expression of the genes for class II histocompatibility I-A beta, tumor necrosis factor (TNF) and complement component C3 was assayed after treating bone marrow macrophages with IFN-gamma. Each gene displayed a characteristic pattern of regulation. First, the increase in the level of RNA for each gene followed different kinetics. The level of TNF RNA increased within 15 min after IFN-gamma treatment and reached a plateau after 4 h. In contrast, there was a lag of about 4 h before the level of I-A beta RNA began to rise and a plateau was not reached until 48 h after the IFN-gamma treatment began. C3 gene expression followed an intermediate time course between that for TNF and I-A beta. Second, the expression of I-A beta was inhibited when cells were treated with both IFN-gamma and cycloheximide, while the expression of TNF and C3 was not. Interestingly, the sensitivity to cycloheximide only lasted 30 min following the addition of IFN-gamma, after which cycloheximide had no effect on the expression of I-A beta. Third, lipopolysaccharide abolished the IFN-gamma-induced expression of I-A beta, but enhanced the expression of TNF. Based on these observations, we conclude that IFN-gamma must activate multiple pathways to regulate gene expression in macrophages.

Animals↗
PubMed1989

DNA binding of the mouse class II major histocompatibility CCAAT factor depends on two components.

A CCAAT-binding factor that recognizes a CCAAT sequence (Y box) located upstream of the major histocompatibility class II gene I-A beta has been partially purified. This CCAAT-binding factor was found to consist of two components, designated factors A and B, both of which were required for efficient binding to the DNA. Factor A had an apparent molecular size of 34 kilodaltons, and factor B had an apparent molecular size of 42 to 46 kilodaltons.

Animals↗
PubMed1988

Identification of a nuclear factor that binds to a conserved sequence of the I-A beta gene.

Human and murine class II genes of the MHC show a striking homology 50 to 120 bp upstream of the transcription start site. This area is composed of two conserved sequences (a 13-mer and an 8-mer separated by 19 to 20 bp). Recently, these conserved sequences have been identified as cis-acting transcriptional regulatory elements. We have sought nuclear factors that bind specifically to an upstream fragment (-245 to +75 bp) of the murine I-A beta chain gene that contains the conserved sequences by application of a modified gel electrophoresis DNA binding assay. We report here the identification of a nuclear factor whose binding site overlaps the 8-mer conserved sequence. This factor is present in murine B and T lymphocytes, macrophages, mastocytes, fibroblasts, and human B lymphocytes and macrophages. The binding site was defined by using DNase I and dimethylsulfate protection assays. The putative binding sequence is closely related to the sequence, CCAAT, which is often found associated with the promoter of a gene and is recognized by the transcriptional factors CCAAT-binding transcription factor and nuclear factor I. Oligonucleotides that contain the binding site sequences for nuclear factor I and the alpha-globin CCAAT element, however, do not compete for binding of the nuclear factor to the sequence identified here, suggesting that, in spite of the similarity of the binding sequence, the nuclear factor identified in this report may be different. This nuclear protein may be one of the trans-acting factors that mediate transcription of class II genes.

Animals↗
PubMed1988

The interferon gamma receptor.

Interferon-gamma (IFN gamma) can regulate in vitro and in vivo a number of functions on a variety of cell types including some with no known immune functions. Recent work has indicated that IFN gamma exerts its effects upon interaction with a specific receptor on the cell surface. An IFN gamma receptor has been detected in a large variety of cell types of human and murine origins. Species specificity of IFN gamma appears to be a function of ligand-receptor interaction. In the human system, the IFN gamma receptor is a protein of 90 kd molecular weight. IFN gamma is internalized and degraded at a constant rate. The continuous uptake of IFN gamma is due to the presence of an intracellular receptor pool and to a mechanism of receptor recycling. By comparing the relationship between receptor occupancy and biologic response induction in macrophages, two activation mechanisms became apparent. Induction of certain functions such as H2O2 secretion, expression of Fc receptor or IA appeared to require only a single round of receptor occupancy. However, induction of more complex functions such as non-specific tumoricidal activity appeared to require three to four rounds of receptor occupancy. These results thus support the concept that IFN gamma induces different activities by triggering a different pathway at the cell surface and/or inside of the cell.

Animals↗
PubMed1988

Immune-complex inhibition of macrophage activation is not due to an interaction with the binding or processing of IFN-gamma.

Preincubation of macrophages with immune complexes suppresses the interferon-gamma (IFN-gamma) induction of tumouricidal activity and Ia surface expression. The studies reported in this manuscript were designed to test if immune complexes alter the interaction of IFN-gamma with macrophages. The binding of IFN-gamma to its specific cellular receptor, the uptake or the degradation were not affected by preincubation of macrophages with immune complexes. Preincubation of macrophages with high doses of phorbol esters mimics the inhibitory effect of immune complexes. This suggests that a strong activation of protein kinase C suppresses a subsequent activation of macrophages by IFN-gamma.

Animals↗
PubMed1987

Internalization and degradation of receptor-bound interferon-gamma by murine macrophages. Demonstration of receptor recycling.

Although the interferon-gamma (IFN-gamma) receptor on murine and human mononuclear phagocytes has been defined and partially characterized, very little data exists which describes the ultimate fate of receptor-bound ligand. The current studies were specifically designed to define the metabolic processes which act on murine recombinant IFN-gamma following its interaction with murine macrophages at physiologic temperatures. Ligand internalization was demonstrated by comparing binding of [125I]IFN-gamma to macrophages at 4 degrees C and 37 degrees C. When binding was carried out at 4 degrees C, 96% of the cell-associated [125I]IFN-gamma remained accessible at the plasma membrane and could be stripped from the cell by exposure to pronase. In contrast, at 37 degrees C, only 35% of the cell-associated radioactivity was pronase strippable. Macrophages degraded [125I]IFN-gamma into trichloroacetic acid-soluble material at 37 degrees C at a constant rate of 7000 molecules/cell/hr over a 12-hr time period. The amount of IFN-gamma degraded correlated with the amount of IFN-gamma bound to the cell surface. The receptor was neither up- nor down-regulated by ligand or by other agents known to regulate macrophage functional activity such as IFN-alpha, IFN-beta, lipopolysaccharide, or phorbol myristate acetate. The constant uptake of IFN-gamma by macrophages was due to the presence of an intracellular receptor pool (62% of the total receptor number) and to a mechanism of receptor recycling. Evidence for the latter was obtained using lysosomotropic agents which blocked degradation but not binding and internalization of ligand and caused the intracellular accumulation of receptor. By comparing the relationship between receptor occupancy and biologic response induction, two activation mechanisms became apparent. Induction of certain functions, such as H2O2 secretion, appeared to require only a single round of receptor occupancy. However, induction of more complex functions such as nonspecific tumoricidal activity appeared to require three to four rounds of receptor occupancy. These results thus support the concept that IFN-gamma internalization and receptor recycling are essential in the induction of nonspecific tumoricidal activity by macrophages.

Animals↗
PubMed1986

Analysis of deficiencies in IFN-gamma-mediated priming for tumor cytotoxicity in peritoneal macrophages from A/J mice.

The functional and biochemical responses of macrophages derived from the A/J mouse strain to IFN-gamma have been studied. As compared to macrophages obtained from C57BL/6 strain mice, cells from mice of the A/J strain are deficient in their response to IFN-gamma for acquisition of tumoricidal competence. This deficiency was not due to reduced expression of surface receptors for IFN-gamma or to altered affinity of the receptor for its ligand. IFN-gamma recently has been shown to enhance the potential activity of protein kinase C (PKc) and to modulate the efflux of intracellular Ca2+ in macrophages from C57BL/6 mice. Neither of these two biochemical changes were induced in macrophages derived from A/J mice. Functional competence could, however, be pharmacologically induced in both C57BL/6- and A/J-derived macrophages by combined treatment with an ionophore plus phorbol myristic acetate, which increase intracellular Ca2+ and stimulate PKc, respectively. Although the exact nature of the deficit in A/J strain mice has not been defined, the present findings indicate that it lies between the expression of receptor and the modulation of PKc activity and Ca2+ levels. Furthermore, the data provide support for the notion that these molecular changes are important components of the stimulus-response coupling process in IFN-gamma-mediated activation of macrophages.

Animals↗
PubMed1986

Role of protein kinase C and intracellular calcium mobilization in the induction of macrophage tumoricidal activity by interferon-gamma.

These studies were designed to test the hypothesis that changes in intracellular Ca2+ levels and activation of the calcium ion- and phospholipid-dependent protein kinase C were required for the induction of macrophage tumoricidal activity by interferon-gamma (IFN-gamma). Phenothiazines and R24571, known antagonists of calcium-binding proteins and therefore nonspecific inhibitors of protein kinase C, blocked in a dose-dependent manner the induction of macrophage cytocidal activity by either natural or recombinant IFN-gamma. Macrophages depleted of intracellular Ca2+ by chelation with Quin 2, were also unresponsive to IFN-gamma. These treatments effected neither the binding of IFN-gamma to its cell surface receptor nor the normal intracellular processing of IFN-gamma. Activators of protein kinase C (such as phorbol esters) and Ca2+ ionophores when added alone did not effect the activation state of the macrophage population. However, macrophages exposed to both drugs in combination were elevated into the primed activation state such that in the presence of a second signal (lipopolysaccharide or heat killed Listeria monocytogenes), the cells were triggered to express full levels of tumoricidal activity. The capacity of phorbol esters to induce cellular activation correlated with their ability to bind and to activate protein kinase C. No synergistic effect was observed between IFN-gamma and protein kinase C activators and/or Ca2+ ionophores, indicating that the drugs could only prime and could not trigger macrophages for tumor cell killing. These results thus support the concept that protein kinase C activation and mobilization of intracellular Ca2+ are essential steps in the pathway of IFN-gamma-dependent induction of non-specific tumoricidal activity in macrophages.

Aminoquinolines↗
PubMed1986

[Effect of acute anemia or polycythemia on blood flow and iron transport in bone marrow].

Iron has been shown to be the limiting factor for erythropoiesis. The anemia and polycythemia effect on iron supplied to the bone marrow has been studied in a group of rabbits, by modifying the hematocrit without altering of the blood volume. The cardiac output and the percentage of blood flow to the skeleton was measured using 57Co and 113Sn radiolabelled microspheres, before and after the exchange of blood by plasma or red blood cells concentrates. In addition, ferrokinetic measurements were performed with 55Fe and 59Fe. The production of an acute anemia induced an increase in the cardiac output from 156 +/- 35 to 239 +/- 89 ml/min/kg and a decrease in the percentage of the total blood flow to the skeleton from 7.58 +/- 2.51 to 4.63 +/- 1.8. The production of an acute polycythemia induced a decrease in the cardiac output (97 +/- 28 ml/min/kg) and an increase in the percentage of the total blood flow to the bone marrow (11.69 +/- 4.03). However, in both cases, the absolute amount of blood flow and iron flow to the bone marrow were similar to the controls. These studies demonstrate that anemia or polycythemia per se do not determine the iron supply to the bone marrow.

Acute Disease↗