bluuebunny/crossref_metadata_2025 huggingface dataset:
Isolation of monocytic serine esterase and evaluation of its proteolytic activity
Authors: Salmassi Ali , Kreipe Hans , Radzun Heinz J , Lilischkis Richard , Charchinajad-Amoey Madjid , Zschunke Frank , Buck Friedrich , Lottspeich Friedrich , Parwaresch M Reza
Published: January 2018

Monocytes are characterized by high activity of α-naphthyl acetate esterase (ANAE), distinguishing them from all other blood cells. The physiological function of this monocyte marker enzyme has not yet been elucidated. In this study ANAE's potential proteolytic activity was anlayzed because serine esterases/proteases can function as effector molecules in cell-mediated cytotoxicity and because monocytes-macrophages are known to exert cytotoxic effects on tumor cells. This enzyme was purified from the monocytic cell line U-937 by preparative isoelectric focusing and a three-step high-performance liquid chromatography that conserved its catalytic activity. It has a molecular mass of 60 kd, and partial amino acid sequence revealed that the enzyme is not identical to known serine esterases/proteases. The purified enzyme failed to digest a couple of peptides, indicating lack of protease activity. In addition, the esterolytic activity of ANAE was not inhibited by protease inhibitors. The isolation and purification of ANAE enable further studies concerning its function in monocytes-macrophages and its relation to monocytic cytotoxicity.
Effector function of hepatocytes and Kupffer cells in the resolution of systemic bacterial infections
Authors: Gregory Stephen H , Barczynski Lisa K , Wing Edward J
Published: January 2018

It has been suggested that mononuclear phagocytes serve as the principal site of replication for a number of intracellular pathogens including Listeria monocytogenes. To determine the role of the tissue macrophages (Kupffer cells) in the proliferation of Listeria taken up in the liver, the hepatic cell populations were purified and the associated bacteria were quantified at periodic intervals postinfection. Here we report that the bulk of Listeria injected intravenously into nonimmune mice replicated within hepatocytes rather than Kupffer cells. Whereas a 200-fold increase in the number of hepatocyte-associated Listeria occurred during the first 3 days following infection, a relatively small (less than 2-fold) increase in number of Kupffer cell-associated Listeria was observed. The Listeria injected intravenously into immune animals, on the other hand, were eliminated rapidly from the hepatocyte as well as the Kupffer cell population. The latter findings suggest that uptake and elimination of pathogenic organisms by “nonprofessional phagocytes” in the liver (i.e., hepatocytes) may be an important effector mechanism in host defenses.
Cationic defensins arise from charge-neutralized propeptides: a mechanism for avoiding leukocyte autocytotoxicity?
Authors: Michaelson David , Rayner John , Couto Marcelo , Ganz Tomas
Published: January 2018

Defensins, small cationic polypeptides with antimicrobial and cytotoxic properties, are among the principal constituents of cytoplasmic granules of mammalian neutrophils and certain macrophages. To identify conserved structural features of defensin precursors that may be important for their targeting to cytoplasmic granules or for prevention of autocytotoxicity, we isolated and sequenced three neutrophil-specific rabbit defensin cDNAs that code for preproprotein precursors to the mature defensins NP-3a, NP-4, and NP-5. The preprodefensins NP-3a, NP-4, and NP-5, like the previously characterized preprodefensins, lack consensus sequences for N-linked glycosylation, suggesting that defensins are targeted to lysosome-like granules by a mechanism not dependent on the mannose-6-phosphate receptor. Analysis of all seven known myeloid prodefensins revealed a structure wherein an anionic propiece neutralizes the cationicity of the mature peptide. Because defensins apparently require cationic epitopes for cell membrane permeabilization and cytotoxicity, charge neutralization of mature peptides by their anionic propieces may prevent autocytotoxicity during defensin synthesis and processing.
Regulatory role of cytokines in IgE-mediated allergy
Authors: Tan Henkie P , Lebeck Lauralynn K , Nehlsen-Cannarella Sandra L
Published: January 2018

The discovery of immunoglobulin E (IgE) is considered the most important contribution, to date, in the field of clinical allergy. Studies in rodents and humans have suggested that IgE production could be regulated by antigen-specific helper and suppressor T cells and by isotype-specific factors having affinity for IgE. In recent years, the synthesis of IgE has been shown to be regulated, in part, by a cytokine network. This review summarizes the cytokines that up-regulate (interleukins-4, 5, and 6) and down-regulate (interferon-γ and interleukin-2) the production of IgE. Emphasis is placed on IL-4 and IFN-γ, two lymphokines known to play a major, but reciprocal, role in IgE synthesis. Increased insight into the various mechanisms of IgE control by cytokines and their receptors will eventually lead to improved treatment strategies in the clinical management of IgE-mediated allergy.
Tissue-specific expression of murine IP-10 mRNA following systemic treatment with interferon γ
Authors: Narumi Shosaku , Wyner Lawrence M , Stoler Mark H , Tannenbaum Charles S , Hamilton Thomas A
Published: January 2018

We have examined the tissue distribution of 10-kd inflammatory protein (IP-10) mRNA expression in C57Bl/6 mice injected intravenously (i.v.) with various inflammatory stimuli. IP-10 mRNA was strongly induced by interferon-γ (IFN-γ) in liver and kidney but only poorly in skin, heart, and lung. IFN-γ had nearly equivalent access to these tissues as indicated by the distribution of radiolabeled recombinant IFN-γ 1 h after injection. The time course of IP-10 mRNA appearance was rapid and transient in both liver and kidney; maximal expression in the liver (2 h) preceded that in the kidney (3 h) and declined rapidly thereafter in both tissues. Expression of IP-10 mRNA in the liver and kidney was highly sensitive to IFN-γ treatment; nearly maximal stimulation occurred with injection of 500 U of IFN-γ per mouse. Comparable stimulation of IP-10 mRNA expression in splenic macrophages required 10,000 U of IFN-γ administered i.v., indicating that liver and kidney responses are 10- to 20-fold more sensitive. IP-10 mRNA expression in both tissues was not restricted to stimulation by IFN-γ but was also seen with injection of lipopolysaccharide (LPS) (25 μg/mouse) or IFN-β (100,000 U/mouse). Two other members of the IP-10 gene family, KC (gro) and JE (MCP-1), were expressed at lower levels under similar treatment conditions. Analysis of IP-10 mRNA distribution in the liver and kidney by in situ hybridization indicated that expression in both tissues was most prominent in the reticuloendothelial cell system, particularly in the endothelial lining of the microvascular circulation. Although the function of the IP-10 gene product has not been defined, these results suggest that it may play an important role in the response of both the liver and kidney to systemic inflammation.
Impaired stimulus-response coupling in association with increased growth rate of HL60 cells
Authors: Hadjokas Nicholas , Bayer Clifford , Nielson Christopher P
Published: January 2018

Impairments in the respiratory burst and stimulus-response coupling were studied with respect to the increased rate of cell replication that occurred in HL60 cells during repetitive passages in cell culture. During a 45-week period of culture, HL60 cells developed a progressive increase in rate of replication. Concomitantly, undifferentiated cells developed an impairment in ATP-induced calcium mobilization. The percentage of cells that could be differentiated with dimethyl sulfoxide progressively diminished. Differentiated cells developed an impairment in both the respiratory burst and secretion of β-glucuronidase. In addition, regulation of the respiratory burst by cAMP agonists including isoproterenol, adenosine, and prostaglandin E2 was reduced in rapidly proliferating cells. Thus, multiple changes in stimulus-response coupling occur during cell culture in association with an increase in rate of cell replication. It may be important to recognize progressive impairments in cell function in studies using repetitive samples of HL60 cells from a continuously maintained cell population. The observed impairments in stimulus-response coupling may be relevant to unregulated cell growth in neoplastic disease.
Effect of lipopolysaccharide on transport and metabolism of arginine in mouse peritoneal macrophages
Authors: Sato Hideyo , Fujiwara Mari , Bannai Shiro
Published: January 2018

The transport activity of arginine in mouse peritoneal macrophages was strongly induced when they were cultured with 1 ng/ml bacterial lipopolysaccharide (LPS) for 12 h. Arginine in the medium decreased whereas ornithine in the medium increased during the culture. This time-dependent change of arginine to ornithine was accelerated by LPS. However, the activity of arginase in the macrophages did not change during the culture with or without LPS and release of arginase from the cells to the medium was not detected. It is suggested that the transport of arginine and ornithine was a rate-limiting step in arginine-to-ornithine conversion in the macrophage culture medium. A possible role of the induction of arginine transport activity in the macrophage cytocidal activity due to arginine depletion and nitric oxide production is discussed.
The myeloid leukemia cell line HL-60 produces a factor that induces chloride secretion in cultured epithelial cells
Authors: Colgan Sean P , Matthews Jeffrey B , Parkos Charles A , Delp Charlene , Awtrey Christopher S , Madara James L
Published: January 2018

When activated, polymorphonuclear leukocytes (PMNs) produce a small soluble factor, termed neutrophil-derived secretagogue (NDS), that elicits electrogenic Cl- secretion — the transport event responsible for hydration of mucosal surfaces. Work toward purification of this factor has been hampered by variability in activity of PMN-derived NDS. Using a human-derived intestinal epithelial cell line (T84) that serves as a model for studies of the regulation of electrogenic Cl- secretion, we find that the human promyelocytic leukemia cell line HL-60 secretes a factor with NDS-like activity. Buffer conditioned by HL-60 cells (107 cells/ml for 1 h), when applied to T84 monolayers grown on permeable supports and analyzed by routine electrophysiological techniques, elicited a short-circuit current (Isc) of 11.7 ± 1.02 μA/cm2. This short-circuit current was sensitive to bumetanide, an inhibitor of the basolateral Na-K-2CI cotransporter, and was dependent on the presence of chloride in the assay buffers. Such data indicate that buffer conditioned by HL-60 cells stimulates electrogenic Cl- secretion. Like NDS, the active factor in HL-60–conditioned buffer has a nominal molecular weight of less than 500 and was increased by activation of cells with phorbol ester. 125I and 86Rb efflux assays confirmed that the secretagogue released by stimulated HL-60 cells, similar to PMN-derived NDS, preferentially stimulates opening of Cl- rather than K+ channels in T84 cells. Lastly, release of NDS-like bioactivity increases when HL-60 cells are differentiated toward granulocytes compared to cells differentiated toward monocytes.
Lytic effects of Actinobacillus actinomycetemcomitans leukotoxin on human neutrophil cytoplasts
Authors: Iwase Masayasu , Korchak Helen M , Lally Edward T , Berthold Peter , Taichman Norton S
Published: January 2018

Actinobacillus actinomycetemcomitans leukotoxin lysed human neutrophil cytoplasts. The reaction was associated with a rapid influx of extracellular calcium, a collapse in membrane potential, release of lactate dehydrogenase, and overt disintegration of the plasma membrane. These functional and structural alterations in the plasmalemma of neutrophil cytoplasts reinforce the hypothesis that A. actinomycetemcomitans leukotoxin acts as a pore-forming, membranolytic agent and indicate that neutrophil cytoplasts are useful tools in studying the biology of membrane-active toxins.
Macrophage function in response to PGE2, L-arginine deprivation, and activation by colony-stimulating factors is dependent on hematopoietic stimulus
Authors: Rutherford Mark S , Schook Lawrence B
Published: January 2018

Different macrophage preparations were compared for functional capacity in conditions of high prostaglandin E2 (PGE2) or low l-arginine concentrations. Macrophages derived in vitro from bone marrow progenitor cells (bone marrow-derived macrophages, BMDMs) using colony-stimulating factor 1 (CSF-1) as the myelopoietic stimulus displayed a greater sensitivity to PGE2-induced suppression of tumor necrosis factor a (TNF-α) secretion than did macrophages derived using granulocyte-macrophage colony-stimulating factor (GM-GSF). Neither BMDM population was inhibited by PGE2 for the direct cytolysis of L929 cells (TNF-α sensitive), and only GM-CSF–derived macrophages showed decreased killing of TNF-α–resistant K562 targets. Exogenous cAMP inhibited TNF-α secretion, but not nitrite secretion, by both BMDM populations. GM-GSF–derived macrophages accumulated less cAMP following PGE2 treatment than did CSF-1–derived macrophages. Removing l-arginine from the medium did not inhibit cytotoxicity or PGE2 secretion, but the listeriacidal activity specific to interferon-γ plus lipopolysaccharide (LPS)-activated GM-CSF–derived macrophages was blocked by removal of l-arginine. Treatment with CSF-1 or GM-CSF alone did not activate the macrophages, but GM-CSF efficiently primed both BMDM populations for augmented TNF-α secretion in response to secondary stimulation using LPS. However, GM-CSF augmented the LPS-induced production of nitrite and PGE2 by CSF-1–derived macrophages only. These results demonstrate the potential for differential macrophage function within inflammatory sites based on the hematopoietic stimulus under which the macrophage is derived and the specific conditions present in the lesion.
Liposomes containing muramyl tripeptide phosphatidylethanolamine (MTP-PE) are excellent adjuvants for induction of an immune response to protein and tumor antigens
Authors: Ullrich Stephen E , Fidler Isaiah J
Published: January 2018

Liposomes containing the synthetic lipophilic analog of muramyl dipeptide, muramyl tripeptide phosphatidylethanolamine (MTP-PE), were used as adjuvants for the induction of humoral and cellular immune responses following immunization with protein or tumor antigens. Cellular immune reactions, including delayed- type hypersensitivity and lymphoproliferation in vitro, were observed following immunization of mice with a mixture of antigen and liposome-MTP-PE. Immunization with murine melanoma K1735 cells, admixed with liposomal MTP-PE, induced a protective immune response as demonstrated by the rejection of transplanted tumor cells. Antibody production was also induced following immunization with protein antigens admixed with liposome-MTP-PE. The efficacy of adjuvant activity following immunization with antigens admixed with liposome-MTP-PE was equal to or better than that of complete Freund's adjuvant (CFA). Moreover, liposome- MTP-PE did not have the toxic side effects associated with CFA. These data suggest that phospholipid liposomes containing MTP-PE are superior adjuvants and should receive consideration for vaccine therapy.
Activation of human monocyte chemiluminescence response by acylpoly(1,3)galactosides derived from Kiebsiella pneumoniae
Authors: Kouassi Edouard , Hmama Zakaria , Una Gérard , Vial Jean , Faure-Barba Florence , Normier Gérard , Binz Hans , Revillard Jean-Pierre
Published: January 2018

The stimulating activity of several preparations isolated from a membrane proteoglycan of a nonen- capsulated smooth strain of Klebsiella pneumoniae (Kp- MPG) on the oxidative burst of human blood monocytes was assessed by luminol-enhanced chemiluminescence (CL). Five Kp derivatives were studied: a 34-kd acyl- poly(l,3)galactoside (APG), obtained by drastic alkaline hydrolysis and purified by chromatography; an APG preparation subjected to acid hydrolysis that removed the core part and all fatty acids, leaving intact the galactose chain of APG (GC-APG); an APG preparation subjected to mild oxidation (ox APG); a preparation obtained by mild alkaline hydrolysis of Kp-MPG, containing additional ester-linked C14 and C16 fatty acids bound to the APG molecule (EFA-APG); and a polymer of the latter compound, APG pol. EFA-APG directly stimulated monocyte CL, whereas Kp-MPG, APG pol, and the whole bacterial cells had little or no activity. APG itself and ox APG induced a weaker response than EFA-APG. Polymyxin B sulfate completely inhibited the CL response to bacterial lipopolysaccharide (LPS) but not to EFA-APG. The stimulating action of EFA-APG on blood monocytes was dependent on the extracellular levels of both calcium and magnesium. Preincubation of monocytes with monoclonal antibody anti-Mac-1 directed against CDllb, the αchain of complement receptor type 3 (CR3; CDllb/CD18), strongly inhibited CL activation by EFA-APG and to a lesser extent CL activation by un- opsonized zymosan and rough LPS. Altogether, these findings provide indirect evidence for the contribution of the CDllb/CD18 integrin in the functional interaction of EFA-APG with monocyte membranes. They demonstrate the role of fatty acids in the triggering of monocyte oxidative burst, while the polysaccharide chain itself does not contribute to induction of the CL response in this model. In keeping with the effects of EFA-APG and APG, we show that the monocyte CL response was triggered by bacterial LPS from the rough strain of Salmonella minnesota Re 595 and its lipid A, but not by LPS from smooth strains, again suggesting a critical role for the lipid moiety.
Role of CD4+ T lymphocytes and interleukin-5 in antigen-induced eosinophil recruitment into the site of cutaneous late-phase reaction in mice
Authors: Iwamoto Itsuo , Tomoe Sanae , Tomioka Hisao , Takatsu Kiyoshi , Yoshida Sho
Published: January 2018

Previous studies suggested that the eosinophil recruitment into the site of cutaneous late-phase reaction (LPR) was dependent on IgE antibody and mast cells. In this study, we determined the role of CD4+ T cells and CDS+, T cells in causing antigen-induced eosinophil recruitment of LPR in mouse skin. Eosinophil infiltration into the subcutaneous tissue of ovalbumin (OVA)- sensitized BALB/c mice was biphasic, reaching the first peak at 6 h after the subcutaneous challenge with OVA and the second peak at 24 to 48 h. The in vivo depletion of CD4+ T cells by pretreatment with anti-L3T4 monoclonal antibody (mAb) significantly decreased the second peak (at 24 h and 48 h), but not the first peak (at 6 h), of OVA-induced eosinophil infiltration into the skin of OVA-sensitized mice. However, the depletion of CD8+ T cells by pretreatment with anti-Lyt-2 mAb had no significant effect on either the first peak or second peak of OVA-induced cutaneous eosinophilia. Pretreatment with anti-murine interleukin-5 (IL-5) mAb also decreased the second peak, but not the first peak, of OVA- induced cutaneous eosinophilia. In contrast to the inhibitory effects of depletion of CD4+ T cells and of anti-IL-5 mAb on the second peak of antigen-induced cutaneous eosinophilia, disodium cromoglycate and a selective antagonist for platelet activating factor (PAF) CV-6209 decreased the first peak of OVA-induced cutaneous eosinophilia in the mouse. These results indicate that CD4+ T cells, but not CD8+ T cells, cause the second peak of antigen-induced eosinophil recruitment of cutaneous LPR and that IL-5 mediates this eosinophil recruitment. In contrast, the first peak of antigen-induced eosinophil recruitment of cutaneous LPR is mediated by mast cells and PAF.
Eosinophil granule proteins in peripheral blood granulocytes
Authors: Abu-Ghazaleh Randa I , Dunnette Sandra L , Loegering David A , Checked James L , Kita Hirohito , Thomas Larry L , Gleich Gerald J
Published: January 2018

Eosinophils contain four principal cationic proteins, major basic protein (MBP), eosinophil-derived neurotoxin (EDN), eosinophil cationic protein (ECP), and eosinophil peroxidase (EPO). To determine the quantities of these proteins in granulocytes and whether they are specific to eosinophils, their concentrations in ly- sates of human granulocytes were measured using specific radioimmunoassays. The effect of different methods for eosinophil lysis on the recovery of the proteins was also studied. Maximal recovery occurred at pH 2 for MBP and pH 5.6 for the other granule proteins. The proteins cosedimented with eosinophils and their concentrations (X ± SEM) in ng/106 eosinophils (and in nM/106 eosinophils) were: MBP, 8,982 ± 611 (641.6); EDN, 3,283 ± 116 (178.4); ECP, 5,269 ± 283 (250.9); and EPO, 12,174 ± 859 (171.5). Basophils from a normal person contained (in ng/106 cells) MBP, 2,374; EDN, 214; ECP, 77; and EPO, 17. Highly purified neutrophils contained (in ng/106 cells) MBP, 3 ± 0.5; EDN, 72 ± 9; and ECP, 50 ± 12. Therefore we conclude that these proteins are mainly expressed in eosinophils, but that certain ones are present in basophils and neutrophils.
A new, one-step assay on whole cell suspensions for peroxidase secretion by human neutrophils and eosinophils
Authors: Menegazzi Renzo , Zabucchi Giuliano , Knowles Alessandra , Cramer Rita , Patriarca Pierluigi
Published: January 2018

The degranulation of neutrophils and eosinophils is frequently monitored by assaying myeloperoxidase (MPO) and eosinophil peroxidase (EPO) activity in the cell-free supernatant of degranulating cells, after removal of the cells by centrifugation. This procedure leads to underestimation of the extent of degranulation, since both peroxidases tend to stick to cell surfaces, to test tube walls, and to particulate stimuli used to elicit degranulation, because of their highly cationic nature. In this paper we describe a method for assaying MPO and EPO secretion in whole cell suspensions that avoids separation of the cells from the incubation medium. The least toxic and thus safest among the sensitive peroxidase substrates, 3,3′,5,5′-tetramethylbenzidine (TMB), was employed for peroxidase assay. The method we describe here is applied to the detection of peroxidase release by neutrophil and eosinophil cell suspensions incubated in either polypropylene test tubes or flat- bottomed microtiter plate wells. Because of the omission of the centrifugation step, the TMB method offers two major advantages over the currently used techniques: (1) higher estimates of degranulation, which permits the use of a smaller number of cells (in the microassay version, 150,000 neutrophils and 50,000 eosinophils) and smaller amounts of the secretagogues, and (2) rapidity, since the degranulation assay can be performed immediately on completion of the cell incubation with the secretagogue.
Tumor necrosis factor-alpha decreases neutrophil chemotaxis to N-formyl-1-methionyl-1-leucyl-1-phenylalanine: analysis of single cell movement
Authors: Vollmer Kelly L , Alberts Jeffrey S , Carper Holliday T , Mandell Gerald L
Published: January 2018

Tumor necrosis factor-alpha (TNF-α), a cytokine produced by mononuclear cells in response to endotoxin, inhibits neutrophil chemotaxis. We analyzed the effects of TNF-α on the orientation and movement of individual neutrophils in a chemoattractant gradient. Neutrophils, treated or untreated with TNF-α, were observed migrating in a gradient of the chemotactic peptide N-formyl-l-methionyl-l-leucyl-l-phenylalanine (fMLP) on a specially constructed chamber (Zigmond bridge). The movement of these cells was videotaped, digitized, and then tracked using a newly designed computer algorithm. The data obtained from this algorithm were then utilized to calculate distance traveled, speed and ability to polarize and migrate in a directed manner for each individual cell. TNF-a-treated cells behaved like cells not exposed to fMLP in that they failed to orient in a chemotactic gradient and moved in a manner similar to randomly migrating cells. This study provides unique observations of the effect of TNF-α on multiple parameters of PMN migration.
Expression of the DAF (CD55) and CD59 antigens during normal hematopoietic cell differentiation
Authors: Terstappen Leon WMM , Nguyen Mai , Lazarus Hillard M , Medof M Edward
Published: January 2018

Expression of decay-accelerating factor (DAF or CD55) and of CD59 during hematopoietic cell development in normal bone marrow and on peripheral blood leukocytes were characterized by three-color immunofluorescence experiments. With this technique cell subsets were identified by forward light scatter, orthogonal light scatter, and two cell-surface antigens. For each cell lineage, specific combinations of two monoclonal antibodies labeled with different fluorochromes were used. DAF or GD59 were then quantitated on the defined cell subsets from the fluorescence signal of the respective antibody conjugated with a third fluorochrome. Early uncommitted hematopoietic progenitor cells (CD34+, CD38+) all expressed both proteins homogeneously. Initial commitment to the eryth- roid (CD71+, CD45dim), myeloid (CD33+), or B lymphocyte (CD10+) lineages was not associated with changes in DAF or CD59 levels. With erythroid development, i.e., after loss of CD45 and decrease of CD71, expression of both proteins decreased. With myeloid maturation, expression of GD59 remained constant and expression of DAF varied. During neutrophil maturation, DAF decreased initially and then reemeiged on maturing neutrophils concurrently with the appearance of GD16 (FcγRIII), whereas during monocyte maturation, DAF increased concurrently with up- regulation of CD14. With B cell development, expression of DAF increased concurrently with down-regulation of GD10 and up-regulation of CD20, whereas expression of CD59 diminished slightly late in B cell maturation. Analysis of peripheral blood elements showed that monocytes, neutrophils, and B lymphocytes expressed both proteins homogeneously, but that in contrast to other cell subsets, which all expressed CD59, only a subset of (CD3+) T lymphocytes and (GDI 6+) Natural killer cells expressed DAF. The absence of DAF was not related to CD4 or CD8 expression or to the presence of activation markers (CD25+, CD38+), memory cell markers (CD58+, CD45RO+), or virgin T cell markers (CD45RA+), but was correlated with expression of CDllb (CR3) and CDllc (gpl50/95). Although CD21” (CR2) and CD35* (CR1) cells all expressed DAF, CDlla (LFA-1) levels correlated inversely with those of DAF. Although the presence of CD55 and CD59 on early progenitor cells and throughout hematopoietic cell development is consistent with the requirements for both proteins in protection of host cells from complement-mediated injury, the physiological relevance of the unique patterns of variation for each cell lineage is unclear. Nevertheless, the availability of a detailed DAF and CD59 expression map in normal marrow will facilitate analyses of alterations during hematopoietic development that may occur in hematological disorders including paroxysmal nocturnal hemoglobinuria (PNH).
Liposome-encapsulated hemoglobin inhibits tumor necrosis factor release from rabbit alveolar macrophages by a posttranscriptional mechanism
Authors: Langdale Lorrie A , Maier Ronald V , Wilson Lynne , Pohlman Timothy H , Williams John G , Rice Charles L
Published: January 2018

Macrophages contribute to the systemic inflammatory response that characterizes the sepsis syndrome through the production of inflammatory cytokines such as tumor necrosis factor (TNF). Liposome- encapsulated hemoglobin (LEH), a potential red cell substitute, is cleared by fixed tissue macrophages. In these studies, in vitro incubation of alveolar macrophages with stored LEH was shown to inhibit the expression of TNF induced by endotoxin (lipopolysaccharide, LPS) stimulation. This effect was dependent on LEH dose but independent of the period of exposure to the LEH. Despite inhibition of TNF expression, Northern blot analysis of total cellular RNA from LPS-stimulated macrophages revealed accumulations of TNF-specific transcripts in cells treated with or without LEH. Thus the mechanism of LEH inhibition of TNF expression appears to involve a posttranscriptional event. Although these results suggest a potential advantage of resuscitation with LEH when sepsis complicates hemorrhagic shock, immunomodulation in vivo remains to be defined.
Role of PAF in the allergic pleurisy caused by ovalbumin in actively sensitized rats
Authors: Martins Marco A , Neto Hugo C Castro Faria , Bozza Patricia T , Silva Patricia MR e , Lima Marcia CR , Cordeiro Renato SB , Vargaftig B Boris
Published: January 2018

Selective platelet-activating factor (PAF) antagonists and autodesensitization to this lipid were used to investigate the role of PAF in antigen-induced pleurisy in the rat. Pleural inflammation was triggered by the intrathoracic (i.t.) injection of ovalbumin (12 μg/cavity) into animals actively sensitized 14 days before. Successive daily i.t. injections of PAF (μg/cavity) led to selective autodesensitization, which was apparent after the third injection and maximal after the fifth. The PAF antagonists BN 52021 and WEB 2086 inhibited the late pleural eosinophil accumulation caused by antigen but, as also noted with WEB 2170, failed to modify the early antigen-induced plasma exudation and leukocyte infiltration. In contrast to the antagonists, desensitization to PAF was clearly effective against these early alterations. To further investigate this discrepancy, the antigenic challenge was performed 24 h after a single prestimulation with PAF, when sensitivity to the lipid was still intact. Under this condition, plasma exudation and cellular influx triggered by the antigen were also abrogated, indicating that this protective effect was accounted for by a mechanism other than refractoriness to PAF. Because 24 h after PAF injection only eosinophil counts remained elevated, an alternative eosinophilotactic substance was used to further study the mechanism of PAF versus antigen-induced pleural inflammation. Prior treatment with the peptide Ala-Gly-Ser-Glu (ECF-A, 20 fig/cavity) also inhibited the allergic pleurisy, whereas the noneosinophilotactic substances histamine (200 μg/cavity) and serotonin (100 μg/cavity) were inactive. Furthermore, drugs that share the ability to impair PAF-induced eo- sinophilia, including azelastine and cetirizine, prevented the inhibitory effect of PAF on the antigen-induced pleurisy. These findings suggest that PAF may account for the late eosinophilia, but not for the acute phase of the rat allergic pleurisy, which is clearly attenuated by PAF or ECF-A pretreatment. J. Leukoc. Biol. 53: 104–111; 1993.
Vitamin D3 compounds regulate human immunodeficiency virus type 1 replication in U937 monoblastoid cells and in monocyte-derived macrophages
Authors: Pauza C David , Kornbluth Richard , Emau Peter , Richman Douglas D , Deftos Leonard J
Published: January 2018

We studied the effects of vitamin D3 compounds on the replication of human immunodeficiency virus type 1 (HIV-1) in the monoblastoid cell line U937 and in primary monocyte-derived macrophage cultures to understand how modulators of monocyte/macrophage effector function might affect the pathogenesis of HIV-1 infection. U937 cell cultures exposed to 1,α25-dihydro- xyvitamin D3 prior to HIV-1 infection showed enhanced virus replication that was apparently due to increased cellular resistance to viral cytopathic effects; a marked inhibition of virus replication was noted in cells exposed to 1,α25-dihydroxyvitamin D3 subsequent to infection. Exposure of blood-derived monocyte/macrophages to vitamin D3 compounds prior to infection also affected virus growth; in most cases, substantial inhibition of HIV- 1 replication was noted in vitamin D3-treated macrophage cultures. Our results demonstrate that vitamin D3 compounds with recognized abilities to induce cellular differentiation can modulate HIV-1 infection of human macrophages. J. Leukoc. Biol. 53: 157–164; 1993.
Oxidative priming of neutrophils by interferon-γ
Authors: Tennenberg Steven D , Fey Debra E , Lieser Mark J
Published: January 2018

Priming of neutrophil (PMN) oxidative responses is an integral component of host defense and inflammation and may contribute to cell-mediated tissue injury. The characteristics and mechanisms of interferon- γ (EFN-γ)-induced oxidative priming of PMNs were explored in vitro. Following pretreatment of human PMNs with recombinant IFN-γ, superoxide anion release was selectively primed toward the receptor-initiated stimulants f-Met-Leu-Phe (fMLP) and C5a but not toward the transduction-mediated stimulants phorbol myristate acetate and A23187, a calcium ionophore. IFN-γ also induced priming toward the stimulant NaF, a direct activator of guanine nucleotide regulatory proteins. Priming was not associated with changes in fMLP surface receptor expression or degranulation. Priming was dependent on de novo mRNA and protein synthesis. The immuno- regulatory lymphokine IFN-γ primes PMN oxidative responses, apparently via production of proteins that are involved in the early postreceptor transductional pathways of oxidative metabolism. J. Leukoc. Biol. 53: 301–308; 1993.
Cytokine priming of the respiratory burst in human eosinophils is Ca2+ independent and accompanied by induction of tyrosine kinase activity
Authors: van der Bruggen Tjomme , Kok Paul TM , Raaijmakers Jan AM , Verhoeven Arthur J , Kessels Roger GC , Lammers Jan WJ , Koenderman Leo
Published: January 2018

We report that pretreatment of human eosinophils with GM-CSF, IL-3, or IL-5 enhanced the respiratory burst induced by opsonized particles. In order to gain more insight into the intracellular mechanism(s) involved in cytokine priming, the role of [Ca2+], and tyrosine kinases was studied. Optimal priming concentrations of GM-CSF, IL-3, and IL-5 did not induce a rise in [Ca2+]i, and Ca2+-depleted eosinophils ([Ca2+]i < 20 nM) were still primed after preincubation with these cytokines. GM-CSF, IL-3, and IL-5 induced phosphorylation of two proteins (102 and 122 kd) on tyrosine residues, as deduced from Western blot analysis with an antiphospho- tyrosine monoclonal antibody (4G10). This cytokine- stimulated tyrosine phosphorylation was not inhibited under Ca2+-depleted conditions. In conclusion, this study demonstrates that GM-CSF, IL-3, and IL-5 priming of the opsonized particle-induced respiratory burst in human eosinophils is completely Ca2+ independent. Moreover the tyrosine phosphorylation of a 102-kd and a 122-kd protein is Ca2+ independent, suggesting that this event might be involved in cytokine priming. J. Leukoc. Biol. 53: 347–353; 1993.
The mouse macrophage activation-associated marker protein, p71/73, is an inducible prostaglandin endoperoxide synthase (cyclooxygenase)
Authors: Phillips Teresa A , Kuiubu Dean A , MacKay Robert J , Herschman Harvey R , Russell Stephen W , Pace Judith L
Published: January 2018

The inducible protein p71/73 marks the response of mouse macrophages to one of several stimuli (e.g., bacterial lipopolysaccharide or poly I:C) that trigger the expression of cytolytic activity when these cells have previously been primed for tumor cell killing by interferon-γ (IFN-γ). The results reported here identify this marker protein as the inducible prostaglandin endoperoxide synthase (PES), TIS10/PES-2. Identification was based on four findings: (1) p71/73, like the TIS10/ PES-2 protein, was associated with cellular membranes; (2) the sequence of amino acids in the NH2 terminus of both p71 and p73 was 96% identical to the predicted NH2-terminal sequence of the TIS10/PES-2 protein; (3) a polyclonal antiserum raised against the COOH-terminal region of the TIS10/PES-2 gene product recognized p71/73 in immunoblots; and (4) dexamethasone, which blocks induction of TIS10/PES-2 expression, inhibited the induction of both p71/73 synthesis and tumoricidal activity in macrophage. Several regulatory roles for this protein in the activation process are possible. J. Leukoc. Biol. 53: 411–419; 1993.
Mast cell granules inhibit macrophage-mediated lysis of mastocytoma cells (P815) and nitric oxide production
Authors: Dileepan Kottarappat N , Lorsbach Robert B , Stechschulte Daniel J
Published: January 2018

The effects of mast cell granules (MCGs) on macrophage-mediated lysis of P815 mastocytoma cells and nitric oxide (NO) production were studied. Murine peritoneal macrophages exhibited tumor cell killing and NO production only when activated with lipopolysaccha- ride (LPS) or interferon-γ (IFN-γ). Coincubation of macrophages with MCGs during LPS activation dose-depen- dently inhibited macrophage-mediated tumor cell lysis. The MCG effect was not due to inactivation or removal of LPS by MCG. The inhibitory effect was also not due to histamine or serotonin present in the MCGs. The granules were not toxic to macrophages or P815 mastocytoma cells. The effect of MCGs on macrophage-mediated tumor cell killing was evident whether MCGs were added before or after a 4-h exposure of macrophages to LPS. However, the inhibitory effect was not seen if MCGs were added after macrophages had been exposed to LPS for 24 h. To assess whether MCGs could inhibit a non-LPS trigger, MCGs were tested on macrophages activated with IFN-γ. In these experiments, MCGs dose-dependently inhibited macrophage-mediated tumor cell killing induced by IFN- γ, LPS, or IFN-γ plus LPS. Furthermore, in parallel experiments, MCGs significantly inhibited macrophage NO production induced by LPS, IFN-γ, or IFN-γ plus LPS. Pretreatment of MCGs with diisopropylfluorophosphate, a serine protease inhibitor, only partially abrogated the effects of MCGs. The results demonstrate that MCGs inhibit both LPS- and IFN-γ-induced macrophage killing of P815 cells and the inhibition is associated with the decrease of NO production. J. Leukoc. Biol. 53: 446–453; 1993.
Isoprenaline inhibits thromboxane B2 release from U937 cells
Authors: Brightling CE , Baker AJ , Fuller RW
Published: January 2018

The U937 monocytic cell line and its differentiated macrophage-like form have been well characterized, illustrating many similarities with their analogous in vivo cells. We examined the release of thromboxane B2 (TXB2) from undifferentiated and differentiated cells after stimulation with opsonized zymosan and investigated whether the release of this mediator could be modified by isoprenaline. After stimulation, the U937 cells released TXB2 in a dose-dependent manner, with the release greater from the differentiated cells. The TXB2 released was inhibited by flurbiprofen (>10−8M;P < .01) and isoprenaline (>10−6 M; P < .01), and the inhibition was reversed by propranolol (10−6 M; P <.02). Thus, it is clear that undifferentiated and differentiated U937 cells release TXB2, which can be inhibited by β2- adrenoceptor stimulation. These findings illustrate an important functional difference between the in vivo macrophages and differentiated U937 cells because β2- adrenoceptor stimulation does not inhibit mediator release from macrophages. J. Leukoc. Biol. 53: 559–562; 1993.
Monocyte urokinase expression: modulation by interleukins
Authors: Gyetko Margaret R , Wilkinson Camille C , Sitrin Robert G
Published: January 2018

This study delineates the regulatory effect of interleukin-1 (IL-1) and interleukin-2 (IL-2) on monocyte plasminogen activator (PA) activity. Mononuclear phagocytes regulate net PA activity by modulating the expression of urokinase-type PA (uPA) and a specific plasminogen activator inhibitor, PA1-2. To understand the regulation of mononuclear phagocyte PA activity, it is important to compare the expression of uPA and PAI-2. In this study, we determined the relative abundance of secreted PA and PA inhibitor activity in human monocyte-conditioned medium after stimulation with human recombinant IL-1 or IL-2. In agreement with our previous description of tumor necrosis factor-α and interferon- γ stimulation of mononuclear phagocytes, we found no detectable PA activity in conditioned medium. Both IL-1 and IL-2 had dose-dependent effects, significantly up- regulating PA inhibitor activity in monocyte-conditioned medium (up to 11-fold). To further investigate the mechanism underlying this effect, Northern blot analysis was done to measure steady-state mRNA for uPA and PAI-2. Consistent with the increase in secreted PA inhibitor activity, we found that both IL-1 and IL-2 significantly increased steady-state mRNA for PAI-2. In addition, however, both IL-1 and IL-2 increased steady-state mRNA for uPA. IL-1 appears to increase mRNA for uPA to a greater extent than does IL-2. We conclude that IL-1 and IL-2 modulate monocyte proteolytic activity by increasing expression of uPA and PAI-2 with a resultant predominance of PAI-2. We further conclude that cytokine-specific regulation of plasminogen activity is achieved partly by varying the proportionate expression of uPA and PAI-2. J. Leukoc. Biol. 53: 598–601; 1993.
Mechanisms generating functionally heterogeneous macrophages: chaos revisited
Authors: Rutherford Mark S , Witsell Alice , Schook Lawrence B
Published: January 2018

Macrophage populations exhibit a wide range of antigenic and functional phenotypes, including cytokine production, response to immunomodulatory stimuli, and clearance of pathogens. The expanding clinical exploitation of recombinant growth factors and cytokines with the potential to regulate the production and function of peripheral macrophage populations necessitates an increased understanding of the mechanisms by which functionally distinct macrophage populations arise as well as the ramifications of macrophage heterogeneity. The present review summarizes recent data which supports multiple mechanisms by which heterogeneous macrophage populations arise: 1) differential signals experienced within diverse tissue microenvironments; 2) developmentally-staged expression of specific functions; 3) clonal variation of myeloid progenitor cells; and 4) alternate hematopoietic stimulation. These data show that the above processes are not mutally exclusive and that each likely contributes to the observed heterogeneity of peripheral macrophage populations. J. Leukoc. Biol. 53: 602–618; 1993.
Phospholipase C and phospholipase D are activated independently of each other in chemotactic peptide-stimulated human neutrophils
Authors: Mullmann Theodore J , Cheewatrakoolpong Boonlert , Anthes John C , Siegel Marvin I , Egan Robert W , Billah M Motasim
Published: January 2018

When cytochalasin B-treated neutrophils were stimulated with fMet-Leu-Phe (fMLP) in the presence of Ca2+, phospholipase C (PLC) activity, as measured by inositol-1,4,5-trisphosphate (IP3) formation, preceded phospholipase D (PLD)-catalyzed breakdown of choline- containing phosphoglycerides to form choline and dirady 1-sn -glycero-3-phosphate (phosphatidic acid), suggesting a possible link between PLC and PLD. However, in the absence of cytochalasin B or extracellular Ca2+, PLC was fully activated by fMLP with minimal activation of PLD, indicating that PLC activation alone is not sufficient for PLD activation. Full activation of PLD by fMLP required the simultaneous presence of both Ca2+ and cytochalasin B, a condition that caused no further enhancement of PLC. This result suggests that PLD products are not involved in the regulation of PLC activation. Furthermore, under conditions of complete inhibition of PLC by phorbol 12-myristate 13-acetate (PMA), there was no inhibition of PLD, showing that fMLP can activate PLD in the absence of PLC. Treatment of intact neutrophils with pertussis toxin inhibited both PLC and PLD, with PLC inhibition occurring at lower concentrations than PLD inhibition. These differential effects of pertussis toxin and the observed lack of inhibition of fMLP-stimulated PLD by PMA, which is believed to inactivate G-proteins involved in PLC activation, imply that PLC and PLD are linked to fMLP receptors through distinct G-proteins. Taken together, these observations suggests that, in fMLP-stimulated neutrophils, PLC and PLD are activated through independent mechanisms. J. Leukoc. Biol. 53: 630–635; 1993.
Stimulation of the bactericidal activity of polymorphonuclear leukocytes by manganese
Authors: Klebanoff Seymour J , Schlechte Kristie G , Waltersdorph Ann M
Published: January 2018

An antioxidant effect of manganese (Mn) complexes due to the scavenging of the superoxide anion (Of) or to the decomposition of hydrogen peroxide (H2O2) has been described. We report here that Mn also can exert a prooxidant effect under certain experimental conditions. Thus Mn2+ in phosphate buffer increased the bactericidal effect of phorbol myristate acetate-stimulated polymorphonuclear leukocytes (PMNs) on extracellular Escherichia coli. This effect was inhibited by azide, catalase, and a decrease in chloride concentration and was not observed when normal PMNs were replaced by those of patients with chronic granulomatous disease or myeloperoxidase (MPO) deficiency. Mn2+ could be replaced by Mn3+ or by superoxide dismutase (SOD). These findings suggest that Mn (or SOD), by increasing the conversion of Of to H2O2, can increase the activity of the MPO-H2O2-chloride antimicrobial system released by stimulated PMNs. J. Leukoc. Biol. 53: 666–672; 1993.
Chemotactic peptide–induced cytoplasmic pH changes in incubated human monocytes
Authors: Bernardo John , Brennan Lisa , Brink Holly F , Ortiz Maria F , Newburger Peter E , Simons Elizabeth R
Published: January 2018

Stimulation of phagocytic leukocytes with chemotactic factors results in transient acidification, followed by alkalinization of the cytosol. Human monocytes are known to alter their functional responses to the chemotactic peptide N-formylmethionyl-leucyl-phenylalanine (fMLP) in a complex fashion as they mature in vitro to macrophages. To examine the evolution of the cytoplasmic pH (pHi) response of monocytes to fMLP as they mature into macrophages, we incubated cells for 0, 24, 48, and 96 h (Medium-199 + 10% fetal bovine serum; 37°C) and examined pHi using the fluorescent probe 2′, 7′-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF; 1 μΜ) and a Perkin-Elmer 650/10 spectrofluorimeter (λem = 530 nm, λ_ex = 500, 450 nm) as previously described. The resting pH; of fresh (0 h) monocytes was 7.07 ± 0.16 (SD) and was unchanged after incubation for 24, 48, or 96 h (7.09, 7.11, 7.05, respectively). Cells exhibited an fMLP dose-dependent cytoplasmic acidification, with maximal δpHi occurring 30-60 s after exposure to 10−7 M fMLP. The response to fMLP did not change with the duration of incubation and, as with neutrophils, cytoplasmic realkalinization was blocked by dimethyl- amiloride (20 μΜ). Incubation with 2-deoxyglucose (10 min, 5 mM), sufficient to inhibit by more than 90% the formyl peptide-stimulated superoxide generation by monocytes, slowed fMLP-induced acidification and abrogated the alkalinization. In addition, monocytes isolated from the blood of a patient with X-linked chronic granulomatous disease (CGD) underwent fMLP-induced acidification that was unmasked further by coincubation with di- methylamiloride, in a manner quantitatively similar to that of normal monocytes, despite the inability of the CGD cells to produce superoxide. The chemotactic factor-induced cytoplasmic pH responses of mono- cytes/macrophages remained constant as the cells matured in vitro and exhibited a dimethylamiloride-independent acidification and dependent alkalinization, as did the response in neutrophils. The cytoplasmic acidification of these cells thus did not correlate with the cells’ production of superoxide and with the concomitant hexose monophosphate shunt activation, as has been suggested for other leukocyte types. J. Leukoc. Biol. 53: 673–678; 1993.