bluuebunny/crossref_metadata_2025 huggingface dataset:
Protease inhibitors (gabexate mesylate and ulinastatin) stimulate intracellular chemiluminescence in human neutrophils
Authors: Nishijima Junichi , Hiraoka Nobuaki , Murata Atsuo , Oka Yoshio , Kitagawa Kazunori , Tanaka Nobuo , Toda Hitoshi , Mori Takesada
Published: January 2018

The effect of protease inhibitors on the intracellular production of free radicals was investigated by measuring chemiluminescence (CL) elicited from phagocytosed luminol-bound microspheres (Lumispheres) in human neutrophils stimulated with formylmethionyl-leucyl-phenylalanine (fMLP), interleukin-8 (IL-8), phorbol 12-myristate 13-acetate, or diacylglycerol. Both gabexate mesylate (Foy) and ulinastatin (Miraclid), urinary trypsin inhibitor, increased intracellular CL in a dose dependent manner. Compared to control buffer without protease inhibitor, gabexate mesylate (322 μg/ml) caused about a 10-fold increase in intracellular CL in stimulated neutrophils, and ulinastatin (3100 U/ml) a twofold increase in neutrophils stimulated with fMLP or IL-8. When the protease inhibitors were added to the cell suspension after the phagocytosis of lumispheres, CL responses rapidly increased again to the level which was observed when both protease inhibitors and neutrophil stimulants were incubated simultaneously. In contrast, extracellular release of oxygen metabolites from stimulated neutrophils, assayed by a conventional measurement of luminol-dependent CL, was reduced by the protease inhibitors in a dose dependent fashion. When luminol-unbound microspheres were incubated with neutrophils stimulated by fMLP in luminol solution, extracellular CL was almost completely inhibited by gabexate mesylate. These results indicate that the protease inhibitors enhance the generation of intracellular CL and suppress the extracellular release of free radicals.
Levamisole causes differential cytokine expression by elicited mouse peritoneal macrophages
Authors: Kimball Edward S , Schneider Craig R , Fisher M Carolyn , Clark M Christina
Published: January 2018

Thioglycolate-elicited peritoneal macrophages from normal C57B1/6J mice were examined in vitro for bacterial lipopolysaccharide (LPS)-stimulated interleukin-1 (IL-1), IL-6, and tumor necrosis factor (TNF) production. Macrophages from mice administered a single oral dose of levamisole (3 mg/kg) 1 to 4 days prior to macrophage harvest demonstrated a twofold enhancement of IL-1 production compared to vehicle-treated mice. In contrast, IL-6 production and TNF production by the same macrophages were inhibited up to 36 and 62%, respectively, compared to production by macrophages harvested from vehicle-treated mice. Similar results were observed when IL-1 production and TNF production were followed in peritoneal exidate cells directly stimulated with levamisole in vitro. The ex vivo LPS-stimulated IL-1 production was enhanced 4 days after macrophage elicitation, whereas TNF and IL-6 production returned to baseline by 72 h after macrophage recruitment and augmentation. No evidence could be found for the presence of inhibitors of TNF or IL-6. The specificity of the IL-1, IL-6, and TNF bioactivities was demonstrated by neutralization with specific antisera. Immunoprecipitation studies of supernatants from biosynthetically labeled macrophages also revealed augmented IL-1 production and decreased IL-6 and TNF, indicating that levamisole may have affected cytokine production at the translational level. Kinetics studies revealed that ex vivo release of IL-6 and TNF by macrophages from levamisole-dosed mice was delayed compared to production of these cytokines by macrophages harvested from mice given vehicle only. The results may explain, in part, the reported ability of levamisole to ameliorate cases of rheumatoid arthritis or other autoimmune and inflammatory diseases by affecting the relative levels of cytokines produced by macrophages recruited to sites of injury, which are associated with inflammation and acute-phase protein synthesis.
Interferon-α -dependent and -independent participation of accessory cells in natural killer cell-mediated lysis of HSV-1-infected fibroblasts
Authors: Feldman Michael , Howell Donna , Fitzgerald-Bocarsly Patricia
Published: January 2018

Abstract: Human natural killer (NK) cells require an HLA-DR+ accessory cell (AC) population to lyse herpes simplex virus-type 1 (HSV-l)-infected fibroblasts (HSV-Fs) but not K562 target cells. It has been postulated that ACs may function by producing interferon-α (IFN-α), which stimulates NK cells. Using a sequential enrichment protocol, ACs were .found to coenrich with the interferon- producing cells (IPCs). Treatment of the ACs with a protein synthesis inhibitor, emetine, inhibited both their IFN production and AC function, results that support a central role for IFN in AC activity. In contrast, when the arginine analogue canavanine was added to NK assays, no IFN-a was produced and NK(HSV-Fs) activity was only partially inhibited. Consistent with IFN-independent AC function, treatment with either polyclonal sheep or bovine anti-IFN-α neutralized all the IFN-α produced during the NK assays but caused either no or partial reduction of NK(HSV-Fs) activity, respectively. However, when limiting numbers of ACs were used, the bovine antiserum neutralized both IFN-α and NK(HSV-Fs) activity. We further found that HLA-DR+ cells are required for cell clustering, suggesting a role for cell contact. Finally, fixation of activated ACs prevented the accessory function. Together, these results demonstrate that ACs can provide help to NK cells in both IFN-α -dcpendcnt and -independent manners.
Ontogenic development of macrophage subpopulations and la-positive dendritic cells in fetal and neonatal rat spleen
Authors: Takeya Motohiro , Takahashi Kiyoshi
Published: January 2018

Development, differentiation, and distribution of macrophage subpopulations and la+ dendritic cells in the fetal and neonatal rat spleen were investigated by means of double immunohistochemical staining and im- munoelectron microscopy. To characterize these cell populations, a panel of anti-rat macrophage monoclonal antibodies (RM-1, ED2, ED3, TRPM-3, Ki-M2R) and an anti-rat la antibody (OX6) were used. In the fetal rat spleen, macrophages were first detected by RM-1 at fetal day 15. ED2+ and/or Ki-M2R+ macrophages appeared at fetal day 16. TRPM-3+ and/or ED3+ macrophages appeared a day later. During the fetal and neonatal development, ED2+ and TRPM-3+ macrophages differentiated independently, maturing into red pulp macrophages and marginal metallophilic and marginal zone macrophages respectively. Intimate topographical relations were observed between ED2+ macrophages and hematopoietic cells and between TRPM-3+ macrophages and marginal zone lymphocytes. Ia+ cells were first observed around arterioles at fetal day 15. In the fetal and neonatal period, the number of la+ cells gradually increased, their shape became dendritic, and they matured into interdigitating cells in the inner periarteriolar lymphatic sheath. In ontogeny, la+ dendritic cells were not stained with ED2 or TRPM-3. These results suggest that ED2+ macrophages, TRPM-3+ macrophages, and Ia+ dendritic cells are distinct cell lines that pursue independent developmental processes in spleen ontogeny.
Monocyte diapedesis through an in vitro vessel wall construct: inhibition with monoclonal antibodies to thrombospondin
Authors: Huber Andres R , Ellis Susan , Johnson Kent J , Dixit Vishva M , Varani James
Published: January 2018

Human peripheral blood monocytes were examined for migration across an endothelial cell monolayer in an in vitro vessel wall construct. Few monocytes invaded in the absence of a chemotactic gradient, despite significant adhesion to the endothelial monolayer. However, the addition of zymosan-activated human plasma to the lower compartment, to create a chemotactic gradient across the vessel wall, resulted in significantly enhanced monocyte migration. Pretreatment of the monocytes with monoclonal antibodies to thrombospondin (TSP) dramatically inhibited monocyte diapedesis into the vessel wall. The same treatment inhibited monocyte adhesion to endothelial cells in two-dimensional monolayer cultures as well as in vessel wall constructs (no chemotactic gradient). Of interest, however, the monoclonal antibodies had no inhibitory effect on monocyte migration into collagen gels devoid of endothelial cells in response to the same chemotactic gradient, suggesting the importance of TSP in monocyte-endothelial cell interactions. Monoclonal antibodies to fibronectin and normal mouse immunoglobulin G did not inhibit migration in this model of a vessel wall. Furthermore, monoclonal antibodies to TSP showed no inhibition of human peripheral blood neutrophil migration. Previous studies have shown that monocytes synthesize TSP and express this moiety on their surface. The present data suggest that monocytes may utilize TSP to interact with endothelial cells lining the vessel wall during diapedesis.
Isolation and characterization of macrophages from rat embryonic muscle culture
Authors: Honda Hitoshi , Kimura Hiromitsu , Rostami Abdolmohammad
Published: January 2018

We have previously described the wide distribution of resident macrophages in normal rat skeletal muscle. In this study, we investigated the characteristics of the macrophages that occur in rat embryo muscle cultures. We showed that cells of monocyte-macrophage lineage are present in primary muscle cultures of rat embryos (18 days in gestation) and that these cells form morphologically and phenotypically heterogeneous populations, based on their reaction with monoclonal antibodies EDI, ED2, ED3, and OX43. Constitutively la+ cells with dendritic appearance were also observed. Furthermore, we established the procedure for isolation of macrophages from the primary muscle cultures. The isolated cells, mostly EDI+, expressed class I and CD4 antigens and bore complement (C3) receptors on their surfaces. The fact that cell of monocyte-macrophage lineage occur in the embryonic muscle suggests that during embryogenesis these cells may enter the developing muscle and give rise to a population of tissue-associated macrophages.
Augmented expression and release of nonspecific cross-reacting antigens (NCAs), members of the CEA family, by human neutrophils during cell activation
Authors: Kuroki Motomu , Matsuo Yoshino , Kinugasa Tetsushi , Matsuoka Yuji
Published: January 2018

Nonspecific cross-reacting antigens (NCAs) are a group of human glycoproteins immunologically cross-reactive with carcinoembryonic antigen (CEA). Our previous studies have shown that at least seven NCA glycoproteins different in molecular weight and antigenic reactivity, including a species corresponding to CD67, can be detected in neutrophil granulocytes. In the present paper, it is demonstrated that neutrophil activation induced with soluble stimulators, the calcium ionophore A23187, N-formylmethionyl-leucyl-phenylalanine, and phorbol myristate acetate, results in augmented release and cell surface expression of NCAs. The NCA release was correlated with the discharge of azurophil granules but not with that of specific granules and was attributable to the release of NCA species of 53 and 30 kd. The increased NCA expression on the cell surface was due to increments of the NCAs of 160, 100 (CD67), 95, 90, 30, and 26 kd. These results, together with the previous findings that the CEA family members can mediate intercellular adhesion and bind Escherichia coli in vitro, imply that the neutrophil NCAs participate in the functions of neutrophils such as phagocytosis, chemotaxis, and adherence.
Interleukin-2 signal transduction in human NK cells: multisite phosphorylation and activation of the tyrosine kinase p56lCk
Authors: Einspahr Kregg J , Abraham Robert T , Dick Christopher J , Leibson Paul J
Published: January 2018

Interleukin-2 (IL-2) potently stimulates natural killer (NK) cell proliferation and cytotoxic function. However, the molecular mechanisms by which IL-2 delivers activation signals from the IL-2 receptor to the NK cell interior are incompletely understood. Previous studies demonstrated that IL-2 stimulation induced the tyrosine phosphorylation of multiple proteins in NK cells, together with a prominent reduction in the elec- trophoretic mobility of p56lac. The present studies indicate that IL-2 induces a rapid (≤1 min) increase in the catalytic activity of p56lck, as measured by increases in protein tyrosine kinase activity in vitro. Furthermore, in response to IL-2, p56lck itself undergoes complex alterations in serine and tyrosine phosphorylation. Cyanogen bromide cleavage maps indicate that IL-2 stimulates a pronounced increase in the phosphorvlation of the NH2-terminal region of p56lck containing multiple known sites of serine phosphorylation. In addition, IL-2 induced a marked increase in the phosphorylation of a COOH-terminal peptide containing the regulatory Tyr-505 residue of p56lck. These results suggest that p56lck serves as a substrate for both protein serine and tyrosine kinases activated during stimulation of this cell type with IL-2. Furthermore, these results indicate that the pleiotropic effects of IL-2 on NK cell physiology are initiated and regulated by a complex and multitiered interaction of different protein kinases including p56lck.
The gene for human eosinophil Charcot-Leyden crystal protein directs expression of lysophospholipase activity and spontaneous crystallization in transiently transfected COS cells
Authors: Zhou Zeqi , Teneri Daniel G , Dvorak Ann M , Ackerman Steven J
Published: January 2018

Expression of the gene encoding human eosinophil lysophospholipase, the Charcot-Leyden crystal (CLC) protein, was studied in transiently transfected COS cells. Recombinant CLC (rCLC) protein expression was demonstrated both by Western blot and radioimmunoassay inhibition analyses of transfected COS cell extracts and by immunofluorescent staining and ultrastructural im- munogold analyses of intact cells. The rCLC protein was immunochemically indistinguishable from native eosinophil-derived CLC protein, and each transfected COS cell expressed approximately 11 pg of rCLC protein as determined by radioimmunoassay and assessment of transfection efficiency. Immunofluorescent microscopy and ultrastructural immunogold analyses localized rCLC protein to the nucleus, cytoplasm, and plasma membrane of COS cells. Lysates from transfected COS cells producing CLC protein expressed significant lysophospholipase activity. Furthermore, rCLC protein expressed in COS cells spontaneously formed the distinctive intracytoplas- mic and intranuclear hexagonal bipyramidal crystals characteristic of the native eosinophil and basophil- derived protein. Expression of the CLC gene confirms the authenticity of the CLC cDNA, the expression of lysophospholipase activity by this unique eosinophil and basophil constituent, and will facilitate the routine purification of the active enzyme for in vitro and animal model studies of its role (or roles) in eosinophil and basophil associated allergic inflammation and eosinophil- parasite interactions.
Organic peroxides inhibit neutrophil leukotriene B4 biosynthesis
Authors: Okazaki Ian J , Newman Lisa M , Allen David W
Published: January 2018

Leukotriene B4, an autacoid metabolite of arachidonic acid produced by polymorphonuclear neutrophils, induces chemokinesis, chemotaxis, and adhesion of these cells at sites of inflammation. Because neutrophil infiltration is a self-limited process, we hypothesized that oxidized lipid products of neutrophil-damaged tissue might inhibit leukotriene B4 biosynthesis, thereby preventing additional neutrophil infiltration and limiting peroxidative tissue damage. Erythrocyte ghosts exposed to a hydrogen peroxide-generating system served as a model of peroxidized tissue in inflammation and inhibited neutrophil leukotriene B4 production by 50% compared with unoxidized ghosts. Organic peroxides, including tert-butylhydroperoxide, peracetic acid, and linoleic hydroperoxide, resembling the product(s) of tissue membrane peroxidation in lipid solubility and cata- lase resistance, inhibited leukotriene B4 biosynthesis in a dose-dependent manner (50% inhibitory concentration of 3.9 μM compared to 530 μM for H2O2). Biosynthetic steps prior to the 5-lipoxygenase did not appear to be the site of inhibition. Likewise, the step after the 5-lipoxygenase, the leukotriene A4 hydrolase, was not primarily involved. Thus a possible mechanism for controlling the influx of neutrophils and their oxidative damage during inflammation may be inhibition of the 5-lipoxygenase by catalase-resistant lipid peroxides released by tissue membranes.
Regulation of herpes simplex virus type 1 gene expression in nonpermissive murine resident peritoneal macrophages
Authors: Wu Linxian , Morahan Page S , Leary Kathryn
Published: January 2018

The restriction of herpes simplex virus type 1 (HSV-1) gene expression in nonpermissive murine resident peritoneal macrophages (ResPMO) was analyzed at the level of transcription. Using nuclear run-on assays, we established that transcription of a representative gene from each HSV-1 kinetic class was initiated in both ResPMO and permissive Vero cells. Using transient expression assays, we observed that the promoters of two early genes were activated by the immediate-early protein ICP4 in ResPMO. However, the level of trans- activation of these promoters by another immediate-early protein, ICPO, was markedly reduced in ResPMO. Furthermore, the synergistic trans-activation normally observed between ICPs 4 and O in Vero cells was noticeably absent in ResPMO. The data indicate that the block in viral gene expression in ResPMO occurs after initiation of viral gene transcription and involves dysfunction of a viral immediate-early regulatory protein, ICP0. J. Leukoc. Biol. 53: 61–65; 1993.
Activation of primary lymphocytes requires prolonged lectin stimulation
Authors: Nash Patricia V , Mastro Andrea M
Published: January 2018

T lymphocytes are activated by a complex series of events, but the mechanisms remain unclear. One uncertainty is the time of receptor-ligand interaction necessary for commitment to DNA synthesis and proliferation. Although this issue has broad implications for the interpretation of T cell activation data, it remains unresolved. Therefore, we examined the temporal activation requirements of rat splenocytes stimulated with con- canavalin A (Con A) by measuring proliferation, as well as interleukin-2 (IL-2) production and IL-2 receptor (IL-2R) expression. Splenocytes stimulated with various Con A concentrations for 3 h did not incorporate significantly more [3H] thymidine than unstimulated splenocytes. Some increase occurred after 6 h of lectin exposure but maximum proliferation occurred only after the 52-h stimulation. Furthermore, Con A incubations of 6 h or more were required for significant increases in IL-2 or IL-2R. Maximum lymphokine production and receptor expression were observed after the 52-h stimulation. Thus, activation of some primary lymphocytes required only 6 h of stimulation, but much longer mitogen contact was necessary for maximum recruitment. J. Leukoc. Biol. 53: 73–78; 1993.
Thromboxane A2 receptors in equine monocytes: identification of a new subclass of TXA2 receptors
Authors: Simmons T Robin , Cook James A , Moore James N , Halushka Perry V
Published: January 2018

Thromboxane (TX) A2 has been implicated as an important pathophysiologic mediator of a variety of cardiovascular diseases. Monocytes synthesize TXA2 and it modulates their function. This study sought to characterize monocyte TXA2 receptors. Radioligand binding studies were performed on membranes prepared from equine peripheral blood monocytes using [125I]BOP, a TXA2 receptor agonist. [125I]BOP bound to a single class of binding sites (Kd - 1.0 ± 0.3 nM and Bmax = 389 ± 191 fmol/mg protein; n = 5). Several TXA2 receptor agonists and antagonists competed for binding with [125I]BOP. I- BOP produced a concentration-dependent inhibition of endotoxin-induced tumor necrosis factor (TNF) activity (IC50 = 9.6 ± 2.5 nM; n = 5). In contrast to its effects in platelets and vascular smooth muscle, I-BOP significantly increased cAMP formation in monocytes (EC50 = 22 ± 3.6 nM; n = 4). The TXA2 receptor antagonists SQ29548 (5.6 nM) and L657925 (0.13 μM) significantly blocked I-BOP- stimulated cAMP formation but did not block 250 nM prostaglandin E2-stimulated cAMP formation. These data support the presence of a TXA2 receptor in equine peripheral blood monocytes. Activation of this receptor results in suppression of endotoxin-induced TNF formation and stimulation of cAMP production. Increased cAMP production after receptor activation suggests that this receptor may represent a unique subclass of TXA2 receptors. J. Leukoc. Biol. 53: 173–178; 1993.
Hematopoietic recovery in repeatedly irradiated mice can be enhanced by a repeatedly administered combination of diclofenac and glucan
Authors: Hofer M , Pospíšil M , Viklická š , Vacek A , Pipalovà I , Bartoničková A
Published: January 2018

A combination of diclofenac and glucan administered repeatedly in a protective regimen in the course of repeated gamma irradiation of mice (6 × 2 Gy during 3 weeks) enhanced granulopoiesis and other indices of hematopoietic recovery investigated from 3 to 7 days after the last radiation exposure. Repeated administration of diclofenac or glucan alone or treatment of the mice with the didofenac-glucan combination given once before the first or the last radiation exposure did not induce such effects. The protective effect of the repeatedly administered combination of the drugs was realized despite the fact that the response of the serum colony-stimulating activity to the repeated combined drug administration was decreased at the end of the treatment regimen compared to that of mice given this drug combination only once. The combined treatment is supposed to act via increased proliferation of the hematopoietic stem or progenitor cells. Additivity or even synergism of the hematostimulatory action of glucan and of the strengthening of positive control of cell proliferation achieved by removing negatively acting prostaglandins (diclofenac action) may account for the radioprotective effects observed. J. Leukoc. Biol. 53: 185–189; 1993.
Centrifugal enhancement of human immunodeficiency virus type 1 infection and human cytomegalovirus gene expression in human primary monocyte/macrophages in vitro
Authors: Ho Wen-Zhe , Cherukuri Ravi , Ge Shun-Di , Cutilli Joann R , Song Li , Whitko Stephanie , Douglas Steven D
Published: January 2018

In an effort to facilitate the efficiency of human immunodeficiency virus type 1 (HIV-1) and/or human cytomegalovirus (HCMV) infection in primary monocyte/macrophages in vitro, the effect of low-speed centrifugation was studied. The infectivity of Aree strains (Bai, Ada-M, and IIIB) of HIV-1 tested was significantly enhanced by centrifugal inoculation at a force of 1500g for 60 min. Reverse transcriptase activity and HIV-1 p24 antigen in primary monocyte/macrophages infected by a centrifugal inoculation technique were detectable 3-7 days earlier and were more than 10-fold greater in magnitude (at an early stage of the infection) than those of control cells infected by the conventional inoculation technique. Examination of the cells by indirect immunofluorescence revealed higher expression of HIV-1 p24 protein in the monocyte/macrophages infected by the centrifugal inoculation technique. These differences were directly related to centrifugal inoculation and were evident up to 3 weeks after infection. Enhancement was not observed when centrifugation was carried out before or after HIV-1 infection. Centrifugal inoculation of HCMV also enhanced its immediate-early and early gene expression up to 36- to 50-fold, although neither late nuclear antigens and glycoproteins of HCMV nor infectious virus was detected in HCMV-infected monocyte/ macrophage cultures. These results show that centrifugal inoculation is a useful technique for improving the efficiency of HCMV and HIV-1 infection in vitro. J. Leukoc. Biol. 53: 208–212; 1993.
Recombinant soluble IgA Fc receptor: generation, biochemical characterization, and functional analysis of the recombinant protein
Authors: Maliszewski Charles R , VandenBos Timothy , Shen Li , Schoenbo Michael A , Kubagawa Hiromi , Beckmann M Patricia , Monteiro Renato C
Published: January 2018

We previously described the cloning of a human myeloid cell surface receptor for the Fc region of immunoglobulin A (FcαR). In the present study, a soluble version of the FcαR (solFcαR) was generated by removing the transmembrane and cytoplasmic coding regions from full-length FcocR cDNA and ligating into a mammalian expression vector. COS-7 cells transfected with the solFcαR plasmid secreted a protein that inhibited both immunoglobulin A (IgA) and anti-FcaR monoclonal antibody (mAb) binding to FcαR+ U937 cells. Furthermore, the solFcαR bound specifically to and could be eluted from an anti-FcαR mAb-immunoaffinity column, retaining biological activity. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that the recombinant full-length FcαR migrates over a molecular mass range of approximately 40-60 kd, consistent with the reported size and heterogeneity of the naturally occurring myeloid cell surface FcocR. The solFcocR ran on SDS-PAGE as a smaller band (37-55 kd) that reduced to two bands of 23 and 25 kd following N-glycanase treatment, indicating that the FcαR is a heavily glycosylated protein. The biochemical data, coupled with flow cytometry studies showing that the recombinant FcαRs bind to five different anti-FcαR mAbs, clearly demonstrate that the cloned FcαR corresponds directly to the major FcαR species expressed on human monocytes, neutrophils, and myeloid cell lines. The generation of soluble receptor protein will permit in-vestigations of the role of FcocR in IgA-mediated im-munoregulation, effector functions, and disease. J. Leukoc. Biol. 53: 223–232; 1993.
Silica-exposed macrophages release a growth-promoting activity for type II pneumocytes
Authors: Melloni Boris , Lesur Olivier , Cantin André , Bégin Raymond
Published: January 2018

This study addresses the hypothesis that silica- activated alveolar macrophages could release soluble factors stimulating type II cell proliferation. Macrophages from control sheep were exposed or not in vitro to silica, aluminum-treated silica (Si-Al), or titanium dioxide (TiO2). In addition, macrophages from a model of chronic silica-exposed sheep were studied. Supernatants from unstimulated macrophages were found to double basal type II cell DNA synthesis. Alveolar macrophage-conditioned media (AMCM) collected from cells exposed in vitro to silica induced an additional growth-promoting activity for type II cells. Supernatants from Si-Al- or TiO2 dust-exposed alveolar macrophages had the same effects as unstimulated AMCM. In addition, AMCM from in vivo silicotic sheep cells also contained elevated levels of type II cell mitogenic activity. These results suggest that in vitro as in vivo, silica-activated macrophages produce a type II cell growth factor(s) for type II epithelial cells. J. Leukoc. Biol. 53: 327–335; 1993.
Human pulmonary macrophages utilize prostaglandins and transforming growth factor β1 to suppress lymphocyte activation
Authors: Roth Michael D , Golub Sidney H
Published: January 2018

The ability to activate peripheral blood lymphocytes (PBLs) in vitro with interleukin-2 (IL-2) is suppressed by the presence of autologous human pulmonary alveolar macrophages (AMs). AMs suppress both IL-2-induced proliferation and the induction of lympho- kine-activated killer cell (LAK) activity in a dose- dependent manner (79 ±6% suppression of LAK activity at a 0.25:1 AM/PBL ratio). Increasing the IL-2 concentration increased baseline LAK activity but did not prevent AM-mediated suppression. At least two different mechanisms of suppression were observed, one diffusible in nature and the other contact dependent. Indomethacin prevented the component of inhibition that diffused across porous polycarbonate membranes, indicating prostaglandins as the diffusible inhibitor. In contrast, indomethacin had no effect when added alone into conventional AM- PBL cocultures, but a combination of indomethacin and anti-transforming growth factor β1 (TGF-β1) antibody did prevent inhibition. This result suggests that TGF-ß1 acts as an additional contact-dependent inhibitor. PBLs that were rendered unresponsive to IL-2 completely recovered their responsiveness within 4 days after removing AMs from the coculture. These features suggest that pulmonary macrophages have multiple mechanisms for locally suppressing IL-2 responses and lymphocyte activation. J. Leukoc. Biol. 53: 366–371; 1993.
Transmembrane signaling pathways involved in phagocytosis and associated activation of NADPH oxidase mediated by FcγRs in human neutrophils
Authors: Bianca Vittorina Della , Grzeskowiak Miroslawa , Dusi Stefano , Rossi Filippo
Published: January 2018

We have previously shown that in neutrophils classical transmembrane signaling consisting of increased [Ca2+]i and hydrolysis of phospholipids was not essential for phagocytosis mediated by more than one receptor (yeast-IgG, yeast-C3b/bi, yeast-Con A). This work deals with the role of this transmembrane signaling in phagocytosis of erythrocyte (E) IgG, which is mediated only by receptors for IgG (FcγRs). The ingestion of E-IgG was associated with an increase in [Ca+]i and production of inositol phosphates, phosphatidic acid, diacylglycerol, and arachidonic acid, via activation of phospholipases G, D and A2. Related to the same number of particles ingested, the respiratory burst and the transmembrane signaling during phagocytosis of E-IgG were much smaller than during phagocytosis of yeast-IgG. In Ca2+-depleted neutrophils, where the increase in [Ca2+]i and hydrolysis of phospholipids were lacking, the phagocytosis of E-IgG was depressed by about 60%; the respiratory burst was also depressed due to the decrease of ingestion and of stimulation of NADPH oxidase by residual phagocytosis. Pertussis toxin (PT) did not inhibit the phagocytosis of E- IgG but depressed by about 40% the stimulation of li- pidic transmembrane signaling and the respiratory burst in normal neutrophils. In Ca2+-depleted neutrophils the toxin was without effect on ingestion and respiratory burst. Staurosporine did not inhibit the ingestion of E- IgG in normal and Ca2+-depleted neutrophils but depressed by 30-40% the respiratory burst in normal and not in Ca2+-depleted neutrophils. Genistein, an inhibitor of tyrosine kinase, did not inhibit the ingestion of E-IgG but depressed by 30-40% the respiratory burst both in normal and Ca2+-depleted neutrophils. These results demonstrate the following findings in human neutrophils. (1) Contrary to the phagocytosis mediated by more than one receptor (yeast-IgG, yeast-Con A, yeast-C3b/bi), the transmembrane signaling involving increase in [Ca2+]i and hydrolysis of phospholipids plays a role in the phagocytosis and respiratory burst mediated by FcγRs alone. Thus, different signal transduction pathways can be involved in phagocytosis and associated respiratory burst depending on the receptor or combination of receptors activated. (2) FcγRs alone promote phagocytosis with two signaling pathways independent of and dependent on [Ca2+]i changes and phospholipid hydrolysis and insensitive to PT, staurosporine, and genistein. (3) The signaling pathways promoting phagocytosis triggered by FcγRs alone are in some way, or at some step, different from those that activate the respiratory burst. J. Leukoc. Biol. 53: 427–438; 1993.
Hypoxia reoxygenation-induced injury of cultured pulmonary microvessel endothelial cells
Authors: Wiles Marc E , Hechtman Herbert B , Morel Nicole ML , Shepro David
Published: January 2018

Polymorphonuclear leukocyte (PMN) sequestration within the pulmonary microvasculature is known to occur in association with ischemia/reoxygenation (I/R). This sequestration is dependent on eicosanoids and reactive oxygen species. PMN sequestration within the lungs suggests that pulmonary microvascular endothelial cells (MECs) may in part regulate the I/R response. Simulating I/R, we examined the effect of hypoxia/reoxy- genation (H/R) on pulmonary MECs in vitro, with and without PMNs. Significant cellular injury, assessed by 51Cr release, occurred upon reoxygenation of MECs (P < .01). Addition of PMNs to the H/R-injured monolayers did not increase MEC injury. Reoxygenation of MECs also resulted in increased thromboxane (Tx) B2 production compared to controls (P < .01). Inhibition of Tx secretion by aspirin reduced H/R-induced PMN adhesion to MECs (P< .01). Furthermore, H/R-induced increases in PMN-MEC adhesion were prevented by al- lopurinol and superoxide dismutase (P< .01). These data suggest that the pulmonary response to H/R is mediated by MEC generation of reactive oxygen radical species and Tx, which promotes increased PMN adhesion. J. Leukoc. Biol. 53: 490–497; 1993.
Separation of alveolar macrophages and dendritic cells via autofluorescence: phenotypical and functional characterizatior
Authors: Havenith Carin EG , Breedijk Annette J , van Miert Paula PMC , Blijleven Nico , Calame Wim , Beelen Robert HJ , Hoefsmit Elisabeth CM
Published: January 2018

In a previous study we demonstrated that an increase of monocytes and dendritic cells (MDC) was found in the bronchoalveolar lavage fluid (BAL) after intratracheal instillation of a bacillus Calmette-Guérin suspension. In the current study, the bright autofluorescence of alveolar macrophages (AMs) was used to separate them efficiently from the MDC. Sorting of freshly isolated BAL cells resulted in a high-autofluorescent fraction, consisting predominantly of AMs, and a low-autofluorescent fraction containing the MDC, lymphocytes, and granulocytes. Thus, a clear separation between suppressive (AM) and stimulating (MDC) activity was obtained as shown in antigen-specific T cell responses. Flow cytometric parameters, density fractionation, and a series of ED monoclonal antibodies raised against rat macrophage antigens showed that both AMs and MDC were diverse populations. After overnight culture, more than 80% of an MDC population with a density range of 1.065-1.079 changed to a lower density (< 1.056) and morphologically developed into DCs with many processes. Concomitantly, monoclonal antibody EDI expression changed from a granular pattern to a discrete juxtanuclear spot localization. J. Leukoc. Biol. 53: 504–510; 1993.
A lipopolysaccharide-inducible macrophage gene (D3) is a new member of an interferon-inducible gene cluster and is selectively expressed in mononuclear phagocytes
Authors: Tannenbaum Charles S , Major Jennifer , Ohmori Yoshihiro , Hamilton Thomas A
Published: January 2018

We previously reported the isolation and characterization of cDNA clones encoding novel lipopolysac- charide (LPS)-inducible mRNAs from murine peritoneal macrophages. We now present the complete coding sequence of a cDNA previously termed D3. Analysis of multiple clones from a murine macrophage cDNA library provided a complete cDNA sequence of approximately 1.6 kb. The corresponding RNA contains a single open reading frame encoding a hydrophilic protein composed of 425 amino acids and is characterized by a region including three perfect and two imperfect repeats of a seven- amino-acid sequence. Based on nucleotide and deduced amino acid sequence, this mRNA is a new member of a previously described multigene cluster of interferon- inducible genes termed the Mouse 200 series genes. This new sequence most closely resembles gene 204 because both D3 and 204 genes have segments containing the seven-amino-acid repeat sequence. The Mouse 202 and 204 genes, however, have an approximately 200-amino- acid carboxyl-terminal domain that is absent in the LPS- inducible macrophage-derived cDNA. In addition, D3, 202, and 204 can all be distinguished from one another by virtue of unique 3′ noncoding regions 200-300 base pairs in length. The D3 unique sequence is largely restricted to the smallest of the three size classes of this gene family expressed in macrophages and is not detected in interferon- or platelet-derived growth factor-stimulated fibroblasts. Overall, three separate mRNAs have now been described, each of which has three or more of a possible seven nucleotide sequence domains. Although the functions) of the members of this gene family remains unknown, the multiple forms inducible by diverse stimuli and their restricted cell type expression suggest diverse and important physiologic roles for their products in inflammation. J. Leukoc. Biol. 53: 563–568; 1993.
Evidence for integrins other than β2 on polymorphonuclear neutrophils: expression of α6β1 heterodimer
Authors: Rieu Philippe , Lesavre Philippe , Halbwachs-Mecarelli Lise
Published: January 2018

The expression of non-β2 integrins on polymorphonuclear neutrophils (PMNs) was analyzed by im- munoprecipitation and flow cytometry using platelet-free PMN preparations and anti-FcγR blocking mAbs. No β3 integrin was detected with six anti-β3 mAbs. Conversely, integrin β1 chain was present on PMNs, although at low level, and could be distinguished from platelet β1 by SDS-PAGE. The MW differences disappeared after N- glycanase treatment. PMNs express only 2500 molecules of β1 per cell and this expression is not modulated by agonists such as phorbol myristate acetate, formylmethionyl- leucyl-phenylalanine, granulocyte-macrophage colony- stimulating factor, or tumor necrosis factor α, which enhance CDllb expression, or by interferon-7 or transforming growth factor α. PMNs were found to express α6 associated with β1, and no reactivity was observed with various anti-α1, anti-α2, anti-α3, anti-α4, anti-α5 or anti-αv mAbs. In conclusion, although other leukocytes express various β1 integrins, which mediate cell interactions with ECM proteins, PMNs appear to express only the laminin receptor α6β1 PMN interactions with nonlaminin ECM ligands thus seem to be mediated either exclusively by β2 integrins or by nonintegrin molecules. J. Leukoc. Biol. 53: 576–582; 1993.
Multiple pathways of interferon-induced gene expression in murine macrophages
Authors: Politis Alexander D , Sivo Judit , Vogel Stefanie N
Published: January 2018

Steady-state levels of mRNAs that encode specific FcyR and la antigen genes have been measured in macrophages treated with interferons (IFNs) to examine the induction of these markers at the molecular level. Our previous studies suggested requirement for protein kinase C (PKC) in the IFN induction of these macrophage surface markers, although a difference in PKC dependence was found between IFN-α/β- and IFN-γ-induced FcγR expression. The protein kinase antagonist H 7, used previously to distinguish between the surface induction of FcyR by IFN-α/β and IFN-γ, also distinguishes between the IFN-γ and IFN-γ in the induction of FcyRI mRNA and FcyRI surface expression. Protein kinase inhibitors blocked the IFN-γ induction of la mRNA in a manner similar to that reported previously for cell surface la expression. It is concluded that FcyRI is induced by both IFN-γ and IFN-γ through distinct biochemical pathways, whereas IFN-γ utilizes distinct pathways to induce the two macrophage activation markers, la antigen and FcyRI. J. Leukoc. Biol. 53: 583–590; 1993.
Respective role of polymorphonuclear leukocytes and their integrins (CD-11/18) in the local or systemic toxicity of lipopolysaccharide
Authors: Chang Hernán R , Vesin C , Grau Georges E , Pointaire R , Arsenijevic Denis , Strath Malcolm , Pechdre Jean-Claude , Piguet Pierre-Frangois
Published: January 2018

The role of neutrophils (PMNs) and leukocyte integrins was investigated in two models of lipopolysac- charide (LPS)-induced toxicity: the systemic lethality assay in D-galactosamine-sensitized mice and the local reaction elicited by intradermal injection of LPS and tumor necrosis factor (TNF) at 24-h intervals. In the local reaction, depletion of PMNs with an anti-PMN monoclonal antibody (mAb) and mAbs against CD-11a (or LFA1) and CD-llb (or CR3) completely prevented the hemorrhagic necrosis. Evaluation of histological sections and myeloperoxidase levels suggested different mechanism of protection because PMNs were abundant in anti- CD-11- and absent in anti-PMN-treated mice. In the systemic assay, depletion of PMNs ensured 100% survival, whereas after administration of anti-CD-1 la or b mAb, the percentages of survivors were 6 and 59%, respectively. One hour after LPS injection, the serum TNF-α level was higher in PMN-depleted mice than in controls. These studies provide evidence that neutrophils are essential for the expression of local or systemic LPS-induced injury, whereas the requirement for their leukocytic integrins is obvious only in the local reaction. J. Leukoc. Biol. 53: 636–639; 1993.
The in vivo effects of PIXY321 therapy on human monocyte activity
Authors: Hamilton Raymond F , Vadhan-Raj Saroj , Uthman Margaret , Grey Marjorie , Holian Andrij
Published: January 2018

In this report we describe the time-dependent effects of PIXY321 (a synthetic hybrid cytokine) treatment (500 and 750 μg/m2/day for 14 days) on six sarcoma patients. Blood was taken prior to PIXY321 injection (day 0), on days 1, 7, and 14 of treatment, and 7 days posttreatment (day 21). The number of isolated monocytes quadrupuled by day 7 and sustained a significant increase through day 14. There were significant increases in the percentage of circulating monocytes relative to total mononuclear cells on days 1 and 7 of therapy. There were no significant changes in monocyte cell surface antigens (15 checked), suggesting that the increase in monocyte numbers was not due to increased numbers of immature monocytes. The basal activity of the monocytes was not markedly altered during treatment; however, they were primed for significantly increased phorbol 12,13- dibutyrate-stimulated superoxide anion production and endotoxin-stimulated release of interleukin-1 β (IL-lβ) and tumor necrosis factor-α (TNF-α) on days of 1 and 7 of therapy. There was a significant increase of IL-β mRNA levels (unstimulated cells) on days 1 and 7, but TNF-α mRNA levels increased significantly on day 1 only. Consistent with the increase in superoxide anion production, there were increases in monocyte protein kinase C (PKC) levels on all days of therapy. There was a significant increase in PKCIIβ mRNA only on the first day of treatment. All significant changes in monocyte number and function produced by PIXY321 infusion were reversible, as there were no sustained effects on day 21 (7 days after therapy). These results indicate that the effects of PIXY321 may be mediated through up-regulation of PKC resulting in monocytes primed for increased functional activity in response to an appropriate second stimulus. J. Leukoc. Biol. 53: 640–650; 1993.
Splenic macrophage activation and functions in amyloid enhancing factor-induced secondary amyloidosis Study of phagocytosis, killing, respiratory burst, and MHC class II surface expression
Authors: Reid Caroline , Hébert Lise , Pozzulo Gina , Gervais Francine
Published: January 2018

Secondary amyloidosis is a systemic disease characterized by the extracellular tissue deposition of insoluble fibrillar amyloid A protein. Aberrant metabolism of serum amyloid A protein by reticuloendothelial cells is thought to result in the accumulation of fibrils within the tissue. Treatment of mice with amyloid-enhancing factor (AEF) in conjunction with an inflammatory stimulus (i.e., AgNOs) induced amyloid deposition within 48-72 h. The activation state of a macrophage largely defines its enzymatic capabilities. In the studies reported here, we examined the effect of AEF on spleen macrophage activation using both functional and phenotypic assays. We found that while AEF in the presence or absence of AgNOs has no apparent effect on the ability of spleen and liver macrophages to phagocytose or kill Listeria monocytogenes, it appears to block enhanced respiratory burst function (as measured by O2− production) observed with AgNOs alone. AEF therefore seems capable of inhibiting certain macrophage activation-associated functions while not affecting others. Our activation phenotype studies, using surface Ia expression, reveal that AEF blocks the increase in number of splenic macrophages expressing Ia seen with AgNO3 alone. Treatment with interferon-7 was found to restore decreased Ia expression in animals given AEF + AgNO3 but did not prevent amyloid A fibril deposition. J. Leukoc. Biol. 53: 651–657; 1993.
Comparison of cytosolic calcium shifts, superoxide generation, and lactoferrin secretion in the neutrophils of atopies and nonatopics
Authors: Zweiman Burton , von Allmen Carolyn , Moskovitz Anne , Atkins Paul , Taylor Megan , Korchak Helen
Published: January 2018

We previously found that platelet-activating factor (PAF) stimulated greater lactoferrin secretion by neutrophils of atopies than nonatopics. To help understand underlying mechanisms, we compared agonist- stimulated lactoferrin secretion with another response, superoxide generation, to determine whether there is a global alteration in the reactivity of atopic neutrophils to these stimuli. We also determined Ca2+ mobilization in such neutrophils because such mobilization is a signaling pathway for both superoxide generation and granule content exocytosis. Although PAF again stimulated greater lactoferrin secretion in atopic than in nonatopic neutrophils, the peak superoxide secretion and cytosolic Ca2+ mobilization were not significantly different in atopies and nonatopics. Leukotriene B4-induced superoxide secretion and cytosolic Ca2+ shifts were also similar in atopies and nonatopics. These findings suggest that (1) the enhanced PAF-induced lactoferrin secretion in atopic neutrophils is not a reflection of greater PAF binding or broad-based cell hyperreactivity and (2) a selective signaling pathway in atopic neutrophils may be responsible for the enhanced lactoferrin secretion. These findings may be relevant to in vivo events because we have found increased PAF and lactoferrin release in skin chambers overlying immunoglobulin E-mediated human skin reactions. J. Leukoc. Biol. 53: 727–731; 1993.
Monocyte 1α-hydroxylase regulation: induction by inflammatory cytokines and suppression by dexamethasone and uremia toxin
Authors: Gyetko Margaret R , Hsu Chen Hsaing , Wilkinson Camille C , Patel Sanjeev , Young Eric
Published: January 2018

Abstract: Alveolar macrophages acquire lα-hydroxylase activity in inflammation, and thereby metabolize 25 hydroxyvitamin D3 (25 D3) to the active metabolite, la,25-dihydroxyvitamin D3 (1,25 D3, calcitriol). Cal- citriol is a potent differentiation agent that modulates mononuclear phagocyte activation and effector functions. The mediators that induce macrophage lα-hydroxylase activity are not well delineated. Furthermore, it is unclear whether calcitriol is a product only of terminally differentiated macrophages or whether less mature mononuclear phagocytes can produce it as well. The ability of newly recruited monocytes to produce calcitriol as an autocrine differentiation agent is particularly important in inflammation, as it may substantially expand the functional repertoire of these cells. To assess the effects of cytokines on lα-hydroxylase activity, blood monocytes were cultured in the presence and absence of human recombinant tumor necrosis factor a (TNF-α), interferon-7 (IFN-γ), and interleukins 1 and 2 and then incubated with 25 D3 substrate. The conditioned media were assayed for calcitriol by high-performance liquid chromatography and competitive receptor binding assay. No detectable calcitriol was produced by unstimulated monocytes. However, all the cytokines markedly increased monocyte calcitriol production (range 133-151 pg/mg protein; in all cases P < .001). We then determined whether calcitriol production was suppressed by preincubation with either dexamethasone or the putative uremia toxin guanidinosuccinic acid (GSA). Dexamethasone pretreatment significantly inhibited subsequent cytokine-induced calcitriol production by monocytes, as did GSA (average 69 and 63% of control, respectively).