bluuebunny/crossref_metadata_2025 huggingface dataset:
Lipopolysaccharide modulation of a CD14-like molecule on porcine alveolar macrophages
Authors: Kielian Tammy L , Ross Christopher R , McVey D Scott , Chapes Stephen K , Blecha Frank
Published: January 2018

Cluster of differentiation antigen 14 (GD14) functions as a receptor for lipopolysaccharide (LPS) LPS-binding protein (LBP) complexes. Because LPS has varying effects on CD14 expression in vitro, we evaluated GDI 4 expression in response to LPS with a fully differen tiated macrophage phenotype, the alveolar macrophage. By using flow microfluorometric analysis and a radioim munoassay with an anti-human CD14 monoclonal anti body (My4) that cross-reacts with porcine GD14, we found that macrophages stimulated with LPS for 24 h ex hibited a two- to fivefold increase in CD14-like antigen compared with unstimulated cells. At low concentrations of LPS, up-regulation of the CD14-like antigen was de pendent on scrum; at higher concentrations of LPS, se rum was not required. In the absence of serum a 10-fold higher dose of LPS (10 ng/ml) was required to increase CD14-like expression. In addition, LPS-induced CD14-like up-regulation correlated with secretion of tumor necrosis factor-α, regardless of serum concentration. Blockade with My4 antibody significantly inhibited LPS-induced tumor necrosis factor-α secretion at 1 ng/ml of LPS. However, inhibition decreased as we increased the LPS concentration, suggesting the existence of GDI4-independent pathways of macrophage activation in response to LPS. Alternatively, My4 may have a lower affinity for the porcine CD14 antigen than LPS, which may have only partially blocked the LPS-LBP binding site at high concentrations of LPS. Therefore, these data suggest that LPS activation of porcine alveolar macro phages for 24 h increased CD14-like receptor expression. The degree of CD14-like up-regulation was related to LPS concentration, however, activation did not require the presence of serum at high concentrations of LPS. J. Leukoc. Biol. 57: 581–586; 1995.
Mechanisms of β1 integrin-dependent adherence of granulocytic HL60 to fibronectin
Authors: Bohnsack John F , Chang Jukay , Zhou Xiaoning , Yednock Ted A
Published: January 2018

We investigated the mechanism of β  1 integrin-mediated adherence of stimulated granulocytic HL60 cells to fibronectin using a monoclonal antibody (15/7) that recognizes β  1 integrins only when the recep tors are active for ligand binding. Phorbol myristate ace tate (PMA) stimulated expression of the 15/7 epitope on granulocytic HL60 by nearly fivefold but had an in significant effect on the expression of the epitope on undifferentiated HL60 cells. These results paralleled the effect of PMA on HL60 and granulocytic HL60 adhesion to fibronectin, indicating that activation of β  1 integrins is important for β  1-mediated adherence of granulocytic HL60 cells to fibronectin. Agonists that stimulate α  5  β  1-dependent human polymorphonuclear leukocyte (PMN) adhesion to fibronectin (C5a and PMA) also up-regulated the 15/7 epitope on purified human PMNs. Although PMA rapidly induces increased levels of filamentous actin (F-actin) in granulocytic HL60 cells and a decrease in F-actin levels in undifferentiated HL60 cells, depolymerization of the actin cytoskeleton with cytochalasin B did not affect increased expression of the 15/7 epitope on granulocytic HL60 cells. Cytochalasin B did, however, inhibit granulocytic HL60 adherence to fibronectin by 50%, demonstrating that actin polymeri zation is important for optimal β  1-dependent granulo cytic adherence. J. Leukoc. Biol. 57: 592–599; 1995.
GM-CSF and IL-2 share common control mechanisms in response to costimulatory signals in T cells
Authors: Shannon M Frances , Himes S Roy , Coles Leeanne S
Published: January 2018

Antigen complexed with major histocompatibility complex class I or II molecules on the surface of antigen presenting cells interacts with the T cell receptor (TCR) on the surface of T cells and initiates an activation cascade. So called costimulatory signals, mediated by other cell surface interactions or soluble cytokines produced by antigen presenting cells, are also required for complete T cell activation. High levels of cytokine gene expression in T cells also required both TCR and costimulatory signals. The granulocyte-macrophage colony-stimulating factor requires sequences in the promoter as well as a powerful enhancer located 3kb upstream to respond to TCR-like signals. These promoter and enhancer regions are mainly activated by the transcription factor nuclear factor of activated T cells (NFAT). The activation of NFAT by TCR signals has been well described for interleukin-2 (IL-2) and IL4 gene transcription in T cells. Costimulatory signals, such as activation of the CD28 cell surface molecule on T cells, lead to activation through a distinct region of the granulocyte-macrophage colony-stimulating factor (GM-CSF) promoter. This region is termed the CK-1 or CD28RE and appears to bind specific members of the NF-κB family of transcription factors. Human T leukemia virus type 1 (HTLV-1) infects T cells and can lead to increase GM-CSF expression. We have found that the HTLV-1 transactivator protein, tax, acts as a costimulatory signal for GM-CSF and IL-2 gene transcription, in that it can cooperate with TCR signals to mediate high level gene expression. Tax activates the GM-CSF promoter through the CK-1/CD28RE region and also activates nuclear factor-κB binding to this region. However, other transcription factors or coactivators of NF-κB are required for tax activation but these remain to be identified. The CK-1/CD28RE of GM-CSF shows a high degree of similarity to the IL-2 CD28RE and the IL-3 gene also contains a related region. This observation, together with the fact that both GM-CSF and IL-2 respond to TCR signals via NFAT, implies a high degree of conservation in the regulation of cytokine gene expression in T cells. J. Leukoc. Biol. 57: 767-773; 1995.
Characterization of a non-granule associated pore-forming protein in agranular lymphocytes
Authors: Ortaldo J R , Wiltrout T A , Sayers T J , Yagita H , Winkler-Pickett R T
Published: January 2018

Recently, two populations of small lymphocytes (SL) have exhibited non-major histocompatibility complex (MHC) restricted lysis. Recent studies by numerous laboratories have demonstrated that resting T cells triggered through CD3 and CD28 costimulations can result in immediate, non-MHC restricted killing. Our recent studies with CD3-, CD56+ SL demonstrated that although these cells contained no cytoplasmic granules detected with electron microscopy, they mediated potent NK and ADCC activity. In the present study, we have used a Western blotting technique that allows for the detection and quantitation of total cellular levels of pore-forming protein (PFP). We have found that freshly isolated peripheral non-granulated lymphocytes (both CD3+ and CD3-) contain PFP. In addition, CD3- CD56+ SL contain levels of PFP similar to those of the highly granular CD3- LGL. In search of non-granule PFP, we exploited the rat NK (RNK) cell lines as a source of other potential cytotoxic factors. A membrane associated PFP, based on Western blotting, was isolated from rat RNK cells. Unlike PFP isolated from granules, this PFP was active after culture in Ca2+-containing medium. However, the lytic activity isolated from the non-granule PFP of RNK cells was inhibited by monoclonal antibodies to PFP. Collectively, these studies indicate that PFP is present in small agranular lymphocytes (both NK and T cells) and that it is not stored in large cytoplasmic granules. The implication of our results for the acquisition and activation of lytic ability in NK and T cells will be discussed. J. Leukoc. Biol. 57: 897–903; 1995.
Monocytes cultured in cytokine-defined environments differ from freshly isolated monocytes in their responses to IL-4 and IL-10
Authors: Hart Prue H , Jones Catherine A , Finlay-Jones John J
Published: January 2018

To investigate the regulatory effects of the prototypic Th2 lymphocyte products and potential immunotherapeutic agents interleukin-4 (IL-4) and IL-10 on macrophages differentiated in vitro under different cytokine-defined environments, blood monocytes were incubated for 7 days in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF), macrophage colony-stimulating factor or IL-4. The effect of monocyte culture in the presence or absence of serum was also investigated. Functional responses by 7-day-cultured cells to IL-4, quantified as decreased CD 14 expression and suppression of lipopolysaccharide (LPS)-induced tumor necrosis factor-α (TNF-α) and IL-1β production, and as a positive response, increased CD23 expression, were compared directly with the responses by monocytes from which they were derived. In response to IL-10, decreases in LPS-induced TNF-α and IL-lβ production and reduction in the expression of major histocompatibility complex (MHC) class II antigens were examined. Seven-day cultured monocytes/macrophages showed (1) diminished TNF-α production in response to IL-10 but not EL-4 (2), diminished IL-1β production in response to both IL-4 and IL-10, and compared with fresh monocytes (3), diminished CD14 expression in response to IL-4, and (4) a lesser increase in CD23 expression in response to IL-4. This was the case regardless of the cytokine in the presence of which the cells had been cultured for 7 days. Monocytes cultured for 7 days in GM-CSF expressed increased levels of MHC class II and LPS-induced TNF-α and responded inefficiently to IL-10 for decreased MHC class II. The responses by monocytes cultured for 7 days with GM-CSF resemble the published properties of synovial fluid macrophages from patients with chronic inflammatory arthritis. The study highlights the complexity of monocyte/macrophage responses to the immunoregulatory cytokines IL-4 and IL-10 and concludes that responses to IL-4 and IL-10 by blood monocytes may not be representative of responses by their differentiated or activated counterparts. J. Leukoc. Biol. 57: 909–918; 1995.
The potential role of the macrophage colony-stimulating factor, CSF-1, in inflammatory responses: characterization of macrophage cytokine gene expression
Authors: Evans Robert , Kamdar Sonya J , Fuller Jane A , Krupke Debra M
Published: January 2018

In this report we report that recombinant human monocyte-macrophage colony-stimulating factor-1 (CSF-1) induces resident murine peritoneal cells (PCs) to transcribe several inflammatory cytokine genes, including interleukin (IL)-1α, IL-1β, IL-6, and granulocyte-macrophage CSF in a dose-dependent and time-related manner. Peak mRNA levels were seen between 4 and 6 h. CSF-1 did not modulate the expression of tumor necrosis factor-α mRNA. The serum content of the culture medium appeared to regulate both the extent of CSF-1-induced gene transcription and the adherence properties of the cells. Decreasing the serum concentration significantly reduced CSF-1-induced transcription and was associated with the rapid spreading of the majority of the adherent cells. This reduced sensitivity to CSF-1 was paralleled by a markedly lower levels of c-fms mRNA encoding the CSF-1 receptor. Induced gene transcription was followed by the release of large quantities of IL-6 only. IL-1 activity remained associated with the cells. Neither supernatant nor cell lysate granulocyte-macrophage CSF activity was inducible above the low levels associated with control cultures. Evidence that the mononuclear phagocytes, as opposed to B or T cells, were the targets of CSF-1 was obtained in two ways: (1) PCs from B6 scid/scid and NOD scid/scid mice consisting of 78–86% MAC-1+, F4/80+ cells and few B or T cells, as shown by flow cytometry analysis, released 5- to 10-fold more IL-4 in response to CSF-1 stimulation than B6 PCs, which contained 30% double-positive cells, and (2) pretreatment of B6 PCs with antibodies to the CSF-1 receptor blocked the CSF-1-induced secretion of IL-6. These data suggest that CSF-1 primes noninflammatory mononuclear phagocytes for a role in inflammatory responses but does not provide the necessary signals for either secretion or translation of all cytokines equally. J. Leukoc. Biol. 58: 99–107; 1995.
The role of chemokines in inflammatory joint disease
Authors: Kunkel Steven L , Lukacs Nickolas , Kasama Tsuyoshi , Strieter Robert M
Published: January 2018

Rheumatoid arthritis (RA) is an autoimmune disease characterized by the elicitation and activation of a number of leukocyte populations within both the synovial space and joint tissue. The recruited leukocytes subsequently play an instrumental role in synovial cell proliferation, pannus formation, and bone erosion. Although it is known that leukocytes are important participants in the evolving joint pathology, the mechanism responsible for the successful elicitation of cells to the joint is not clear. A number of studies have identified an association of specific cytokines, including chemokines, with active arthritis, but longitudinal analyses of cytokine expression and the causal role of these mediators have not been defined. Animal models and cell culture systems have proved useful in identifying the expression of various cytokines during the maintenance of chronic joint inflammation. In addition, animal models have provided important information regarding the kinetic production and contribution of specific mediators to the development of experimental arthritis. These studies provide insight into the potential mechanisms for leukocyte involvement in inflammatory joint disease.
Endogenously produced urokinase amplifies tumor necrosis factor-α secretion by THP-1 mononuclear phagocytes
Authors: Sitrin Robert G , Shoilenberger Susan B , Strieter Robert M , Gyetko Margaret R
Published: January 2018

This study examined the effects of endogenous urokinase (uPA) on lipopolysaccharide (LPS)-stimulated tumor necrosis factor α (TNF-α) secretion in THP-1 mononuclear phagocytes. Anti-uPA monoclonal antibody (mAb) suppressed LPS-driven TNF-α secretion by 61.6 ± 5.9% (P < .001), and PAI-1, a uPA inhibitor, suppressed it to 53.1 ± 8.2% of the control value (P < .001). Up-regulation of TNF-α mRNA was suppressed in parallel with secreted TNF-α protein. TNF-α secretion was unaffected by depleting plasminogen or by aprotinin, a plasmin inhibitor. When endogenous uPA was displaced from the cell, exogenous high-molecular-weight (intact) uPA augmented LPS-driven TNF-α secretion. By contrast, a uPA fragment containing the catalytic domain was inhibitory, and the uPA receptor-binding domain had no effect. We conclude that endogenous uPA amplifies TNF-α neosynthesis of UPS-stimulated THP-1 mononuclear phagocytes. The effect requires intact uPA and is independent of plasmin activity. This represents a novel mechanism by which a mononuclear phagocyte–derived protease contributes to generating proinflammatory signals.
A monoclonal antibody to human macrophage gangliosides inhibits macrophage migration
Authors: Berenson Charles S , Patterson Melissa A , Pattoli Mark A , Murphy Timothy F
Published: January 2018

Gangliosides have diverse immunoregulatory properties. The gangliosides endogenous to macrophages may have immunoregulatory properties that distinguish them from other gangliosides. Gangliosides have been indirectly implicated in macrophage migration as putative cell surface receptors for migration inhibitory factor (MIF). In this study, a monoclonal antibody to human macrophage gangliosides (antibody 25F4) was developed and characterized. This is the first report of the development of monoclonal antibodies to gangliosides of macrophages of any species. Thin-layer chromatographic immunostaining indicated that antibody 25F4 recognized major gangliosides of human macrophages but did not recognize those previously identified as containing fucose. Immunofluorescent surface labeling of viable human macrophages indicated that antibody 25F4 recognized a surface-accessible epitope, present on all cells, and that this was abolished with lipid depletion of macrophage membranes. This epitope was not present on several human nonmacrophage cells. Finally, human macrophages pretreated with antibody 25F4 demonstrated striking inhibition of migration in an agarose droplet assay, whereas an irrelevant monoclonal antibody or monoclonal antibodies to nonganglioside surface epitopes of human macrophages had no effect on migration. Migration inhibition occurred even though antibody 25F4 was removed from the extracellular milieu and was not due to formation of cellular aggregates. These studies support a role for human macrophage gangliosides in macrophage migration.
In vitro culture of murine peritoneal and alveolar macrophages modulates phagocytosis of Pseudomonas aeruginosa and glucose transport
Authors: Everett Karin D E , Barghouthi Sameer , Speert David P
Published: January 2018

Phagocytosis by murine peritoneal macrophages (PMφ) of unopsonized Pseudomonas aeruginosa is a novel, glucose-dependent process occurring in concert with glucose or mannose transport via the GLUT1 facilitative transporter. The mechanism by which this transport triggers phagocytosis is not understood. The purpose of these investigations was to improve our understanding of this mechanism by delivery of an alternative sugar (fructose) to PMφ and to murine alveolar macrophages (AMφ). Fructose-cultured PMφ developed fructose-dependent phagocytosis of P. aeruginosa with increased glucose-dependent phagocytosis, GLUT1 expression, and [14C]glucose transport. Freshly explanted AMφ, which were unable to transport [41C]ghicose or to ingest P. aeruginosa acquired the ability to transport glucose and to phagocytose P. aeruginosa with culture in either glucose or fructose. Both fructose- and glucose-cultured AMφ remained viable but incapable of measurable fructose transport or fructose-dependent phagocytosis. These studies suggest that an intracellular metabolite of fructose, glucose, and mannose is involved in triggering macrophage phagocytosis of P. aeruginosa. We demonstrate that delivery of appropriate substrates can substantially improve AMφ phagocytic function and may therefore possibly improve pulmonary host defense against P. aeruginosa.
Differential regulation of Ia expression and antigen presentation by listeriolysin-producing versus non-producing strains of Listeria monocytogenes
Authors: Vazquez M A , Sicher S C , Proctor M L , Crowley J C , Lu C Y
Published: January 2018

listeria monocytogenes is an intracellular bacterial pathogen. A single gene product, listeriolysin (LLO), is critical for the induction of protective immunity. We now show that listeria that produce functional LLO augment Ia expression by macrophages and are better presented to a Th1, CD4+ anti-listeria T cell line. We used two genetically engineered strains of listeria which differed only in their ability (Ly+) or inability (Ly-) to produce functional LLO. Ia-negative murine macrophages ingested either Ly+ or Ly-, and then were stimulated by interferon-γ(IFN-γ). Increasing numbers of live Ly+, but not Ly-, augmented IFN-γ-induced Ia expression. Ly+ by itself did not induce Ia expression. Heat-killed Ly+ and Ly- did not augment IFN-γ-induced Ia expression. The abundance of Ia on the macrophage cell surface is one major determinant of antigen presentation to CD4+ T cells. Consistent with their ability to augment Ia expression, Ly+ were better presented than Ly- to a CD4+, Th1, anti-listeria T cell line. When macrophages and T cells were from different inbred mouse strains, antigen presentation required identity at the Class II region of the MHC gene complex. This indicated that antigen presentation occurred via Ia molecules. The increased ability of macrophages to present Ly+ is a product of the macrophage-listeria interaction, not a property of the T cell line 86. If Ia-negative macrophages ingested listeria and were then stimulated by IFN-γ, Ly+ was presented more efficiently than Ly-. On the other hand, if Ia-positive macrophages ingested Listeria, then Ly+ and Ly- were presented equally well to T cells. Altogether our data is consistent with the hypothesis that macrophages interact differently with Ly+, and that this contributes to the ability of only live Ly+-to induce protective immunity.
Development and characterization of a novel monoclonal antibody (mNI-11) that induces cell adhesion of the LPS-stimulated human monocyte-like cell line U937
Authors: Ikewaki Nobunao , Inoko Hidetoshi
Published: January 2018

A monoclonal immunoglobulin G1 (IgG1) antibody (mAb), designated mNI-11, was produced by immunizing mice with the lipopolysaccharide (LPS)-stimulated monocyte-like cell line U937. The reactivity of mNI-11 was tested by the indirect immunofluorescence method. The antigen defined by mNI-11 was found to be expressed on U937 cells, LPS-stimulated U937 cells, normal CD14+ cells (monocytes/macrophages), and human umbilical vein endothelial cells (HUVECs). Expression of the antigen defined by mNI-11 on HUVECs slightly increased in response to exposure to tumor necrosis factor-α (TNF-α) and phorbol myristate acetate (PMA). When the reactivity of mNI-11 and mAbs binding human differentiation antigens such as CD11a, CD11b, CD11c, CD14, CD16, CD18, CD23, CD28, CD29, CD31, CD43, CD44, CD45RA, CD49d, CD50, CD54, CD58, CD80, CD102, CD106, HLA-class I, or HLA-class II antigen was compared, no mNI-11 reactivity resembling that of these mAbs was found. mNI-11 markedly induced homotypic cell aggregation of U937 cells when they were stimulated with LPS. The mNI-11-induced aggregation of LPS-stimulated U937 cells, referred to as LPS-U937 cells, required neither Fc receptor engagement nor cross-linking of the antigen defined by mNI-11 because aggregation was induced by both F(ab’)2 fragments and monovalent F(ab/) fragments of mNI-11. The mNI-11-induced aggregation was blocked by the addition of ethylenediaminetetraacetate, and also when incubated at 4°. mAbs to CD11a/CD18 (lymphocyte-function associated antigen-1; LFA-1) and CD54 (intercellular adhesion molecule-1; ICAM-1) completely blocked the LPS-U937 cell aggregation induced by mNI-11. The IPS-U937 cell aggregation induced by mNI-11 was partially but not completely blocked by the protein kinase C inhibitors sphingosine and H-7, and was completely blocked by the protein-tyrosine kinase inhibitor genistein. Interestingly, mNI-11 markedly promoted LPS-U937 cell adhesion to HUVECs. The mNI-11-induced LPS-U937 cell adhesion to HUVECs was not reduced in the presence of LFA-1 (CD11a/CD18) or ICAM-1 (CD54) mAbs. On the other hand, IPS-U937 cells, whether treated with mNI-11 or not, sufficiently adhered to the extracellular matrix protein fibronectin, but not to laminin or collagen type I. However, mNI-11 did not markedly promote IPS-U937 cell adhesion to fibronectin. Adhesion of IPS-U937 cells treated with mNI-11 to fibronectin was completely blocked by CD29 (β chain of very late antigens) mAb. The surface antigen recognized by mNI-11 had a molecular size of ~97 kDa under non-reducing conditions and ~117 kDa under reducing conditions, as determined by immunoblotting analysis. We found that mNI-11 recognizes an adhesion-associated molecule distinct from any previously reported in terms of its pattern of cellular distribution and molecular weight, and also found that mNI-11 has activity which induces cell adhesion/aggregation of U937 cells when stimulated with LPS.
Novel actions of aspirin and sodium salicylate: discordant effects on nitric oxide synthesis and induction of nitric oxide synthase mRNA in a murine macrophage cell line
Authors: Kepka-Lenhart Diane , Chen Li-Chun , Morris Sidney M
Published: January 2018

Aspirin and sodium salicylate each inhibit to a similar extent the production of nitric oxide (NO) in the RAW 264.7 murine macrophage cell line following stimulation by either lipopolysaccharide (LPS) or interferon-γ(IFN-γ). The similar potencies of aspirin and sodium salicylate indicate that acetylation of cellular macromolecules is not essential for the observed effects. The failure of added prostaglandin E2 to overcome the effects of aspirin or sodium salicylate indicates that these effects are not simply the result of inhibition of prostaglandin synthesis. The inhibition of NO production occurs irrespective of the effect of these agents on induction of nitric oxide synthase (iNOS) mRNA by LPS or IFN-γ. Aspirin and sodium salicylate inhibit iNOS mRNA induction in LPS-stimulated cells but enhance iNOS mRNA induction in IFN-γ-stimulated cells. In contrast, these agents consistently inhibit induction of argininosuccinate synthetase mRNA in both LPS- and IFN-γ-stimulated cells. Concentrations of aspirin in the 3–10 mM range inhibit induced NO production and expression of iNOS protein without inhibiting induction of iNOS mRNA. Discordances between effects on NO synthesis and induction of iNOS mRNA indicate that aspirin and sodium salicylate have multiple sites of action in their effects on pathways that are involved in the production of NO by stimulated RAW 264.7 cells.
T cell rescue of monocytes from apoptosis: role of the CD40-CD40L interaction and requirement for CD40-mediated induction of protein tyrosine kinase activity
Authors: Suttles Jill , Evans Mike , Miller Robert W , Poe Jonathan C , Stout Robert D , Wahl Larry M
Published: January 2018

Circulating monocytes have a limited life span and will undergo apoptosis in the absence of specific stimuli. Recent studies have demonstrated that monocytes can be rescued from apoptosis via lipopolysaccharide (LPS) activation or stimulation with interleukin-1 or tumor necrosis factor-α. Based on previous studies from our laboratory, we hypothesized that, in nonseptic (e.g., autoimmune) inflammation, the presence of activated T cells may enhance monocyte longevity through T cell contact-dependent signaling. Plasma membranes prepared from 6 h activated (TmA) and resting (TmR) purified CD4+ T cells were added to resting elutriation-purified monocytes cultured in serum-free medium. Cells were assayed for degree of apoptosis occurring over a 72-h incubation using both agarose gel electrophoresis and flow cytometry. The addition of TmA (but not TmR) was capable of blocking monocyte apoptosis and the ability of TmA to rescue monocytes was abrogated by the addition of anti-CD40L antibodies. Rescue of monocytes from apoptosis could also be mediated by direct cross-linking of monocyte CD40. Inhibitors of tyrosine kinase activity blocked both TmA and anti-CD40-mediated rescue of monocytes from apoptosis, suggesting a primary role of a tyrosine kinase signaling pathway in the events controlling monocyte longevity.
MIP-1α contributes to the anticryptococcal delayed-type hypersensitivity reaction and protection against Cryptococcus neoformans
Authors: Doyle Hester A , Murphy Juneann W
Published: January 2018

Leukocyte infiltration into infected tissues is essential for the clearance of microorganisms. In animals with a cell-mediated immune (CMI) response to the infectious agent, as opposed to naive animals, leukocyte migration is greatly enhanced into sites of the organism or antigen. The role of the chemotactic cytokine or chemokine, macrophage inflammatory protein-1α (MIP-1α), in the expression phase of the CMI response and in protection against Cryptococcus neoformans was assessed. With the use of a gelatin sponge model in mice as a means of detecting an anti-cryptococcal delayed-type hypersensitivity (DTH) reaction, we found that MIP-1 α levels in fluids from cryptococcal antigen (CneF)-injected sponges in immunized mice (DTH-reactive sponges) were significantly increased over levels of MIP-1α in fluids from saline-injected control sponges at 12 and 24–30 h after injection. MIP-1α levels peaked before increases in neutrophils and lymphocytes in the DTH-reactive sponges, suggesting that MIP-1α was responsible, at least in part, for attracting these leukocyte types. Immunized mice treated with neutralizing antibody to MIP-lα before sponge injection with CneF had reduced numbers of neutrophils and lymphocytes in the DTH-reactive sponges and showed reduced clearance of C. neoformans from the lungs, spleens, livers, and brains when compared with controls. Furthermore, injection of rmMIP-1α into sponges in naive mice resulted in an increase in the influx of neutrophils and lymphocytes into the sponges compared with saline-injected sponges. Together our findings provide solid evidence that MIP-1α is a component of the anticryptococcal DTH reaction. In addition, MIP-1α influences neutrophil influx and attracts lymphocytes into the DTH reaction site. Finally, we showed that MIP-1α plays a role in protection against C. neoformans.
Monocytes/macrophages rather than PMN are involved in early cartilage degradation in cationic immune complex arthritis in mice
Authors: van Lent P L E M , Blom A , Holthuysen A E M , Jacobs C W M , van De Putte L B A , van Den Berg W B
Published: January 2018

We investigated the involvement of polymorphonuclear granulocytes (PMN) and monocytes in cartilage degradation in immune complex-mediated arthritis (ICA). ICA induced with lysozyme-anti-lysozyme in the murine knee joint is characterized by a major influx of PMNs followed by monocytes and marked cartilage proteoglycan (PG) depletion develops within 2 days. Around 60% of 35S-prelabeled PG is lost at day 2. Influx of cells was manipulated using interleukin-1 (IL-1) receptor antagonist (IL-lra) or anti-bodies to adhesion molecules. Cellular infiltrate was analyzed on hematoxylin-stained joint sections. Early systemic treatment with IL-lra highly reduced PMN influx, whereas monocyte influx was hardly diminished. PG loss was not significantly reduced, declining from 62% in controls to 47% in IL-lra-treated mice. Total blockade of cell influx was found after intravenous treatment with monoclonal antibodies 5C6 (anti-CD11b/CD18:anti-CR3) or NIMP.R14 (25–30 kDa protein mainly present on PMN) and PG loss was reduced to 5–10%. A similar reduction was observed after prior depletion of circulating PMNs with total body irradiation. Because amounts of IL-1 produced in leukopenic and control arthritic joints are comparable, this suggests that IL-1 is only marginally involved in PG loss in the first phase of ICA. This study indicates that monocytes rather than PMN might be involved in PG loss in this form of arthritis, either directly or by local activation of synovial layer cells of the joint.
Monocyte chemotactic protein-4: tissue-specific expression and signaling through CC chemokine receptor-2
Authors: Godiska Ronald , Chantry David , Raport Carol J , Schweickart Vicki L , Le Trong Hai , Gray Patrick W
Published: January 2018

Chemokines constitute a family of low-molecular-weight proteins that attract or activate a variety of cell types, including leukocytes, endothelial cells, and fibroblasts. An electronic search of the GenBank Expressed Sequence Tags database uncovered a partial cDNA sequence with homology to the chemokine monocyte chemotactic protein-1 (MCP-1). Isolation of the full-length clone revealed that it encodes the chemokine MCP-4, an eosinophil chemoattractant recently described by Uguccioni et al. [J. Exp. Med. 183, 2379–2384]. Recombinant MCP-4 was expressed in mammalian cells and purified by heparin-Sepharose chromatography. Sequencing the amino terminus of this protein corroborated the reported sequence of recombinant MCP-4 produced in insect cells. As shown by calcium flux assays, MCP-4 activated the cloned G protein-coupled receptor CCR-2, which also recognizes MCP-1 and MCP-3. Northern hybridization indicated that MCP-4 is constitutively expressed at high levels in the small intestine, colon, and lung. This expression profile is consistent with its role as a chemoattractant for eosinophils, which can be rapidly mobilized to the lung or intestine in response to invading pathogens. In marked contrast to MCP-1, MCP-4 was not induced in cell lines treated with pro-inflammatory stimuli such as lipopolysaccharide or tumor necrosis factor α.
Inflammatory damage following first-generation replication-defective adenovirus controlled by anti-LFA-1
Authors: Guérette B , Moisset P-A , Huard C , Tardif F , Gravel C , Tremblay J P
Published: January 2018

First-generation replication-defective adenoviruses have been reported to lead to transient reporter gene expression due to a specific immune reaction involving T and B lymphocytes. Some recent reports have also demonstrated the presence of a nonspecific inflammatory reaction involving macrophages and neutrophils after both intramuscular injections and viral vectors transduction. To further investigate this nonspecific inflammatory reaction, ΔE1/E3a adenoviruses were injected intramuscularly in immunocompetent mice. Some of these mice were treated with anti-LFA-1. The adenovirus-injected muscles showed abundant CD4+, CD8+, LFA-1+, and Mac-1+ cell infiltration 3 days after the ΔE1/E3a injection. The anti-LFA-1 monoclonal antibody was able to block the nonspecific inflammatory damage due mostly to neutrophils and macrophages. The anti-LFA-1 did not produce this effect by reducing the muscle infiltration by LFA-1+ cells. It may instead have blocked the direct interaction between LFA-1 and ICAM-1 thus preventing the damage produced by the respiratory burst of neutrophils. Blocking the resulting damage of this inflammatory reaction with anti-LFA-1 in animals also treated with FK506, a powerful immunosuppressant for gene therapy, largely increased the long-term transgene expression compared with mice only treated with FK506.
Inhibitory effect of 3,4-dichloropropionaniline on cytokine production by macrophages is associated with LPS-mediated signal transduction
Authors: Xie Y C , Schafer R , Barnett J B
Published: January 2018

Our previous studies demonstrated that both in vivo and in vitro 3,4-dichloropropionanilide (propanil) exposure inhibited interleukin-6 (IL-6) and tumor necrosis factor (TNF) production by adherent thioglycollate-elicited peritoneal cells (macrophages) after lipopolysaccharide (LPS) stimulation. In this study, we report that IL-6 and TNF-α message is reduced by propanil in a concentration-dependent pattern, yet the stability of cytokine mRNA is not affected. In addition, exposure of macrophages to propanil after a relatively short period of LPS stimulation significantly reduced the production of IL-6 and TNF. Determination of the intracellular Ca2+ levels demonstrates that LPS-induced Ca2+ release is abrogated in propaniltreated macrophages. However, the binding of LPS to macrophages is not affected. Measurement of inositol 1,4,5-triphosphate (IP3) demonstrates that propanil significantly increases the level and the duration of IP3 in macrophages. These results suggest that the inhibitory effect of propanil on macrophage cytokine production is associated with the early stages of LPS-mediated signal transduction in macrophages.
Modulation of human neutrophil inflammatory responses by nitric oxide: studies in unprimed and LPS-primed cells
Authors: Nath Jayasree , Powledge Annette
Published: January 2018

Because nitric oxide (NO) can act both as a regulatory and as a toxic molecule, we have studied N-formyl-methionyl-leucyl-phenylalanine (fMLF) -stimulated responses of human neutrophils (PMNs) during various conditions of NO modulation in unprimed and bacterial lipopolysaccharide (LPS) -primed cells. Effects of various NO modulators were assessed on stimulated superoxide (O2-) generation, granule exocytosis, homotypic aggregation, and rises in intracellular free Ca2+ ([Ca2+]i). Significant differences in the effects of various NO modulators on inflammatory responses of PMNs kept in stirred suspension versus those kept under static and/or adherent conditions, were observed. L-arginine, the physiological substrate for NO synthase (NOS), and NG-nitro-L-arginine methyl ester, an inhibitor of NOS, both caused a 40-50% inhibition of LPS-induced priming of O2- generation in PMNs in stirred suspension, but not in LPS-primed PMNs under static or adherent conditions. The NO donors, sodium nitroprusside and S-nitroso-N-acetylpenicillamine, completely abrogated the LPS-induced priming of 02- generation in PMNs in suspension, while causing only a 40-50% inhibition in PMNs under static or adherent conditions. The Ca2+ ionophore, A23187, prevented the LPS-induced priming of 02-generation without affecting 02- generation in unprimed PMNs. LPS priming of PMNs induced about a twofold increase in fMLF-stimulated homotypic aggregation, exocytosis of secondary granules, and rises in [Ca2+]i. In related studies, we also provide definitive evidence for enzymatic formation of NO in human PMNs and demonstrate a significant decrease in NO levels in LPS-primed PMNs. Taken together, these findings indicate that NO modulates PMN inflammatory responses and plays a protective role in priming and activation processes of inflammatory PMNs.
Monocyte chemoattractant protein-2 is a potent agonist of CCR2B
Authors: Moore Una M , Kaplow June M , Pleass Robert D , Castro Sandra W , Naik Kuni , Lynch Carolyn N , Daly Steven , Roach Alan G , Jaye Michael , Williams Robert J
Published: January 2018

The binding and functional activity of the CC chemokines monocyte chemoattractant protein-1 (MCP-1), MCP-2, and MCP-3 have been characterized using Chinese hamster ovary DXB-11 cells transfected with the chemokine receptor CCR2B. Receptor binding studies demonstrated that 125I-labeled MCP-1 bound to a single class of high-affinity receptors with a Kd of 0.14 (0.070.32) nM. In competition studies MCP-1, MCP-2, and MCP-3 completely inhibited 125I-labeled MCP-1 binding with Ki values of 0.3 (0.16-0.46), 8.8 (3.4-26), and 12.2 (0.6-22) nM, respectively. In calcium mobilization studies, MCP-1 and MCP-3 induced robust elevations in intracellular calcium concentrations, whereas MCP-2 was only weakly active. In contrast, using changes in extracellular acidification rate as a functional readout, all three chemokines were identified as potent agonists of CCR2B. These data demonstrate that MCP-2, in addition to MCP-1 and MCP-3, is a potent agonist of CCR2B and furthermore that MCP-2 activates either different or a subset of the signaling pathways activated by MCP-1 and MCP-3.
IL-4 secretion and histamine release by human basophils are differentially regulated by protein kinase C activation
Authors: Schroeder John T , Howard Brian P , Jenkens M Kathleen , Kagey-Sobotka Anne , Lichtenstein Lawrence M , MacGlashan Donald W
Published: January 2018

The role of protein kinase C (PKC) activation was investigated in the secretion of interleukin-4 (IL-4) protein by human basophils. Phorbol myristate acetate (PMA) induced little to no detectable IL-4 protein in culture supernatants, despite being a potent secretagogue of histamine release by basophils. In fact, the secretion of IL-4 by basophils stimulated with ionomycin alone was down-regulated (30-70%) with the simultaneous addition of PMA. In peripheral blood lymphocytes (PBL), however, the combination of ionomycin and PMA were highly synergistic, resulting in maximum IL-4 release but at a slower rate. PKC inhibitors reversed these effects on IL-4 secretion. In sharp contrast to its inhibitory effect on IL-4 protein secretion, PMA did not block the accumulation of IL-4 mRNA in basophils activated by ionomycin. These data suggest that there are marked differences in the regulatory processes for IL-4 transcription, translation, or secretion between basophils and lymphocytes.
Role of complement in in vitro and in vivo lung inflammatory reactions
Authors: Czermak Boris J , Lentsch Alex B , Bless Nicolas M , Schmal Hagen , Friedl Hans Peter , Ward Peter A
Published: January 2018

Complement is one of the integral buttresses of the inflammatory response. In addition to host defense activities, proinflammatory properties of several complement components are described. This overview elucidates the role of complement in inflammatory reactions in vitro and in vivo, focusing on the complement activation products, C5a, and the membrane attack complex, C5b-9. Using several approaches, the impact of these complement components in mechanisms relevant to neutrophil recruitment is emphasized. In addition, the participation of complement in endothelial superoxide generation and its essential requirement for full expression of lung injury is demonstrated, as are the involved intracellular signal transduction pathways. Understanding the mechanisms of complement-induced proinflammatory effects may provide a basis for future therapeutic blockade of complement and/or its activation products. J. Leukoc. Biol. 64: 40–48; 1998.
Complex high affinity interactions occur between MHCI and superantigens
Authors: Chapes Stephen K , Herpich Angela R
Published: January 2018

Staphylococcal enterotoxins A and C1 (SEA or SEC1) bound to major histocompatibility-I (MHCI) molecules with high affinity (binding constants ranging from 1.1 μM to 79 nM). SEA and SEC1 directly bound MHCI molecules that had been captured by monoclonal antibodies specific for H-2Kk, H-2Dk, or both. In addition, MHCI-specific antibodies inhibited the binding of SEC1 to LM929 cells and SEA competitively inhibited SEC1 binding; indicating that the superantigens bound to MHCI on the cell surface. The affinity and number of superantigen binding sites differed depending on whether MHCI was expressed in the membrane of LM929 cells or whether it was captured. These data support the hypothesis that MHCI molecules can serve as superantigen receptors. J. Leukoc. Biol. 64:587–594; 1998.
Oxidized LDL modulates activation of NFκB in mononuclear phagocytes by altering the degradation of IκBs
Authors: Hamilton Thomas A , Major Jennifer A , Armstrong David , Tebo Julie M
Published: January 2018

Oxidized low density lipoprotein (oxLDL) is known to alter the expression of inflammatory gene products in mononuclear phagocytes. The mechanisms involved in this effect were studied by examining the activation of nuclear factor κB (NFκB), a transcription factor known to be important in controlling the expression of such genes. Pretreatment of peritoneal macrophages with oxLDL modulated the activation of NFκB in response to either lipopolysaccharide (LPS) or the combination of interferon-γ (IFN-γ) and interleukin-2 (IL-2). In macrophages pretreated with oxLDL the nuclear translocation of Rel family members (RelA and c-Rel) is delayed (LPS) or markedly diminished (IFN-γ/IL-2) and results in delayed or reduced appearance of κB binding activity within the nucleus. These changes in NFκB activation result from alterations in the stimulus-dependent degradation of IκBα and IκBβ. The effects of oxLDL on NFκB activation depend both on the degree of LDL oxidation (most potent with extensive oxidation) and on the time of exposure of the cells to the lipoprotein preparation (a minimal exposure of 6 h is required before inhibitory effects are observed). The modulation of IκB/NFκB function in cells exposed to oxLDL appears to be responsible for previously reported effects of oxLDL on chemoattractant cytokine gene expression where both inhibition and delay of such stimulus-dependent events has been observed. J. Leukoc. Biol. 64: 667–674; 1998.
Evaluation of the expression of human CAP18 gene during neutrophil maturation in the bone marrow
Authors: Nagaoka Isao , Hirata Michimasa , Sugimoto Koichi , Tsutsumi-Ishii Yuko , Someya Akimasa , Saionji Katsu , Igari Jun
Published: January 2018

To understand the gene expression of CAP18 (18-kDa cationic antibacterial protein), a member of cathelicidins, we evaluated mRNA and protein expression of CAP18 using human bone marrow cells and mature neutrophils. Northern blot analysis revealed that CAP18 mRNA was expressed more abundantly in bone marrow cells than mature neutrophils, whereas Western blot analysis indicated that CAP18 protein was more abundant in mature neutrophils than bone marrow cells. Consistent with this, in situ hybridization using bone marrow cells demonstrated that the expression of CAP18 mRNA was neutrophil lineage-specific and was observed primarily in myelocytes (>95%) with limited expression in more immature cells (promyelocytes) and mature cells (metamyelocytes, band cells, and segmented neutrophils). Furthermore, immunohistochemical study indicated that, coincident with the increase of CAP18 mRNA levels, CAP18-positive cells increased markedly at myelocyte stage, and the increased levels remained almost constant (>95%) in metamyelocytes, band cells, and segmented neutrophils, although the mRNA levels were remarkably reduced in these cells. Together these observations indicate that CAP18 gene transcription likely occurs lineage- and stage-specifically at the myelocyte stage of neutrophil maturation in the bone marrow and results in the synthesis and cytoplasmic accumulation of CAP18, which is present in the subsequent stages of neutrophil maturation. J. Leukoc. Biol. 64:845–852; 1998.
Cytolysis of human dendritic cells by autologous lymphokine-activated killer cells: participation of both T cells and NK cells in the killing
Authors: Parajuli Prahlad , Nishioka Yasuhiko , Nishimura Naoki , Singh Sukh Mahendra , Hanibuchi Masaki , Nokihara Hiroshi , Yanagawa Hiroaki , Sone Saburo
Published: January 2018

Dendritic cells (DC) play a key role in the initiation of immune response by stimulating the naive T cells. The fate of DC after the initiation of immune response is not clearly understood. Although there are few reports implicating natural killer (NK) cells in the elimination of DC, killing of DC by LAK cells, and specifically by T cells, has not been studied. In this study, we observed that DC, generated from monocytes, in vitro in the presence of granulocyte-macrophage colony-stimulating factor, interleukin-4 (IL-4), and tumor necrosis factor α were susceptible to cytolysis by lymphokine-activated killer (LAK) cells induced in the presence of IL-2 and IL-15 but not IL-12 alone. However, LAK cells induced by a combination of IL-12 and suboptimal dose of IL-2 were cytotoxic to DC. When purified lymphocytes were activated with IL-2, the CD8+/CD57− fraction (T-LAK), but not the CD8−/CD57+ fraction (NK-LAK) was cytotoxic to autologous DC. However, when unseparated peripheral blood mononuclear cells were used to generate LAK cells, both T-LAK and NK-LAK fractions showed equal cytotoxicity against autologous DC. Monoclonal antibodies against CD54, CD11a, and CD18 significantly inhibited the cytolysis, indicating that the killing involves the engagement of CD54 with its ligands. J. Leukoc. Biol. 65: 764–770; 1999.
Ubiquitin-proteasome system is involved in induction of LFA-1/ICAM-1-dependent adhesion of HL-60 cells
Authors: Katagiri Koko , Yokosawa Hideyoshi , Kinashi Tatsuo , Kawashima Seiichi , Irie Shinkichi , Tanaka Keiji , Katagiri Takuya
Published: January 2018

Membrane-permeable proteasome inhibitors, lactacystin (LC) and N-acetyl-Leu-Leu-norleucinal (ALLN), but not calpain inhibitor Z-Leuleucinal (ZLL), prevented LFA-1/ICAM-1-dependent cellular adhesion of TPA-stimulated HL-60 cells. These proteasome inhibitors affected neither the induction of monocytic differentiation nor the accompanying protein-tyrosine phosphorylation. They suppressed the increase in the avidity of LFA-1 to ICAM-1 without changing the expression of these molecules. Immunoblotting using monoclonal antibody FK-1, which reacts specifically with polyubiquitinated proteins, demonstrated that the proteasome inhibitors caused the drastic accumulation of the polyubiquitinated proteins in the membrane fraction of TPA-treated HL-60 cells. This indicates that accompanying activation of LFA-1, TPA induces the polyubiquitination of the membrane proteins, which are rapidly degraded by proteasomes. These data taken together show that proteolysis mediated by the ubiquitin-proteasome system is a prerequisite for the induction of LFA-1-dependent adhesion of HL-60 cells. J. Leukoc. Biol. 65: 778–785; 1999.