Platelet-activating factor secreted by DDAVP-treated monocytes mediates von willebrand factor release from endothelial cells

We have previously shown that although DDAVP (1-deamino-8-D-arginine vasopressin), a synthetic analogue of the natural hormone arginine vasopressin, does not directly promote release of vWf from human umbilical vein endothelial cells (ECs), enhanced release does occur when ECs were exposed to either monocytes or to supernatants recovered from DDAVP-treated monocytes. In the present study, we have found that exposure of monocytes to DDAVP did not increase secretion of interleukins (IL)-1β, IL-6, IL-8, tumor necrosis factor (TNF-α), growth factors G-CSF (granulocyte-), GM-CSF (granulocyte, monocyte-colony stimulating factor), prostaglandins (PG) E₂, PGF_2α, or PGI₂ or purine nucleotides such as ATP and ADP. However, increased levels of platelet-activating factor (PAF) were secreted by DDAVP-treated monocytes in a time- and dose-dependent manner that positively correlated with the enhancement in vWf release from ECs. Moreover, this effect could also be elicited when lipid extracts of these supernatants or purified PAF were added directly to ECs. This response could be inhibited with (±)-trans-2,5-Bis(3,4,5-trimethoxyphenyl)-1,3-dioxolane, a specific PAF receptor antagonist, when the ECs were exposed to supernatants from DDAVP-treated monocytes or to pure PAF. The present data indicate that enhanced secretion of PAF from monocytes is one mechanism whereby DDAVP can provoke release of vWf from ECs. © 1993 Wiley-Liss, Inc.     

Stimulation by human interleukin 1 of cartilage breakdown and production of collagenase and proteoglycanase by human chondrocytes but not by human osteoblasts in vitro

Human articular chondrocytes in culture synthesise collagenase and neutral proteoglycanase in response to addition of a 12-17 kDa protein produced by cultured human monocytes. This factor copurifies with interleukin 1, as assessed by lymphocyte activating factor activity, on gel filtration chromatography and isoelectric focusing. The interleukin 1 and chondrocyte-stimulating activities are destroyed by pretreatment of the material with phenylglyoxal. The same materials also promote the release of glycosaminoglycans from cultures of intact bovine nasal cartilage. The proteoglycanase activity release from chondrocytes appears to be a metalloproteinase because it is inhibited by EDTA and not by phenylmethylsulphonyl fluoride (PMSF), and because detection of its activity is dependent on the presence of 4-aminophenylmercuric acetate. Human osteoblast-like cells do not respond to this factor by increased proteinase production, but are stimulated to produce prostaglandins. These results suggest that interleukin 1 has activities upon non-immune cells which promote the degradation of connective tissue matrices. Human osteoblasts do not synthesise neutral collagen- and proteoglycan-degrading enzymes and thus are unlikely to be directly responsible for the matrix degradation which occurs during bone resorption.     

IL-1 receptor antagonist release is regulated differently in human alveolar macrophages than in monocytes.

These studies compared the release of interleukin-1 receptor antagonist (IL-1 RA) from alveolar macrophages and peripheral blood monocytes. The cells were cultured in medium containing various amounts of heat-inactivated fetal calf serum (FCS), granulocyte-macrophage colony-stimulating factor (GM-CSF), lipopolysaccharide (LPS), and immunoglobulin G (IgG). In serum-free medium alone, IL-1 RA release was similar from macrophages and monocytes. Increasing FCS concentration caused a significant upregulation of IL-1 RA release in macrophages but not in monocytes. GM-CSF caused a small increase in both cell types. LPS caused downregulation of IL-1 RA release from monocytes but not from macrophages. IgG did not affect IL-1 RA release in either cell group. These studies demonstrate that regulation of IL-1 RA release is different in monocytes and macrophages.     

Prolactin increases tumor necrosis factor alpha expression in peripheral CD14 monocytes of patients with rheumatoid arthritis

Tumor necrosis factor (TNF)-α is one of the major proinflammatory mediators of rheumatic arthritis (RA); the regulatory factors for TNF-α release is not fully understood. This study aims to investigate the role of prolactin receptor (PRLR) activation in regulating the expression and release of TNF-α from CD14⁺ monocytes. The results showed that the expression of PRLR was detectable in CD14⁺ monocytes of healthy subjects, which was markedly increased in RA patients. Exposure to PRL in the culture increased the expression and release of TNF-α from CD14⁺ monocytes, which was abolished by the PRLR gene silencing or blocking the mitogen activated protein (MAPK) pathway. We conclude that exposure to PRL increases TNF-α release from CD14⁺ monocytes of RA patients, which can be abolished by PRLR gene silencing or treating with MAPK inhibitor.     

Effects of the new adiponectin paralogous protein CTRP-3 and of LPS on cytokine release from monocytes of patients with type 2 diabetes mellitus

Aims/Hypothesis: It was the aim to investigate the hypothesis that the new C1q/TNF-family member CTRP-3 (C1q/TNF-related protein-3) acts anti-inflammatory in human monocytes from healthy controls and patients with type 2 diabetes mellitus (T2D). Methods: Monocytes were isolated from 20 healthy controls and 30 patients with T2D. IL-6 and TNF concentrations were measured by ELISA. CTRP-3 was expressed in insect cells and used for stimulation experiments. Results: Basal IL-6 and TNF were not different in control and in T2D monocytes. LPS-stimulation (1 μg/ml) significantly (p < 0.001) increased IL-6 and TNF in the supernatants of control and in T2D monocytes to a similar extent. CTRP-3 (1 μg/ml) significantly (p = 0.03) inhibited LPS-induced IL-6 in control monocytes but not in T2D monocytes. TNF upon co-stimulation with LPS and CTRP-3 was significantly (p = 0.012) lower in control than in T2D monocytes. LPS-induced TNF concentration was significantly and positively correlated with serum total cholesterol and LDL cholesterol in T2D patients. Conclusions: CTRP-3 inhibits LPS-induced IL-6 and TNF release. This anti-inflammatory effect is lost in T2D. Serum cholesterol concentration affects the pro-inflammatory potential of LPS to induce TNF release from T2D monocytes in the presence or absence of CTRP-3. CTRP-3 might partly account for the pro-inflammatory state in T2D.     

Simplexide Induces CD1d-Dependent Cytokine and Chemokine Production from Human Monocytes

Monocytes are major effector cells of innate immunity and recognize several endogenous and exogenous molecules due to the expression of wide spectrum of receptors. Among them, the MHC class I-like molecule CD1d interacts with glycolipids and presents them to iNKT cells, mediating their activation. Simplexide belongs to a novel class of glycolipids isolated from marine sponges and is structurally distinct from other immunologically active glycolipids. In this study we have examined the effects of simplexide on cytokine and chemokine release from human monocytes. Simplexide induces a concentration- and time-dependent release of IL-6, CXCL8, TNF-α and IL-10 and increases the expression of IL6, CXCL8 and IL10 mRNA. Cytokine and chemokine release induced by simplexide from monocytes is dependent on CD1d since: i) a CD1d antagonist, 1,2-bis (diphenylphosphino) ethane [DPPE]- polyethylene glycolmonomethylether [PEG], specifically blocks simplexide-induced activation of monocytes; ii) CD1d knockdown inhibits monocyte activation by simplexide and iii) simplexide induces cytokine production from CD1d-transfected but not parental C1R cell line Finally, we have shown that simplexide also induces iNKT cell expansion in vitro. Our results demonstrate that simplexide, apart from activating iNKT cells, induces the production of cytokines and chemokines from human monocytes by direct interaction with CD1d.     

Inhibitory effect of estrogen on Rac1-expression in monocytes

Recruitment of circulating monocytes into the vasculature and release of reactive oxygen species (ROS) promote atherogenesis. Rac1-GTPase is an essential component of the superoxide-producing NADPH-oxidase complex. Estrogens inhibit production of vascular reactive oxygen species.Angiotensin II as well as overexpression of the constitutively active mutant RacL61 increased ROS production in monocytes. AngII-mediated ROS release was completely inhibited by overexpression of the dominant negative mutant RacN17 or treatment with 17β-estradiol. 17β-Estradiol reduced Rac1-expression concentration- and time-dependently and decreased basal, as well as AngII-induced Rac1 activity. The effects of 17β-estradiol were receptor-mediated. In vivo, down-regulation of Rac1 by 17β-estradiol was observed in human mononuclear cells of women with elevated 17β-estradiol levels after controlled ovarian hyperstimulation.In summary, the data show that down-regulation of Rac1-GTPase contributes to the inhibition of angiotensin II-mediated superoxide release by 17β-estradiol in monocytes.     

Release of egg development neurosecretory hormone in Aedes aegypti and Aedes taeniorhynchus induced by an ovarian factor

Ovariectomized Aedes aegypti do not synthesize vitellogenin after a blood meal, unless an ovary from a blood-fed donor is implanted. Decapitation, however, prior to implantation inhibits vitellogenin synthesis. A female ovariectomized and decapitated 6 hr after a blood meal, synthesizes vitellogenin if an ovary from a blood-fed donor is implanted. On the other hand, females that are fed on blood and immediately decapitated can not be stimulated to synthesize vitellogenin with implanted ovaries removed from blood-fed donors. These experiments led to the hypothesis that the blood meal stimulates the ovary to secrete a corpus cardiacum stimulating factor, that in turn promotes release of egg development neurosecretory hormone stored in the corpus cardiacum.Injection of 20-hydroxy-ecdysone or ovarian extract prepared from ovaries removed from unfed females does not release egg development neurosecretory hormone. Thus corpus cardiacum stimulating factor is not 20-hydroxy-ecdysone, and ovaries removed from unfed females do not store it.The rate of inactivation of egg development neurosecretory hormone released from the corpus cardiacum after a blood meal was investigated by implanting an ovary into females that were blood fed for various intervals than decapitated and ovariectomized. Seventy per cent of implants grow when the operation is done 18 hr after feeding, and 30% when the operation is done between 18 and 24 hr after feeding, indicating that egg development neurosecretory hormone is stable for the first 18 hr after a blood meal.Aedes taeniorhynchus females ovariectomized 24 hr after adult emergence do not synthesize vitellogenin. When such a female is implanted with an ovary removed from a sugar-fed or blood-fed Aedes aegypti donor vitellogenin synthesis is initiated, and the implant grows. Decapitation prior to implantation inhibit vitellogenin synthesis and implants do not grow. These results indicate that corpus cardiacum stimulating factor is not species specific.     

Processing of interleukin-1 in cells of monocytic lineage is differentiation-dependent.

The interleukin-1 (IL-1) alpha and beta precursor proteins are processed and released from several cell types in the absence of a canonical signal peptide. To gain some insight into the mechanisms that allow the production of IL-1 alpha and beta, we have investigated by immunoprecipitation the synthesis, their release and processing in a promyeloblastic cell line of tumoral origin, U937, and in peripheral blood monocytes. We show that U937 monocytic cells, on induction with a tumor-promoting agent, synthesize and release into the culture medium proIL-1 beta but do not process it. Similarly, peripheral blood monocytes left in adherence for 24 h or longer, prior to addition of lipopolysaccharide, synthesize and release proIL-1 alpha and beta without detectable processing of either cytokine. Processing and release of IL-1 alpha and beta by peripheral blood monocytes can be observed when monocytes are left to adhere for periods less than 15 h before lipopolysaccharide addition. IL-1 alpha and beta show similar kinetics of release from the cells, suggesting the existence of a common mechanism regulating their secretion. Since peripheral blood monocytes left in adherence in the presence of lipopolysaccharide differentiate into macrophages, we conclude that release and processing of IL-1 can occur independently and that processing depends on the stage of differentiation of monocytes, i.e. only the monocytes at an early stage of differentiation produce 17-kDa IL-1 alpha and beta.     

Presentation of nitric oxide regulates monocyte survival through effects on caspase-9 and caspase-3 activation

In the absence of survival factors, blood monocytes undergo spontaneous apoptosis, which involves the activation of caspase-3. Although nitric oxide can block caspase-3 activation and promote cell survival, it can also induce apoptosis. We hypothesized that nitrosothiols that promote protein S-nitrosylation would reduce caspase-3 activation and cell survival, whereas nitric oxide donors (such as 1-propamine 3-(2-hydroxy-2-nitroso-1-propylhydrazine (PAPA) NONOate and diethylamine (DEA) NONOate) that do not target thiol residues would not. Using human monocytes as a model, we observed that nitrosothiol donors S-nitrosoglutathione and S-nitroso-N-acetylpenicillamine suppressed caspase-9 and caspase-3 activity and DNA fragmentation. In contrast, PAPA or DEA NONOate did not promote monocyte survival events and appeared to inhibit monocyte survival induced by macrophage colony-stimulating factor. The caspase-3-selective inhibitor DEVD-fluoromethyl ketone reversed DNA fragmentation events, and the caspase-9 inhibitor LEHD-fluoromethyl ketone reversed caspase-3 activity in monocytes treated with PAPA or DEA NONOate in the presence of macrophage colony-stimulating factor. These results were not caused by differences in glutathione levels or the kinetics of nitric oxide release. Moreover, S-nitrosoglutathione and S-nitroso-N-acetylpenicillamine directly blocked the activity of recombinant caspase-3, which was reversed by the reducing agent dithiothreitol, whereas PAPA or DEA NONOate did not block the enzymatic activity of caspase-3. These data support the hypothesis that nitrosylation of protein thiol residues by nitric oxide is critical for promoting the survival of human monocytes.     

Downregulation of nuclear factor-kappa B (NF-κB) pathway by silibinin in human monocytes challenged with Paracoccidioides brasiliensis

AimsSilibinin is the major active component of silymarin, a polyphenolic plant flavonoid that has anti-inflammatory effects. The modulatory effect of silibinin on monocyte function against Paracoccidioides brasiliensis (Pb18) has not yet been demonstrated. The present study investigated whether the effect of silibinin on nuclear factor-kappa B (NF-κB) pathways may affect the production of tumor necrosis factor-alpha (TNF-α), interleukin-10 (IL-10), transforming growth factor beta (TGF-β1), prostaglandin E₂ (PGE₂), nitric oxide (NO) and fungicidal activity of human monocytes challenged in vitro with Pb18.Main methodsPeripheral blood monocytes from healthy individuals were treated with silibinin and challenged with Pb18 for 18 h. TNF-α, IL-10, TGF-β1 and PGE₂ expression were determined by immunoenzymatic assay (ELISA) and NO release was determined by the accumulation of nitrite in culture supernatants. Fungicidal activity of monocytes was analyzed after treatment with interferon-gamma plus silibinin and challenge with Pb18. NF-κB activation in cultured monocytes was evaluated by flow cytometry and ELISA.Key findingsSilibinin partially inhibited p65NF-κB activation as the number of cells expressing this factor was reduced and the concentration of nuclear p65NF-κB was low, compared to untreated controls. The addition of silibinin also resulted in suppression of TNF-α, IL-10, TGF-β1, PGE₂ and NO production but did not affect the fungicidal activity of monocytes against Pb18.SignificanceSilibinin exerts anti-inflammatory and anti-fibrotic effects on CD14± human monocytes challenged by Pb18 by partial inhibition of p65NF-κB activation.     

Tissue factor as a proinflammatory agent

Tissue factor (TF) is a transmembrane glycoprotein and the main triggering element of blood coagulation. TF expression on monocytes and endothelial cells is induced by exposure to endotoxin, tumor necrosis factor, and IL-1 and is considered to appear in consequence of inflammation. In order to assess the proinflammatory capacity of TF itself, the recombinant extracellular domain of TF was injected intra-articularly into healthy mice. To characterize the role of immune cells in the TF-induced arthritis, mice deprived of lymphocytes, neutrophils and monocytes were used. Histomorphological analysis of the joints with respect to inflammatory cell infiltration, pannus formation and erosion formation revealed development of arthritis in 80% of animals injected with TF. In most of the cases synovial proliferation was accompanied by pannus formation and cartilage destruction. Inflammatory cell infiltrate consisted of CD4^-Mac1⁺ macrophages. Depletion of monocytes was, however, not enough to abolish inflammation. Indeed, combined deficiency of monocytes and lymphocytes was required to prevent inflammation following the injection of TF. We observed that TF induced chemokine production (MIP-1α and RANTES), but did not induce a proliferative response nor cytokine release by mouse spleen cells. TF has strong inflammatogenic properties mediated predominantly by monocytes and their release of chemokines. Our study shows that TF can simultaneously trigger the immune and coagulation systems.     

Interactions between cAMP- and cGMP-dependent protein kinase inhibitors and phosphodiesterase IV inhibitors on arachidonate release from human monocytes

The effects of specific inhibitors of cAMP-dependent protein kinase (PKA) and cGMP-dependent protein kinase (PKG) on the inhibitory activity of phosphodiesterase (PDE) type IV inhibitors and of the cell permeable analogue of cAMP, db-cAMP, were investigated on fMLP-induced arachidonate release from human monocytes. When monocytes were preincubated with the combined PKA/PKG inhibitor H8 (10⁻⁶ to 10⁻⁴ M) or the selective PKG inhibitor Rp-8-cpt-cGMPs (10⁻⁶ to 10⁻⁴ M) a concentration-dependent reduction of the inhibitory effect of db-cAMP (10⁻ M), rolipram (10⁻⁵ M) and Ro 20-1724 (10⁻⁵ M) was noted. When monocytes were preincubated with the selective PKA inhibitor H89 (10⁻⁶ to 10⁻⁴ M), only a small inhibition of the effect of db-cAMP and no inhibition of the effects of rolipram and Ro 20–1724 were observed. The present data indicate that db-cAMP and PDE IV inhibitors elicit an in vitro anti-inflammatory activity by a PKA-independent mechanism, which do not appear to be mainly mediated via the PKG activation.     

Extracellular Histones Induce Chemokine Production in Whole Blood Ex Vivo and Leukocyte Recruitment In Vivo

The innate immune system relies to a great deal on the interaction of pattern recognition receptors with pathogen- or damage-associated molecular pattern molecules. Extracellular histones belong to the latter group and their release has been described to contribute to the induction of systemic inflammatory reactions. However, little is known about their functions in the early immune response to an invading pathogen. Here we show that extracellular histones specifically target monocytes in human blood and this evokes the mobilization of the chemotactic chemokines CXCL9 and CXCL10 from these cells. The chemokine induction involves the toll-like receptor 4/myeloid differentiation factor 2 complex on monocytes, and is under the control of interferon-γ. Consequently, subcutaneous challenge with extracellular histones results in elevated levels of CXCL10 in a murine air pouch model and an influx of leukocytes to the site of injection in a TLR4 dependent manner. When analyzing tissue biopsies from patients with necrotizing fasciitis caused by Streptococcus pyogenes, extracellular histone H4 and CXCL10 are immunostained in necrotic, but not healthy tissue. Collectively, these results show for the first time that extracellular histones have an important function as chemoattractants as their local release triggers the recruitment of immune cells to the site of infection.     

Monokines released during short-term Fc gamma receptor phagocytosis up-regulate polymorphonuclear leukocytes and monocyte-phagocytic function.

Freshly explanted monocytes phagocytosing IgG antibody-coated erythrocyte targets (EIgG) release a factor(s) that stimulates phagocytosis by neighboring monocytes and polymorphonuclear leukocytes (PMN). Culture supernatants obtained after 30-min incubation of adherent monocytes with EIgG, but not unopsonized sheep erythrocytes, markedly up-regulated the extent of PMN phagocytosis and enhanced the rate at which monocytes ingested EIgG. The presence of this factor(s) was first evident in phagocytic studies in which monocytes were prepared by a colloidal silica-based continuous gradient technique (Sepracell-Mn). After introduction of erythrocyte targets, there was a 20- to 30-min delay before initiation of phagocytosis that was not observed with monocytes prepared by the standard Percoll-gradient technique. Experiments suggest that, when compared with monocytes prepared by the Percoll-gradient method, Sepracell-Mn monocytes are closer to a base line state of activation with regard to the expression of Fc gamma RI and the ability to ingest EIgG. The mechanism of PMN upregulation by the monocyte factor(s) was explored. Monocyte supernatants did not induce an increase in the surface expression of PMN Fc gamma RI, II, or III. Neither anti-TNF, anti-IL-2, nor anti-GM-CSF had any significant effect on monocyte supernatant activity. Neutrophil activating protein-1 was not detected by ELISA. In contrast, anti-IL-1 completely blocked the effect of the supernatant on subsequent monocyte phagocytosis, and partially inhibited its effect on PMN phagocytosis. Furthermore, it was shown that RIL-1 as well as TNF markedly enhanced monocyte and PMN ingestion of EIgG. These results suggest that monocytes, after Fc gamma R-mediated phagocytosis, release monokines, including at least IL-1, which enhance the phagocytic function of neighboring PMN and monocytes to augment the host defense process.     

Ex Vivo Cytokine Release and Pattern Recognition Receptor Expression of Subjects Exposed to Dampness: Pilot Study to Assess the Outcome of Mould Exposure to the Innate Immune System

In rooms with moisture damage, the indoor air can be enriched with microorganisms causing a variety of symptoms. Due to the highly diverse composition of bioaerosols and the multiple effects on humans, an assessment of the health risk is not sufficiently possible. The aim of this study was to characterize the features of innate immunity using blood from subjects exposed to moisture damage compared to control subjects living in houses without visible moisture damage. We investigated the expression of TLR-2, TLR-4 and dectin-1 on the surface of monocytes from both fresh blood and after in vitro stimulation with the model substances E. coli endotoxin, zymosan A, Pam₃Cys and Aspergillus versicolor in 25 exposed subjects and 25 control subjects. In vitro stimulation of whole blood with the same components was performed for 20 h and the release of inflammatory mediators IL-8 and IL-1β were quantified. In addition to an enhanced number of blood leucocytes, the expression of the receptors TLR-2, TLR-4 and dectin-1 on blood monocytes was significantly enhanced in exposed subjects. In contrast, no different alteration in expression was detected between exposed and control group after in vitro stimulation with the model substances. The release of IL-8 and IL-1β after stimulation of whole blood with A. versicolor was increased in subjects exposed to moisture damage. Furthermore, in the exposed subjects the IL-1β release was significantly enhanced after in vitro stimulation with E. coli endotoxin (1000 pg/mL). In conclusion, features of the innate immune system (receptor expression and mediator release of monocytes) are altered in subjects exposed to moisture damage which may be a potential explanation for the increased incidence of respiratory health diseases observed in these populations.     

CD14-Dependent Monocyte Isolation Enhances Phagocytosis of Listeria monocytogenes by Proinflammatory,GM-CSF-Derived Macrophages

Macrophages are an important line of defence against invading pathogens. Human macrophages derived by different methods were tested for their suitability as models to investigate Listeria monocytogenes (Lm) infection and compared to macrophage-like THP-1 cells. Human primary monocytes were isolated by either positive or negative immunomagnetic selection and differentiated in the presence of granulocyte macrophage colony-stimulating factor (GM-CSF) or macrophage colony-stimulating factor (M-CSF) into pro- or anti-inflammatory macrophages, respectively. Regardless of the isolation method, GM-CSF-derived macrophages (GM-Mφ) stained positive for CD206 and M-CSF-derived macrophages (M-Mφ) for CD163. THP-1 cells did not express CD206 or CD163 following incubation with PMA, M- or GM-CSF alone or in combination. Upon infection with Lm, all primary macrophages showed good survival at high multiplicities of infection whereas viability of THP-1 was severely reduced even at lower bacterial numbers. M-Mφ generally showed high phagocytosis of Lm. Strikingly, phagocytosis of Lm by GM-Mφ was markedly influenced by the method used for isolation of monocytes. GM-Mφ derived from negatively isolated monocytes showed low phagocytosis of Lm whereas GM-Mφ generated from positively selected monocytes displayed high phagocytosis of Lm. Moreover, incubation with CD14 antibody was sufficient to enhance phagocytosis of Lm by GM-Mφ generated from negatively isolated monocytes. By contrast, non-specific phagocytosis of latex beads by GM-Mφ was not influenced by treatment with CD14 antibody. Furthermore, phagocytosis of Lactococcus lactis, Escherichia coli, human cytomegalovirus and the protozoan parasite Leishmania major by GM-Mφ was not enhanced upon treatment with CD14 antibody indicating that this effect is specific for Lm. Based on these observations, we propose macrophages derived by ex vivo differentiation of negatively selected human primary monocytes as the most suitable model to study Lm infection of macrophages.     

Alveolar macrophages have greater amounts of the enzyme 5-lipoxygenase than do monocytes.

Alveolar macrophages release greater amounts of leukotriene B4 (LTB4) and 5-hydroxyeicosatetraenoic acid (5-HETE) after A23187 stimulation than do blood monocytes. The mechanisms for this enhanced 5-lipoxygenase activity in alveolar macrophages are unknown. In these studies, we determined whether alveolar macrophages have greater amounts of the enzyme 5-lipoxygenase than do blood monocytes. We confirmed that alveolar macrophages released greater amounts of LTB4 after A23187 stimulation than did equivalent numbers of blood monocytes. In both the presence and absence of A23187, alveolar macrophages had greater amounts of immunoreactive 5-lipoxygenase, determined by Western analysis, on a per cell and a per protein basis than did blood monocytes. The amounts of 5-lipoxygenase enzyme in the cells roughly correlated with the amounts of LTB4 released by both types of cells. These observations suggest that A23187 stimulates alveolar macrophages to release greater amounts of LTB4 and 5-HETE than blood monocytes, in part, due to the greater amounts of 5-lipoxygenase.     

Experimentelle Untersuchungen zum Freisetzungsmechanismus von Neurohormonen nach elektrischer Reizung der Corpora cardiaca von Periplaneta americana in vitro

Releasing of neurohormones by in vitro stimulation of the corpora cardiaca of Periplaneta americana by means of suction electrodes and by simultaneous application of sympathicomimetics and -lytics as well as parasympathicomimetics and -lytics was investigated. The release of neurohormone D by stimulation of N.c.c. I in the presence of atropine in bath-fluid is inhibited. Contrary to this, the presence of eserine stimulates release. Application of reserpine, as well as of the sympathicolytics tolazolin, retinin, and guanethidin, does not influence release of this hormone in connexion with the stimulation of N.c.c. I. These results indicate that the release of neurohormone D by stimulation of N.c.c. I is regulated by cholinergic components. On the other hand, release of the hyperlgykaemic factor by electrical stimulation of N.c.c. II is controlled by adrenergic components. This conclusion results from the increase of the release rate caused by reserpine. Sympathicolytics decreased the release rate. Atropine and eserine do not influence hormone release in connexion with the stimulation of N.c.c. II.     

The production and characterisation of products of the lipoxygenase enzyme system released by rat peritoneal macrophages

Rat peritoneal monocytes and macrophages when exposed to the ionophore A23187 release products of the lipoxygenase pathway of arachidonic acid metabolism which cause the aggregation and chemokinesis of polymorphonuclear leucocytes suspensions. The major biologically active compound released was leukotriene B which accounted for greater than 80% of the activity. The remaining biological activity was due to the release of a more polar as yet unidentified compound. In addition rat macrophages release 5, 12 and 15-HETE but these mono-HETEs do not significantly contribute to the biological activity.