Receptor Expression In Monocytes Research Articles

AB0128 Total Calcium-Sensing Receptor Expression in Circulating Monocytes is Increased in Rheumatoid Arthritis Patients with Severe Coronary Artery Calcification

BackgroundHuman circulating monocytes express the calcium-sensing receptor (CaSR) and are involved in atherosclerosis. Vascular calcification is commonly used as a subclinical marker of atherosclerosis and has been linked to increased...

Determination and Modulation of Total and Surface Calcium-Sensing Receptor Expression in Monocytes In Vivo and In Vitro

Expression of the calcium-sensing receptor (CaSR) has previously been demonstrated in human circulating monocytes (HCM). The present study was designed to measure CaSR expression in HCM and to examine its potential modulation by pro-inflammatory cytokines, Ca2+, vitamin D sterols in U937 cell line. Twenty healthy volunteers underwent blood sampling with subsequent isolation of peripheral blood mononuclear cells (PBMC) at 3 visits. Flow cytometry analysis (FACS) was performed initially (V1) and 19 days later (V2) to examine intra- and intersubject fluctuations of total and surface CaSR expression in HCM and 15 weeks later (V3) to study the effect of vitamin D supplementation. In vitro experiments were conducted to assess the effects of pro-inflammatory cytokines, calcidiol, calcitriol and Ca2+ on CaSR expression in U937 cell line. By FACS analysis, more than 95% of HCM exhibited cell surface CaSR staining. In contrast, CaSR staining failed to detect surface CaSR expression in other PBMC. After cell permeabilization, total CaSR expression was observed in more than 95% of all types of PBMC. Both total and surface CaSR expression in HCM showed a high degree of intra-assay reproducibility (<3%) and a moderate intersubject fluctuation. In response to vitamin D supplementation, there was no significant change for both total and surface CaSR expression. In the in vitro study, U937 cells showed strong total and surface CaSR expression, and both were moderately increased in response to calcitriol exposure. Neither total nor surface CaSR expression was modified by increasing Ca2+ concentrations. Total CaSR expression was concentration dependently decreased by TNFα exposure. In conclusion, CaSR expression can be easily measured by flow cytometry in human circulating monocytes. In the in vitro study, total and surface CaSR expression in the U937 cell line were increased by calcitriol but total CaSR expression was decreased by TNFα stimulation.

Interleukin-17 (IL-17) Expression Is Reduced during Acute Myocardial Infarction: Role on Chemokine Receptor Expression in Monocytes and Their in Vitro Chemotaxis towards Chemokines

The roles of immune cells and their soluble products during myocardial infarction (MI) are not completely understood. Here, we observed that the percentages of IL-17, but not IL-22, producing cells are reduced in mice splenocytes after developing MI. To correlate this finding with the functional activity of IL-17, we sought to determine its effect on monocytes. In particular, we presumed that this cytokine might affect the chemotaxis of monocytes important for cardiac inflammation and remodeling. We observed that IL-17 tends to reduce the expression of two major chemokine receptors involved in monocyte chemotaxis, namely CCR2 and CXCR4. Further analysis showed that monocytes pretreated with IL-17 have reduced in vitro chemotaxis towards the ligand for CCR2, i.e., MCP-1/CCL2, and the ligand for CXCR4, i.e., SDF-1α/CXCL12. Our results support the possibility that IL-17 may be beneficial in MI, and this could be due to its ability to inhibit the migration of monocytes.

Recurrent major depressive disorder: Imbalance of neurokinin (NK)-1 and NK-2 receptor expression in monocytes

Increasing evidence suggests that tachykinins are involved in the control of different pathological conditions, including psychiatric disorders. In this study we evaluated the expression of NK1 and NK2 receptors (NK-1R and NK-2R), as well as the effects of substance P (SP) and neurokinin A (NKA), in monocytes isolated from 15 healthy subjects and 15 patients with recurrent major depressive disorder (RMDD), under stable antidepressant therapy. NK-1R expression in monocytes from RMDD patients was significantly decreased as compared to healthy subjects, whereas NK-2R expression was markedly increased. Both NK-1R and NK-2R expression correlated with HAM-D, but not HAM-A, score. SP, NKA and selective NK-1R and NK-2R agonists stimulated TNF-α release in monocytes of both groups, with a significant higher effect observed in RMDD. Moreover they induced NF-κB activation, which was reversed by selective NK-1R and NK-2R antagonists, so demonstrating that it was receptor-mediated. The occurrence of a profound alteration in NK receptor expression in RMDD is a novel finding that suggests NK-1R and NK-2R pathways as possible relevant players in major depressive disorder, so improving our understanding of the complex pathogenesis of the disease.

P015 TSLP enhances type 1 immune response in cutaneous leishmaniasis patients

Introduction Leucocytes from cutaneous leishmaniasis (CL) patients due to L. braziliensis infection secrete high levels of IFN-gamma and TNF-alpha in response to soluble Leishmania antigen (SLA) and at lesion site. This type of immune response helps to control parasite multiplication but also mediates tissue damage and lesion development. Thus, we think CL patients would benefit from a balanced Th1/Th2 immune response. It has been shown that cells that do not classically participate in immune response, such as epithelial cells, can secrete soluble factors interfering on T cells responses to pathogens. Thymic stromal lymphopoietin (TSLP) is a recently described cytokine secreted by epithelial cells. TSLP-activated macrophages and dendritic cells enhance IL-4 and IL-13 secretion by memory CD4+ T cells in asthma and helminthes infections. Methods Peripheral blood mononuclear cells were obtained from CL individuals. Cells were stimulated with soluble Leishmania antigen or D. pteronyssinus in presence or absence of recombinant TSLP for 72 h. Cytokines were measured in cell culture supernatants by ELISA. For immunohistochemistry, biopsies were obtained from CL patients and stained for TSLP. Results Since TSLP induces Th2 responses, our hypothesis was that presence of TSLP would conditionate antigen presenting cells to enhance SLA-induced Th2 responses in CL patients decreasing levels of Th1 cytokines. Surprisingly, our data shows that in presence of TSLP, SLA-stimulated PBMC from CL individuals enhance IFN-gamma production, instead. No change in SLA-induced IL-5, TNF-alpha and IL-10 was observed. To test the effect of TSLP on antigens known to induce Th2 responses, we stimulated cells from the same patients with D. pteronyssinus (Derp) antigen in presence or absence of TSLP. As expected, presence of TSLP enhanced Derp-induced IL-5 production. The fact that TSLP can be expressed by skin epithelial cells, led us to investigate whether there was presence of TSLP in lesion of CL patients. Immunohistochemistry reveled TSLP expression in epithelial cells, endothelial cells and macrophages from CL individuals. We finally asked if Leishmania infection would interfere on TSLP receptor expression in monocytes. L. braziliensis-infected monocytes from healthy individuals up-regulated TSLP receptor expression. Conclusion Our results show a novel role for TSLP in inducing up-regulation of IFN-gamma production in CL patients. These data suggest that presence of TSLP may contribute for parasite killing and also maintenance of inflammatory response in these patients.

Preoperative Toll‐like Receptor Expression in Monocytes is Associated with Host Immunological Responses Following Gastrointestinal Surgery

The present study was designed to evaluate the relationship between preoperative Toll-like receptor (TLR) expression in peripheral blood monocytes (PBMC) and the postoperative inflammatory cytokine secretion from PBMC, as well as the postoperative infectious complications in patients with gastrointestinal cancer. Peripheral blood monocytes were isolated from 26 patients on the preoperative day and on postoperative day 1 (POD1). The monocytes were stained for CD14, TLR2, and TLR4 surface expression and stimulated ex vivo with lipopolysaccharide (LPS), after which the culture supernatant tumor necrosis factor alpha (TNFα), interleukin (IL)-1β, and interferon gamma (IFNγ) concentrations were measured in the culture supernatant. Postoperative clinical courses were monitored. There was a significant positive correlation between preoperative TLR4 expression and clinical stage in patients with gastrointestinal cancer. A significant negative correlation between preoperative TLR2 or TLR4 expression and the preoperative TNFα and IL-1β production was found. Furthermore, there was a significant negative correlation between preoperative TLR2 or TLR4 expression and postoperative inflammatory cytokine production. Preoperative high expression of TLR on monocytes reduces the capacity for LPS-induced inflammatory cytokine production from monocytes. An analysis of preoperative TLR expression may therefore help to elucidate the systemic immunological response and the clinical course following gastrointestinal surgery.

Toll-like receptor expression in monocytes in patients with chronic kidney disease and haemodialysis: relation with inflammation

Inflammation is one of the main contributors to atherosclerosis in haemodialysis (HD) patients. Activation of Toll-like receptors (TLRs) leads to inflammatory response. In this study, we aimed to evaluate the expression of TLRs on monocytes and relate their expression with inflammation in chronic kidney disease (CKD) and HD patients. Thirty-four age- and gender-matched controls and stage 3-4 CKD patients and thirty-two HD patients were included in each study group. The effect of HD on the expression of Toll-like receptor-2 (TLR-2) and Toll-like receptor-4 (TLR-4) on CD14( +) monocytes was determined at the beginning (baseline), during (120 min) and following (300 min and 24 h) HD and compared with control and stage 3-4 CKD groups. The HD procedure was performed by using low-flux polysulphone dialysers. In addition, serum IL-6 levels were evaluated in both groups at baseline and after a HD session. The percentage of CD14( +) monocytes expressing TLR-2 were similar in all of the study groups, whereas the percentage of CD14( +) monocytes expressing TLR-4 were significantly lower in both stage 3-4 CKD and HD patients at baseline than in controls. The mean fluorescence intensities (MFI) of TLR-2 were significantly lower in controls than in stage 3-4 CKD and HD patients at baseline. The MFI of TLR-4 was similar in all of the groups. The percentage of CD14( +) monocytes expressing TLR-2 did not change during and after HD. The MFI of TLR-2 decreased at 120 min of HD compared with baseline (1837 ± 672 vs 1650 ± 578, P < 0.05), and recovered back to baseline values at 300 min and at 24 h post-HD. MFI of TLR-4 increased at 24 h compared with baseline (941 ± 294 vs 1087 ± 441, P < 0.05). Serum IL-6 levels correlated with MFI of TLR-2 and TLR-4 in stage 3-4 CKD patients and in HD patients at baseline and after HD in univariate analysis. Stepwise multiple regression analysis revealed that MFI of TLR-2 was an independent determinant of serum IL-6 concentrations in stage 3-4 CKD and in HD patients at baseline, at 300 min and at 24 h post-HD. Conclusions. Our study demonstrates that TLR-2 is associated with the inflammatory response of non-dialysed and dialysed CKD patients.

Anti-inflammatory haemoglobin scavenging monocytes are induced following coronary artery bypass surgery

Circulating monocytes may counter systemic pro-inflammatory and haemolytic insults through the expression and shedding of the haemoglobin scavenger receptor (CD163). This prospective study aims to assess the effects of coronary artery bypass grafting with and without cardiopulmonary bypass (CPB) on haemoglobin scavenging and anti-inflammatory monocyte behaviour. Forty consecutive patients underwent coronary surgery using CPB (n=20) and off-pump (n=20) techniques. Peri-operative blood samples were taken until the fifth day following surgery and statistical comparison performed to baseline using group- and subject-based analysis. Monocyte receptor expression and plasma concentrations of anti-inflammatory molecules were measured by flow cytometry and enzyme-linked immunosorbent assay, respectively. Monocyte CD163 expression was significantly elevated post-operatively in both surgical groups with (p=0.001) and without CPB (p=0.000) at 24-48h. By contrast, shed CD163 (p=0.02) and haemoglobin-haptoglobin complexes (p=0.000) in plasma were only significantly elevated in the on-pump group at 2-4h. Subject-based analysis demonstrated that CPB is an independent predictor of monocyte CD163 (p=0.002) and plasma sCD163 (p=0.01) concentration adjusting for the measurement timing. Significant downregulation of the monocyte major histocompatibility complex II receptor was observed in both surgical groups, whereas lipopolysaccharide receptor (CD14) expression only declined following on-pump surgery. The anti-inflammatory cytokine interleukin (IL)-10 was elevated post-operatively in both surgical groups peaking earlier in the on-pump group (2h) versus off-pump group (4h). The immunoregulatory IL-6 cytokine was significantly upregulated at 4h in both groups. Coronary artery bypass surgery induces monocyte-associated anti-inflammatory and immunoregulatory responses. There was no significant difference in haemolysis although the effect on monocyte CD163 and plasma sCD163 concentration was significantly different between the two groups. Compensation for varying pro-inflammatory and haemolytic insults may explain the differences observed. Therapeutic strategies for inducing such desirable circulating monocytes may potentially improve surgical outcome and patient recovery.

GM-CSF and IL-4 induce dendritic cell differentiation and disrupt osteoclastogenesis through M-CSF receptor shedding by up-regulation of TNF-α converting enzyme (TACE)

AbstractMonocytes give rise to macrophages, osteoclasts (OCs), and dendritic cells (DCs). Macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-kappaB (RANK) ligand induce OC differentiation from monocytes, whereas granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4) trigger monocytic differentiation into DCs. However, regulatory mechanisms for the polarization of monocytic differentiation are still unclear. The present study was undertaken to clarify the mechanism of triggering the deflection of OC and DC differentiation from monocytes. GM-CSF and IL-4 abolished monocytic differentiation into OCs while inducing DC differentiation even in the presence of M-CSF and RANK ligand. GM-CSF and IL-4 in combination potently up-regulate tumor necrosis factor-α (TNF-α) converting enzyme (TACE) and activity in monocytes, causing ectodomain shedding of M-CSF receptor, resulting in the disruption of its phosphorylation by M-CSF as well as the induction of osteoclastogenesis from monocytes by M-CSF and RANK ligand. Interestingly, TACE inhibition robustly causes the resumption of the surface expression of M-CSF receptor on monocytes, facilitating M-CSF–mediated phosphorylation of M-CSF receptor and macrophage/OC differentiation while impairing GM-CSF– and IL-4–mediated DC differentiation from monocytes. These results reveal a novel proteolytic regulation of M-CSF receptor expression in monocytes to control M-CSF signaling and monocytic differentiation into macrophage/OC-lineage cells or DCs.

Interleukin-18 enhances cytokine secretion by monocytes activated through direct contact with T lymphocytes

Objective:To study whether interleukin (IL)-18 is involved in the activation of monocytes through direct contact with T cells and the related intracellular mechanism. Methods:T cells and monocytes were isolated and purified from the peripheral blood of healthy donors by magnetic beads. Phytohemagglutinin (PHA) pre-stimulated T cells were fixed by 1% paraformaldehyde and were then co-cultured with monocytes at a T cellmonocyte ratio of 41. TNF-α and IL-18 levels in the supernatants were assayed by ELISA. Expression of IL-18 receptor α chain (IL-18Rα) on the surface of monocytes was analyzed by flow cytometry.Results:Monocytes activated by PHA-stimulated T cells produced significantly more TNF-α than by unstimulated T cells; non-cultured T cells or monocytes hardly produced any TNF-α. Upon direct cellular contact,PHA pre-stimulated T cells also up-regulated IL-18Rα expression on the surface of monocytes and induced IL-18 production in monocytes,which could be suppressed by nuclear factor (NF)-κB inhibitor (N-acetyl-L-cysteine,NAC) or phosphatidyl-inositol (PI) 3 kinase inhibitor (LY294002),but not by mitogen activated protein kinase (MAPK) inhibitor (SB203580). Neutralizing anti-IL-18 monoclonal antibody dose-dependently inhibited the production of TNF-α by monocyte-stimulated T cells. IL-18 failed to induce TNF-α production by cultured monocytes alone,while dose-dependently enhanced TNF-α production in monocyte-stimulated T cells,which could be inhibited by NAC or LY294002,but not by SB203580.Conclusion:By direct cellular contact T cells can stimulate monocytes to produce TNF-α and IL-18,up-regulate IL-18 receptor expression in monocytes,and activate intracellular NF-κB and PI3 kinase pathways. IL-18 can enhance T cell ability to stimulate TNF-α production by monocytes,which is dependent on the activation of NF-κB and PI3 kinase pathways.

M1632 Interferon-γ Induces CCK2 Receptor Expression in Monocytes and Bone Marrow-Derived Cells

M1632 Interferon-γ Induces CCK2 Receptor Expression in Monocytes and Bone Marrow-Derived Cells

A novel activity for substance P: stimulation of peroxisome proliferator‐activated receptor‐γ protein expression in human monocytes and macrophages

Substance P (SP) and peroxisome proliferator-activated receptor-gamma (PPAR-gamma) play important roles in different inflammatory conditions and are both expressed in human monocytes and macrophages. However, it is not known whether or not they interact. This study was undertaken to evaluate the effects of SP on PPAR-gamma protein expression in monocytes and macrophages (MDMs: monocyte-derived macrophages) from healthy smokers and non-smokers. PPAR-gamma protein was detected by western blot and quantified by calculating the ratio between PPAR-gamma and beta-actin protein expression. Constitutive tachykinin NK(1) receptor expression in monocytes and MDMs from healthy smokers and non-smokers was evaluated by western blot. Cytokine release was evaluated by ELISA. In the concentration range 10(-10)-10(-6) M, SP stimulated PPAR-gamma protein expression in monocytes and MDMs, being more effective in cells from healthy smokers. Moreover, in these cells there was a constitutively increased expression of NK(1) receptors. SP-induced expression of the PPAR-gamma protein was receptor-mediated, as it was reproduced by the NK(1) selective agonist [Sar(9)Met(O(2))(11)]SP and reversed by the competitive NK(1) antagonist GR71251. SP-induced maximal effects were similar to those evoked by 15-deoxy-Delta(12,14)-prostaglandin J(2); an endogenous PPAR-gamma agonist, and were significantly reduced by a PPAR-gamma antagonist. NK(1) and PPAR-gamma agonists exerted opposite effects on TNF-alpha release from monocytes and MDMs. Enhancement of PPAR-gamma protein expression represents a novel activity for SP, which could contribute to a range of chronic inflammatory disorders.

Neutrophil and monocyte receptor expression in uncomplicated and complicated influenza A infection with pneumonia

Following influenza, the elderly and those with chronic heart/lung diseases are often affected by bacterial complications such as pneumonia. Whether neutrophil and monocyte functions are affected differently in patients with or without complications is less well known. Therefore, blood neutrophil and monocyte surface receptor expressions were measured in patients with influenza A, with or without complications, by means of flow cytometry. Neutrophil expressions of the adhesion molecules CD11b and CD66b were increased in influenza A, with the highest expression of CD11b in uncomplicated influenza. Monocyte expressions of CD11b and CD18 were also higher in influenza compared with bacterial infection and healthy controls. Neutrophil expressions of the phagocyte receptors CD64 and CD32 and the complement receptor CD35 were impaired in influenza with and without pneumonia compared with bacterial infection, whereas the expressions in monocytes were increased in all infected groups. The expression of the phagocyte receptor CD16 on neutrophils was impaired in all infected groups. Our results suggest increased recruitment of neutrophils and monocytes to infected areas by up-regulation of adhesion molecules in influenza that may be involved in the inflammatory response during infection. In contrast, depression of phagocyte receptor expression on neutrophils in patients with influenza pneumonia may contribute to increased susceptibility to bacterial infections and impaired clearance of encapsulated bacteria such as pneumococci.

Defective Receptor Expression and Dendritic Cell Differentiation of Monocytes in Glioblastomas

Better characterization of the changes that occur in the circulating monocytes of patients with glioblastoma has become more important recently as monocyte-derived dendritic cells are used as adjuvants in the development of glioma vaccines. This study seeks to develop understanding of the phenotypic changes that occur in circulating monocytes of patients with intracranial cancer and to assess the ability of these cells to differentiate into mature dendritic cells. Monocyte expression levels of HLA-ABC, HLA-DR, CD86, ICAM-1, TNFRII, and GMCSFR were compared between three cohorts: patients with intracranial glioblastoma (n = 15), patients with intracranial metastases (n = 9), and a group of healthy controls (n = 10). Monocytes were then tested for their ability to differentiate into mature dentritic cells based on morphology, CD83 expression and high levels of co-stimulatory molecules. Comprehensive analysis of monocyte receptor expression demonstrated significantly reduced HLA-ABC, HLA-DR, CD86, ICAM-1, and TNFRII in patients with glioblastoma but not in patients with intracranial metastases compared with a group of healthy controls. GMCSFR expression was significantly reduced in both patients with glioblastoma and intracranial metastases. Additionally, the monocytes of patients with glioblastoma showed a reduced capacity to differentiate into mature dendritic cells as identified by CD83 expression, receptor expression, and morphology. Peripheral monocytes are phenotypically altered in the setting of glioblastoma and display a reduced functional capacity to differentiate into mature dendritic cells.

Distinct toll-like receptor expression in monocytes and T cells in chronic HCV infection

Hepatitis C virus often establishes chronic infections. Recent studies suggest that viral and bacterial infections are more common in HCV-infected patients compared to controls. Pathogens are recognized by Toll-like receptors (TLRs) to shape adaptive and innate immune responses. In this study, to assess the ability of HCV-infected host to recognize invading pathogens, we investigated Toll-like receptor expression in innate (monocytes) and adaptive (T cells) immune cells by real-time PCR. We determined that RNA levels for TLRs 2, 6. 7, 8, 9 and 10 mRNA levels were upregulated in both monocytes and T cells in HCV-infected patients compared to controls. TLR4 was only upregulated in T lymphocytes, while TLR5 was selectively increased in monocytes of HCV-infected patients. MD-2, a TLR4 co-receptor, was increased in patients' monocytes and T cells while CD14 and MyD88 were increased only in monocytes. Our data reveal novel details on TLR expression that likely relates to innate recognition of pathogens and immune defense in HCV-infected individuals.

90 Neutrophil and monocyte receptor expression in uncomplicated and complicated influenza A with pneumonia

90 Neutrophil and monocyte receptor expression in uncomplicated and complicated influenza A with pneumonia

Human intestinal macrophages display profound inflammatory anergy despite avid phagocytic and bacteriocidal activity

Intestinal macrophages, which are thought to orchestrate mucosal inflammatory responses, have received little investigative attention compared with macrophages from other tissues. Here we show that human intestinal macrophages do not express innate response receptors, including the receptors for LPS (CD14), Fcα (CD89), Fcγ (CD64, CD32, CD16), CR3 (CD11b/CD18), and CR4 (CD11c/CD18); the growth factor receptors IL-2 (CD25) and IL-3 (CD123); and the integrin LFA-1 (CD11a/CD18). Moreover, resident intestinal macrophages also do not produce proinflammatory cytokines, including IL-1, IL-6, IL-10, IL-12, RANTES, TGF-β, and TNF-α, in response to an array of inflammatory stimuli but retain avid phagocytic and bacteriocidal activity. Thus, intestinal macrophages are markedly distinct in phenotype and function from blood monocytes, although intestinal macrophages are derived from blood monocytes. To explain this paradox, we show that intestinal stromal cell–derived products downregulate both monocyte receptor expression and, via TGF-β, cytokine production but not phagocytic or bacteriocidal activity, eliciting the phenotype and functional profile of intestinal macrophages. These findings indicate a mechanism in which blood monocytes recruited to the intestinal mucosa retain avid scavenger and host defense functions but acquire profound “inflammatory anergy,” thereby promoting the absence of inflammation characteristic of normal intestinal mucosa despite the close proximity of immunostimulatory bacteria.

Human intestinal macrophages display profound inflammatory anergy despite avid phagocytic and bacteriocidal activity

Intestinal macrophages, which are thought to orchestrate mucosal inflammatory responses, have received little investigative attention compared with macrophages from other tissues. Here we show that human intestinal macrophages do not express innate response receptors, including the receptors for LPS (CD14), Fcalpha (CD89), Fcgamma (CD64, CD32, CD16), CR3 (CD11b/CD18), and CR4 (CD11c/CD18); the growth factor receptors IL-2 (CD25) and IL-3 (CD123); and the integrin LFA-1 (CD11a/CD18). Moreover, resident intestinal macrophages also do not produce proinflammatory cytokines, including IL-1, IL-6, IL-10, IL-12, RANTES, TGF-beta, and TNF-alpha, in response to an array of inflammatory stimuli but retain avid phagocytic and bacteriocidal activity. Thus, intestinal macrophages are markedly distinct in phenotype and function from blood monocytes, although intestinal macrophages are derived from blood monocytes. To explain this paradox, we show that intestinal stromal cell-derived products downregulate both monocyte receptor expression and, via TGF-beta, cytokine production but not phagocytic or bacteriocidal activity, eliciting the phenotype and functional profile of intestinal macrophages. These findings indicate a mechanism in which blood monocytes recruited to the intestinal mucosa retain avid scavenger and host defense functions but acquire profound "inflammatory anergy," thereby promoting the absence of inflammation characteristic of normal intestinal mucosa despite the close proximity of immunostimulatory bacteria.

Differential Expression of the C5a Receptor and Complement Receptors 1 and 3 after LPS Stimulation of Neutrophils and Monocytes

Animal experiments recently suggested that administration of anti-C5a, anti-C5a receptor or soluble complement receptor type-1 may be of value in the treatment of septic shock. Because results regarding C5a receptor expression (C5a-R, CD-88) have been found to differ between septic animals and patients, the aim of this study was to investigate the neutrophil and monocyte receptor expression of CD-88 and complement receptor-1 (CR-1, CD-35) after stimulation with lipopolysaccharide (LPS) ex vivo. Whole blood or isolated neutrophils and monocytes from healthy people were incubated with LPS in a dose range of 0.1-1000 ng/ml. The expressions of CD-88 and CD-35 were analysed by means of flow cytometry. For comparison, the expressions of complement receptor-3 (CR-3, CD-11b/CD-18), Fc-gamma receptor type-I (CD-64) and CEACAM-8 (CD-66b) were also investigated. In whole blood, CD-88 expression on neutrophils was reduced (P < 0.05). The expressions of CD-35 and CD-11b were increased both on neutrophils (P < 0.001; P < 0.05) and on monocytes (P < 0.001; P < 0.001). No effect was observed on isolated cells. In agreement with the findings in septic patients, LPS reduced the neutrophil C5a-R expression, whereas the expressions of CR-1 and CR-3 were increased. The effects of LPS were indirect and were mediated via factors in the blood. The clinical significance of this is not known, but may be associated with decreased chemotaxis.

Interleukin-1 upregulates anaphylatoxin receptors on mononuclear cells

Background The anaphylatoxins, C3a and C5a, that are generated during trauma, major surgery, or infection are potent proinflammatory mediators that increase interleukin (IL-1) cytokine synthesis. We investigated the effects of IL-1 on anaphylatoxin receptor expression in monocytes. Methods A human monocytic cell line, MONO-MAC-6, was used. C3a and C5a binding sites were assayed by competitive binding. Levels of messenger RNA for the C3a and C5a receptors were analyzed by reverse transcriptase-polymerase chain reaction. Changes of free cytosolic Ca 2+ concentration ([Ca 2+]i) in response to C3a and C5a were measured. Results Basal MONO-MAC-6 cell sites for C3a and C5a binding were 10,900 C3aR/cell (K d = 2.0 nmol/L), 8700 C5aR/cell (K d = 0.9 nmol/L). IL-1α increased sites for both C3a (61% increase; P < .01) and C5a (71% increase; P < .001). Levels of C3aR and C5aR messenger RNA also increased in IL-1α-stimulated cells. Receptors were coupled to functional responses, which were demonstrated by C3a- or C5a-induced [Ca 2+]i increases. IL-1 receptor antagonist blocked the effects of IL-1α upregulation of anaphylatoxin receptors. Conclusion These results suggest that there is an additional link between IL-1 and anaphylatoxins to amplify proinflammatory effects through monocytes and macrophages. Although C3a and C5a can increase the monocyte production of IL-1, IL-1 increases monocyte expression of receptors for these anaphylatoxins, which further amplifies inflammation.