Vanillic acid inhibits inflammatory mediators by suppressing NF-κB in lipopolysaccharide-stimulated mouse peritoneal macrophages

Vanillic acid is a benzoic acid derivative that is used as a flavoring agent. It is an oxidized form of vanillin. At present, the mechanisms by which vanillic acid exerts its anti-inflammatory effects are incompletely understood. In this study, we attempted to determine the effects of vanillic acid on lipopolysaccharide (LPS)-induced inflammatory responses in mouse peritoneal macrophages. Our findings indicate that vanillic acid inhibits LPS-induced production of tumor necrosis factor (TNF)-α and interleukin (IL)-6. During the inflammatory process, the levels of cyclooxygenase (COX)-2 and nitric oxide (NO) increased in mouse peritoneal macrophages, but vanillic acid suppressed both the enhanced levels of COX-2 and the production of prostaglandin E₂ and NO. Moreover, vanillic acid suppressed the activation of nuclear factor-kappa B (NF-κB) and caspase-1. These results provide novel insights into the pharmacological actions of vanillic acid and are indicative of the potential use of this molecule in the treatment of inflammatory diseases.     

Sulfasalazine prevents T-helper 1 immune response by suppressing interleukin-12 production in macrophages

Interleukin-12 (IL-12) plays a pivotal role in the development of T-helper 1 (Th1) immune response, which may be involved in the pathogenesis of chronic inflammatory autoimmune disorders. In this study we investigated the effects of sulfasalazine, a drug for treating inflammatory bowel disease and rheumatoid arthritis, on the production of IL-12 from mouse macrophages stimulated with lipopolysaccharide (LPS). Sulfasalazine potently inhibited the production of IL-12 in a dose-dependent manner, in part through the down-regulation of nuclear factor κB (NFκB) activation in IL-12 p40 gene. Activation of macrophages by LPS resulted in markedly enhanced binding activities to the κB site, which significantly decreased upon addition of sulfasalazine as demonstrated by an electrophoretic gel shift assay. Importantly, macrophages pretreated with sulfasalazine either in vitro or in vivo reduced their ability to induce interferon-γ (IFN-γ) and increased the ability to induce IL-4 in antigen-primed CD4⁺ T cells. From these results, sulfasalazine may induce the Th2 cytokine profile in CD4⁺ T cells by suppressing IL-12 production in macrophages, and sulfasalazine-induced inhibition of IL-12 production in macrophages may explain some of the known biological effects of sulfasalazine.     

Ephedrine hydrochloride inhibits PGN-induced inflammatory responses by promoting IL-10 production and decreasing proinflammatory cytokine secretion via the PI3K/Akt/GSK3β pathway

Approaches for controlling inflammatory responses and reducing the mortality rate of septic patients remain clinically ineffective; new drugs need to be identified that can induce anti-inflammatory responses. Ephedrine hydrochloride (EH) is a compound that is widely used in cardiovascular diseases, especially to treat hypotension caused by either anesthesia or overdose of antihypertensive drugs. In this study, we reported that EH also plays an important role in the control of the inflammatory response. EH increased IL-10 and decreased proinflammatory cytokine (IL-6, tumor-necrosis factor (TNF)-α, IL-12 and IL-1β) expression in primary peritoneal macrophages and Raw264.7 cells treated with peptidoglycan (PGN), a Gram-positive cell wall component. The anti-inflammatory role of EH was also demonstrated in an experimental mouse model of peritonitis induced by intraperitoneal PGN injection. The phosphatidylinositol 3-kinase (PI3K)/Akt pathway was found to be responsible for the EH-mediated increase in IL-10 production and decrease in IL-6 expression. Therefore, our results illustrated that EH can help maintain immune equilibrium and diminish host damage by balancing the production of pro- and anti-inflammatory cytokines after PGN challenge. EH may be a new potential anti-inflammatory drug that can be useful for treating severe invasive Gram-positive bacterial infection.     

Subtilase Cytotoxin Enhances Escherichia coli Survival in Macrophages by Suppression of Nitric Oxide Production through the Inhibition of NF-κB Activation

Subtilase cytotoxin (SubAB), which is produced by certain strains of Shiga-toxigenic Escherichia coli (STEC), cleaves an endoplasmic reticulum (ER) chaperone, BiP/Grp78, leading to induction of ER stress and caspase-dependent apoptosis. SubAB alters the innate immune response. SubAB pretreatment of macrophages inhibited lipopolysaccharide (LPS)-induced production of both monocyte chemoattractant protein 1 (MCP-1) and tumor necrosis factor α (TNF-α). We investigated here the mechanism by which SubAB inhibits nitric oxide (NO) production by mouse macrophages. SubAB suppressed LPS-induced NO production through inhibition of inducible NO synthase (iNOS) mRNA and protein expression. Further, SubAB inhibited LPS-induced IκB-α phosphorylation and nuclear localization of the nuclear factor-κB (NF-κB) p65/p50 heterodimer. Reporter gene and chromatin immunoprecipitation (ChIP) assays revealed that SubAB reduced LPS-induced NF-κB p65/p50 heterodimer binding to an NF-κB binding site on the iNOS promoter. In contrast to the native toxin, a catalytically inactivated SubAB mutant slightly enhanced LPS-induced iNOS expression and binding of NF-κB subunits to the iNOS promoter. The SubAB effect on LPS-induced iNOS expression was significantly reduced in macrophages from NF-κB1 (p50)-deficient mice, which lacked a DNA-binding subunit of the p65/p50 heterodimer, suggesting that p50 was involved in SubAB-mediated inhibition of iNOS expression. Treatment of macrophages with an NOS inhibitor or expression of SubAB by E. coli increased E. coli survival in macrophages, suggesting that NO generated by macrophages resulted in efficient killing of the bacteria and SubAB contributed to E. coli survival in macrophages. Thus, we hypothesize that SubAB might represent a novel bacterial strategy to circumvent host defense during STEC infection.     

Low and moderate doses of ionizing radiation up to 2 Gy modulate transmigration and chemotaxis of activated macrophages,provoke an anti-inflammatory cytokine milieu,but do not impact upon viability and phagocytic function

Benign painful and inflammatory diseases have been treated for decades with low/moderate doses of ionizing radiation (LD-X-irradiation). Tissue macrophages regulate initiation and resolution of inflammation by the secretion of cytokines and by acting as professional phagocytes. Having these pivotal functions, we were interested in how activated macrophages are modulated by LD-X-irradiation, also with regard to radiation protection issues and carcinogenesis. We set up an ex-vivo model in which lipopolysaccharide pre-activated peritoneal macrophages (pMΦ) of radiosensitive BALB/c mice, mimicking activated macrophages under inflammatory conditions, were exposed to X-irradiation from 0·01 Gy up to 2 Gy. Afterwards, the viability of the pMΦ, their transmigration and chemotaxis, the phagocytic behaviour, the secretion of inflammatory cytokines and underlying signalling pathways were determined. Exposure of pMΦ up to a single dose of 2 Gy did not influence their viability and phagocytic function, an important fact regarding radiation protection. However, significantly reduced migration, but increased chemotaxis of pMΦ after exposure to 0·1 or 0·5 Gy, was detected. Both might relate to the resolution of inflammation. Cytokine analyses revealed that, in particular, the moderate dose of 0·5 Gy applied in low-dose radiotherapy for inflammatory diseases results in an anti-inflammatory cytokine microenvironment of pMΦ, as the secretion of the proinflammatory cytokine interleukin (IL)-1β was reduced and that of the anti-inflammatory cytokine transforming growth factor (TGF)-β increased. Further, the reduced secretion of IL-1β correlated with reduced nuclear translocation of nuclear factor (NF)-κB p65, starting at exposure of pMΦ to 0·5 Gy of X-irradiation. We conclude that inflammation is modulated by LD-X-irradiation via changing the inflammatory phenotype of macrophages.     

连翘提取物对小鼠腹腔巨噬细胞体外吞噬和NO释放的影响

目的:分析连翘(FS)对小鼠腹腔巨噬细胞的体外吞噬和体外NO释放的影响.方法:无菌收集小鼠腹腔巨噬细胞和羧基荧光素乙酰乙酸琥珀酰亚胺酯(CFDA-SE)标记大肠杆菌DH5α,短期培养3 h后流式细胞术(FCM)分析FS对腹腔巨噬细胞体外吞噬的影响.用LPS体外刺激活化腹腔巨噬细胞,Griess Reagent试剂盒检测并分析FS对巨噬细胞体外释放NO的影响.结果:FCM分析显示,终浓度为40、80、160 mg/L的FS对小鼠腹腔巨噬细胞体外吞噬具有明显的促进作用(P<0.05).不同终质量浓度的FS对LPS诱导小鼠腹腔巨噬细胞体外NO的释放均有抑制作用(P<0.05).结论:FS可以促进小鼠腹腔巨噬细胞的体外吞噬和抑制NO体外的释放.     

The plant hormone zeatin riboside inhibits T lymphocyte activity via adenosine A2A receptor activation

Cytokinins are plant hormones that play an integral role in multiple aspects of plant growth and development. The biological functions of cytokinins in mammalian systems are, however, largely uncharacterized. The naturally occurring cytokinin zeatin riboside has recently been demonstrated to activate the mammalian adenosine A_2A receptor, which is broadly expressed by various cell types including immune system cells, with the activation of the A_2AR playing a role in the regulation of cells involved in both innate and adaptive immunity. We show for the first time that zeatin riboside modulates mammalian immune system activity via an A_2AR-dependent mechanism. Specifically, zeatin riboside treatment induces the production of cyclic adenosine monophosphate (cAMP) by T lymphocytes and inhibits the production by CD3⁺CD4⁺ T cells of interferon (IFN)-γ, IL-2, tumor-necrosis factor (TNF)-α, IL-4 and IL-13, and the production by CD3⁺CD8⁺ T cells of IFN-γ, IL-2 and TNF-α. Additionally, the upregulation of CD25, CD69 and CD40L by activated T lymphocytes is modulated by zeatin riboside. Zeatin riboside treatment also potently inhibits thioglycollate-induced peritoneal leukocytosis. The immunomodulatory activities of zeatin riboside are blocked by co-treatment with the selective A_2AR antagonist ZM241385. These data suggest that zeatin riboside possesses therapeutic potential as a mammalian immunomodulatory agent.     

Vanillic acid inhibits inflammatory mediators by suppressing NF-κB in lipopolysaccharide-stimulated mouse peritoneal macrophages

Vanillic acid is a benzoic acid derivative that is used as a flavoring agent. It is an oxidized form of vanillin. At present, the mechanisms by which vanillic acid exerts its anti-inflammatory effects are incompletely understood. In this study, we attempted to determine the effects of vanillic acid on lipopolysaccharide (LPS)-induced inflammatory responses in mouse peritoneal macrophages. Our findings indicate that vanillic acid inhibits LPS-induced production of tumor necrosis factor (TNF)-α and interleukin (IL)-6. During the inflammatory process, the levels of cyclooxygenase (COX)-2 and nitric oxide (NO) increased in mouse peritoneal macrophages, but vanillic acid suppressed both the enhanced levels of COX-2 and the production of prostaglandin E(2) and NO. Moreover, vanillic acid suppressed the activation of nuclear factor-kappa B (NF-κB) and caspase-1. These results provide novel insights into the pharmacological actions of vanillic acid and are indicative of the potential use of this molecule in the treatment of inflammatory diseases.     

Activation of the RON receptor tyrosine kinase protects murine macrophages from apoptotic death induced by bacterial lipopolysaccharide

RON is a receptor tyrosine kinase activated by macrophage-stimulating protein. We demonstrate here that RON activation inhibits LPS-induced apoptosis of mouse peritoneal macrophages and Raw264.7 cells expressing RON or a constitutively active RON mutant. The antiapoptotic effect of RON was accompanied with the inhibition of LPS-induced production of nitric oxide (NO), a molecule responsible for LPS-induced cell apoptosis. This conclusion is supported by experiments using a chemical NO donor GSNO, in which RON activation directly blocked GSNO-induced apoptotic death of Raw264.7 cells and inhibited LPS-induced p53 accumulation. Furthermore, we showed that treatment of cells with wortmannin, which inhibits phosphatidylinositol (PI)-3 kinase, prevents the inhibitory effect of RON on LPS-induced macrophage apoptosis. These results were confirmed further by expression of a dominant inhibitory PI-3 kinase p85 subunit. These data suggest that by activating PI-3 kinase and inhibiting p53 accumulation, RON protects macrophage from apoptosis induced by LPS and NO. The antiapoptotic effect of RON might represent a novel mechanism for the survival of activated macrophages during inflammation.     

Enhancing effect of oxygen radical scavengers on murine macrophage anticryptococcal activity through production of nitric oxide

We examined the roles of reactive nitrogen intermediates (RNI) and reactive oxygen intermediates (ROI) in interferon-gamma (IFN-γ)-induced cryptococcostatic activity of murine peritoneal macrophages using N^G-monomethyl-L-arginine (L-NMMA), a competitive inhibitor of RNI synthesis, and superoxide dismutase (SOD) and catalase, oxygen radical scavengers. IFN-γ-activated macrophages produced nitric oxide (NO) in a dose-dependent manner, as measured by increased nitrite concentration in the culture supernatant. IFN-γ also enhanced the suppressive effect on cryptococcal growth in a similar dose-dependent manner. The induction of killing activity and NO production by an optimal dose of IFN-γ (100 U/ml) was virtually suppressed by 500 μM L-NMMA. These results confirmed the importance of the RNI-mediated effector mechanism in anticryptococcal activity of macrophages. SOD and catalase significantly enhanced the cryptococcostatic activity of macrophages induced by a suboptimal dose of IFN-γ (20 U/ml). The augmenting effect of these reagents was mediated by NO, since they potentiated the production of NO by macrophages and their effects were totally blocked by L-NMMA. Our results indicate that the IFN-γ-induced anticryptococcal activity of macrophages is dependent mostly on RNI, and suggest that the ROI system down-regulates the effector mechanism for cryptococcostasis by suppressing the RNI system.     

Effects of Strains of Lactococcus lactis on the Production of Nitric Oxide and Cytokines in Murine Macrophages

Nitric oxide (NO) is a multifunctional mediator that is involved in a variety of pathologic and physiologic processes. Few studies have addressed the effect of lactic acid bacteria (LAB), especially Lactococcus lactis strains used in dairy products, on inducible nitric oxide synthase (iNOS) induction as a component of the host’s gastrointestinal immune response. We investigated the ability of L. lactis strains to induce NO synthesis in the murine macrophage-like cell line J774.1 and in peritoneal macrophages from mice. The degree of NO induction was specific to the L. lactis strain used. Compared with the no-treatment control, heat treatment of L. lactis cells decreased NO and TNF-α levels but further stimulated interleukin (IL)-12 production. Adding L. lactis cells to peritoneal macrophages dose-dependently increased the production of NO and IL-10 but decreased that of IL-12p70. Adding L. lactis cells to interferon-γ-stimulated J774.1 cells enhanced cell death and the production of NO and IL-12p40, whereas addition of 1400W, a specific inhibitor of iNOS, decreased NO production and cell death. Conversely, adding 1400W to J774.1 cells further enhanced IL-12p40 production, suggesting that IL-12 production is perturbed by excess endogenous NO. IL-12 production is thought to be a marker of improved immunostimulation. Our results suggest that IL-12 production could be increased by limiting endogenous NO production.     

Differential nitric oxide (NO) production by macrophages from mice and guinea pigs infected with virulent and avirulent Legionella pneumophila serogroup 1

l-arginine-dependent reactive nitrogen intermediates have been identified as macrophage cytotoxic effector molecules against intracellular pathogens. To determine its role, ex vivo production of NO by peritoneal macrophages of C3H/HeN mice and Dunkin–Hartley guinea pigs infected intraperitoneally with a virulent and isogenic avirulent Legionella pneumophila serogroup 1 strain was compared with bacterial clearance from the lungs. While the virulent strain was cleared from mice lungs, the guinea pigs died within 96 h. In vivo infection with both strains resulted in the production of NO by mouse peritoneal macrophages ex vivo. In contrast, guinea pig macrophages did not produce detectable NO. In addition, infection by the avirulent strain led to the production of significantly more NO by mouse macrophages than the virulent parent strain, irrespective of stimulation with lipopolysaccharide (LPS) and/or interferon-gamma (IFN-γ). These results suggest that resistance to Leg. pneumophila infection may depend on the production of NO by host macrophages.     

CD4 lymphocytes in the blood of HIV(+) individuals migrate rapidly to lymph nodes and bone marrow: support for homing theory of CD4 cell depletion

Recent studies have provided evidence that macrophages from Th1-prone mouse strains respond with an M1 profile, and macrophages from Th2-prone mouse strains respond with an M2 profile, characterized by the dominant production of NO or TGF-beta 1, respectively. We have shown that peritoneal macrophages from IL-12p40 gene knockout mice have a bias toward the M2 profile, spontaneously secreting large amounts of TGF-beta 1 and responding to rIFN-gamma with weak NO production. Moreover, IL-12p40KO macrophages are more permissive to Trypanosoma cruzi replication than their wild-type littermate cells. Prolonged incubation with rIL-12 fails to reverse the M2 polarization of IL-12p40KO macrophages. However, TGF-beta 1 is directly implicated in sustaining the M2 profile because its inhibition increases NO release from IL-12p40KO macrophages. IFN-gamma deficiency is apparently not the reason for TGF-beta 1 up-regulation, because rIFN-gamma KO macrophages produce normal amounts of this cytokine. These findings raise the possibility that IL-12 has a central role in driving macrophage polarization, regulating their intrinsic ability to respond against intracellular parasites.     

The expression of interleukin-12 is increased by MAP kinase phosphatase-1 through a mechanism related to interferon regulatory factor 1

Mitogen-activated protein kinase phosphatase-1 (MKP-1) is a nuclear tyrosine/threonine phosphatase that inhibits p38 mitogen-activated protein kinase (MAPK) activity. We and others have shown that MKP-1 deficiency leads to excessive activation of innate immunity and inflammatory gene expression. Surprisingly, the present study shows that MKP-1 is a positive regulator of IL-12 expression in macrophages suggesting a stimulatory effect on Th1 type immune response. In the present study, we found that LPS-induced expression of IL-12p40 was lower in primary mouse peritoneal macrophages (PMs) and bone marrow-derived macrophages from MKP-1 deficient mice than in cells from wild-type mice whereas TNF expression was enhanced as expected. Correspondingly, the inhibition of p38 MAPK by pharmacologic inhibitors BIRB 796 and SB 202190 enhanced LPS-induced IL-12p40 production. Silencing of interferon regulatory factor 1 (IRF1) by siRNA inhibited the expression of IL-12p40 in J774 macrophages, showing that IRF1 is an important factor regulating IL-12p40 expression. BIRB 796 enhanced LPS-induced expression of IRF1 in J774 macrophages and in PMs from wild-type mice, and IRF1 expression was reduced in PMs from MKP-1 deficient mice. In conclusions, our results show that MKP-1 increases and p38 MAPK decreases the expression of IL-12 by enhancing the expression of IRF1. MKP-1, through regulation of IRF1 and IL-12, therefore may be an important factor supporting the development of Th1 type of immune response and anti-microbial defense.     

Cellular requirements for immunomodulatory effects caused by cell wall components of Paracoccidioides brasiliensis on antibody production

In a previous study, we reported an increase in the number of immunoglobulin-secreting cells and the augmentation of antibody production (IgM and IgG3) against unrelated antigens (sheep erythrocytes or bovine serum albumin (BSA)) in mice infected with the fungus Paracoccidioides brasiliensis as well as in mice inoculated with its cell wall preparation (CW). The immunomodulatory effect of the live fungus and CW preparation was dose-dependent and mainly restricted to the i.p. inoculation simultaneously to the BSA challenge by the i.v. route. In the present study, we investigated the active component of CW preparation upon the phenotype and also the degree of activation of possible target peritoneal cells involved in those phenomena. An insoluble polysaccharide fraction (F₁ fraction) mainly composed of β-glucan and chitin, and the purified β-glucan (BGPb) behaved as CW in the augmentation of early antibody production. The peritoneal mononuclear inflammatory cells induced by CW, F₁ fraction and BGPb were highly positive to α-naphthyl esterase staining; released low H₂O₂; expressed high levels of MHC-Ia^d molecules and produced inflammatory cytokines such as tumour necrosis factor-alpha (TNF-α) and IL-6. Phenotypic analysis by flow cytometry and immunohistochemical techniques of the inflammatory cells responding to F₁ fraction showed a prevalence of (CD11b/CD18, Mac-1)⁺ peritoneal macrophages. In addition, s.c. inoculation of F₁ fraction resulted in the formation of nodular, localized and not progressive granulomatous lesions with an accumulation of (CD11b/C18)⁺ macrophages. Adoptive transferred Mac-1 macrophages to immunized syngeneic recipient mice were able to cause an increase in anti-BSA antibody production. These results suggest that inflammatory (CD11b/CD18)⁺ macrophages may be related to immunological disturbances, caused by cell wall components of P. brasiliensis.     

Impaired macrophage responses may contribute to exacerbation of blood-stage Plasmodium chabaudi chabaudi malaria in interleukin-12-deficient mice.

Aiming to clarify the role of endogenous interleukin-12 (IL-12) in protective immunity against blood stages of Plasmodium chabaudi chabaudi (AS), we evaluated the course of infection in IL-12p40 gene knockout (IL-12p40KO) and wild-type (WT) C57BL/6 mice, focusing (1) on the ability of T cells to develop adequate type 1 responses and (2) on the potentiality of macrophages to respond to parasites, interferon-gamma (IFN-gamma), or both. We observed that IL-12p40KO mice develop significantly higher parasitemias during the acute infection, although mice from both groups clear the parasites within a month and similarly eliminate a secondary challenge. Thus, fully protective immunity to P. c. chabaudi can be generated in the absence of IL-12. However, this cytokine may promote parasite control during the early phase of infection. The increased acute parasitemia of IL-12p40KO mice was associated with both impaired IFN-gamma and nitric oxide (NO) response by spleen cells. Because stimulation with recombinant IFN-gamma (rIFN-gamma) failed to improve the NO response in IL-12p40KO macrophages, we investigated whether these cells have an intrinsic defect. Analysis of peritoneal macrophages revealed that IL-12p40KO cells produce higher levels of transforming growth factor-beta1 (TGF-beta1) compared with WT cells and respond to infected erythrocytes or rIFN-gamma by releasing little NO. Moreover, IL-12p40KO macrophages had a severely impaired ability to internalize opsonized infected erythrocytes, suggesting that the low effector profile assumed by these cells may compromise antibody-mediated immunity. Taken together, our results support the idea that the absence of IL-12p40 not only affects IFN-gamma production but also has deep consequences in macrophage effector functions that may contribute to exacerbation of the early phase of P. c. chabaudi malaria.     

Evasion of innate immune responses: evidence for mannose binding lectin inhibition of tumor necrosis factor alpha production by macrophages in response to Blastomyces dermatitidis

Serum factors, including mannose binding lectins (MBL), influence innate responses to microbes. Little is known about the effects of serum factors or MBL on the interaction of Blastomyces dermatitidis, a pulmonary fungal pathogen, with macrophages or on tumor necrosis factor alpha (TNF-α) production. Since macrophage production of TNF-α is an important innate immune response, we examined a mouse peritoneal macrophage (PM) cell line (RAW) and resident PM from CD-1 mice to study TNF-α production by PM stimulated with heat-killed (HK) or live B. dermatitidis yeast cells. Mouse serum and heat-inactivated mouse serum inhibited TNF-α production 94% when macrophages were stimulated by B. dermatitidis, whereas mouse immunoglobulin G (IgG) did not have this effect. HK B. dermatitidis incubated with serum and then washed also failed to stimulate significant TNF-α production by PM. By the sandwich immunofluorescent antibody (IFA) method with anti-mouse MBL (MBL-A or -C), we showed that serum MBL bound to B. dermatitidis. When serum was absorbed with HK B. dermatitidis or live B. dermatitidis, absorbed serum failed to significantly inhibit TNF-α production by RAW cells plus B. dermatitidis, and immunoblotting showed that absorbed serum was depleted of MBL-C. If serum was absorbed with live B. dermatitidis, unbound serum was eluted, and bound serum factor(s) (BS) was released with guanidine buffer, BS inhibited TNF-α production by PM plus B. dermatitidis in a concentration-dependent manner. BS contained MBL-C, which bound B. dermatitidis, as shown by IFA assay. 1,3-β-Glucan stimulated TNF-α production by PM, and this was inhibited by mouse serum. Treatment of B. dermatitidis with anti-1,3-β-glucan antibody inhibited TNF-α production by PM. With anti-1,3-β-glucan antibody, we showed by IFA assay that B. dermatitidis contained 1,3-β-glucan. In an IFA study with B. dermatitidis, serum with an anti-mouse IgG conjugate did not result in fluorescence, yet serum blocked IFA staining of B. dermatitidis by anti-1,3-β-glucan IgG antibody. This indicated that non-IgG serum factors binding to B. dermatitidis prevented access to 1,3-β-glucan by anti-1,3-β-glucan antibody. These results suggest that the mechanism of inhibition of the innate proinflammatory immune response of PM to B. dermatitidis is mediated by serum MBL binding to B. dermatitidis at 1,3-β-glucan sites or sterically masking 1,3-β-glucan sites, thus preventing 1,3-β-glucan stimulation of PM for TNF-α production.