Search

Contains fulltext : 172623.pdf (Publisher’s version ) (Closed access) Coxiella burnetii, the causative agent of Q fever, is recognized by TLR2. TLR10 can act as an inhibitory receptor on TLR2-derived immune responses. Therefore, we investigated the role of TLR10 on C. burnetii-induced cytokine production and assessed whether genetic polymorphisms in TLR10 influences the development of chronic Q fever. HEK293 cells, transfected with TLR2, TLR10 or TLR2/TLR10, and human peripheral blood mononuclear cells (PBMCs) in the presence of anti-TLR10, were stimulated with C. burnetii. In both assays, the absence of TLR10 resulted in increased cytokine responses after C. burnetii stimulation. In addition, the effect of single nucleotide polymorphisms (SNPs) in TLR10 was examined in healthy volunteers whose PBMCs were stimulated with C. burnetii Nine Mile or the Dutch outbreak isolate C. burnetii 3262. Individuals bearing SNPs in TLR10 displayed increased cytokine production upon C. burnetii 3262 stimulation. Furthermore, 139 chronic Q fever patients and 220 controls were genotyped for TLR10 N241H, I775V and I369L. None of these polymorphisms were associated with increased susceptibility to chronic Q fever. In conclusion, TLR10 has an inhibitory effect on in vitro cytokine production by C. burnetii, but the presence of TLR10 polymorphisms does not lead to an increased risk of developing chronic Q fever.

© The Author(s) 2018. Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/., Invasive aspergillosis (IA) is a severe infection that can occur in severely immunocompromised patients. Efficient immune recognition of Aspergillus is crucial to protect against infection, and previous studies suggested a role for NOD2 in this process. However, thorough investigation of the impact of NOD2 on susceptibility to aspergillosis is lacking. Common genetic variations in NOD2 has been associated with Crohn's disease and here we investigated the influence of these genetic variations on the anti-Aspergillus host response. A NOD2 polymorphism reduced the risk of IA after hematopoietic stem-cell transplantation. Mechanistically, absence of NOD2 in monocytes and macrophages increases phagocytosis leading to enhanced fungal killing, conversely, NOD2 activation reduces the antifungal potential of these cells. Crucially, Nod2 deficiency results in resistance to Aspergillus infection in an in vivo model of pulmonary aspergillosis. Collectively, our data demonstrate that genetic deficiency of NOD2 plays a protective role during Aspergillus infection., F.L.v.d.V. was supported by the E-rare project EURO-CMC. M.O. was supported by the NWO, 016.176.006). A.C. and C.C. were supported by the Northern Portugal Regional Operational Programme (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (FEDER) (NORTE-01-0145-FEDER-000013), and the Fundação para a Ciência e Tecnologia (FCT) (IF/00735/2014 to A.C. and SFRH/BPD/96176/2013 to C.C.).

Contains fulltext : 174719.pdf (Publisher’s version ) (Closed access) Coxiella burnetii is the causative agent of Q fever. Although the prevalence of C. burnetii in cattle is much higher than in goats and sheep, infected cattle are rarely associated with human outbreaks. We investigated whether the immune response of humans differs after contact with C. burnetii isolates from different host origins or with different multilocus variable number of tandem repeat analysis (MLVA) genotypes. Cytokine responses were measured in human peripheral blood mononuclear cells (PBMCs) stimulated with 16 C. burnetii isolates with known MLVA genotype from goats, sheep, cattle, acute and chronic Q fever patients. Coxiella burnetii isolates originating from cattle induce significantly more IL-1beta, TNF-alpha and IL-22 than the isolates from goats, sheep or chronic Q fever patients. Comparing the cytokine induction of the isolates based on their MVLA genotype did not reveal differences in response between the MLVA genotypes. The proinflammatory cytokine response induced in human PBMCs by C. burnetii isolates from cattle may explain the low incidence of human Q fever outbreaks caused by cattle. The cytokine profile of PBMCs stimulated with C. burnetii isolates from chronic Q fever patients resembles isolates from goats. Furthermore, cytokine responses seem to be depending on host origin than on MLVA genotype.

Contains fulltext : 171426.pdf (Publisher’s version ) (Open Access) Cells in homeostasis metabolize glucose mainly through the tricarboxylic acid cycle and oxidative phosphorylation, while activated cells switch their basal metabolism to aerobic glycolysis. In this study, we examined whether metabolic reprogramming toward aerobic glycolysis is important for the host response to Mycobacterium tuberculosis (Mtb). Through transcriptional and metabolite analysis we show that Mtb induces a switch in host cellular metabolism toward aerobic glycolysis in human peripheral blood mononuclear cells (PBMCs). The metabolic switch is TLR2 dependent but NOD2 independent, and is mediated in part through activation of the AKT-mTOR (mammalian target of rapamycin) pathway. We show that pharmacological inhibition of the AKT/mTOR pathway inhibits cellular responses to Mtb both in vitro in human PBMCs, and in vivo in a model of murine tuberculosis. Our findings reveal a novel regulatory layer of host responses to Mtb that will aid understanding of host susceptibility to Mtb, and which may be exploited for host-directed therapy.

Chitin is an important cell wall component of Aspergillus fumigatus conidia, of which hundreds are inhaled on a daily basis. Previous studies have shown that chitin has both anti- and proinflammatory properties; however the exact mechanisms determining the inflammatory signature of chitin are poorly understood, especially in human immune cells. Human peripheral blood mononuclear cells were isolated from healthy volunteers and stimulated with chitin from Aspergillus fumigatus. Transcription and production of the proinflammatory cytokine interleukin-1β (IL-1β) and the anti-inflammatory cytokine IL-1 receptor antagonist (IL-1Ra) were measured from the cell culture supernatant by quantitative PCR (qPCR) or enzyme-linked immunosorbent assay (ELISA), respectively. Chitin induced an anti-inflammatory signature characterized by the production of IL-1Ra in the presence of human serum, which was abrogated in immunoglobulin-depleted serum. Fc-γ-receptor-dependent recognition and phagocytosis of IgG-opsonized chitin was identified as a novel IL-1Ra-inducing mechanism by chitin. IL-1Ra production induced by chitin was dependent on Syk kinase and phosphatidylinositol 3-kinase (PI3K) activation. In contrast, costimulation of chitin with the pattern recognition receptor (PRR) ligands lipopolysaccharide, Pam3Cys, or muramyl dipeptide, but not β-glucan, had synergistic effects on the induction of proinflammatory cytokines by human peripheral blood mononuclear cells (PBMCs). In conclusion, chitin can have both pro- and anti-inflammatory properties, depending on the presence of pathogen-associated molecular patterns and immunoglobulins, thus explaining the various inflammatory signatures reported for chitin., IMPORTANCE Invasive aspergillosis and allergic aspergillosis are increasing health care problems. Patients get infected by inhalation of the airborne spores of Aspergillus fumigatus. A profound knowledge of how Aspergillus and its cell wall components are recognized by the host cell and which type of immune response it induces is necessary to develop target-specific treatment options with less severe side effects than the treatment options to date. There is controversy in the literature about the receptor for chitin in human cells. We identified the Fc-γ receptor and Syk/PI3K pathway via which chitin can induce anti-inflammatory immune responses by inducing IL-1 receptor antagonist in the presence of human immunoglobulins but also proinflammatory responses in the presence of bacterial components. This explains why Aspergillus does not induce strong inflammation just by inhalation and rather fulfills an immune-dampening function. While in a lung coinfected with bacteria, Aspergillus augments immune responses by shifting toward a proinflammatory reaction.

Contains fulltext : 153993.pdf (Publisher’s version ) (Open Access) BACKGROUND: Antibiotic treatment of chronic Q fever is cumbersome and of long duration. To monitor treatment, there is a need for alternative biomarkers. Coxiella burnetii-specific interferon (IFN)-gamma and interleukin (IL)-2 production reflect the type of effector and memory T-cell response. In chronic Q fever, C. burnetii-specific IFN-gamma production is higher and IL-2 production is lower than in individuals with past Q fever. Here we explore whether C. burnetii-specific IFN-gamma and IL-2 production correlate to treatment response. METHODS: We studied the longitudinal C. burnetii-specific IFN-gamma/IL-2 ratio in fifteen proven chronic Q fever patients. All patients were followed for at least 18 months during antibiotic treatment. Treatment was considered successful when clinical recovery was observed, a positive PCR for C. burnetii DNA in blood became persistently negative, anti-phase I IgG showed a fourfold decrease or more, and imaging techniques showed disappearance of infectious foci. RESULTS: Overall, the IFN-gamma/IL-2 ratio declined when patients experienced a successful treatment outcome. When treatment failed, IFN-gamma/IL-2 ratios did not significantly decrease. The median (+/-IQR) slope of the longitudinal IFN-gamma/IL-2 ratio with successful treatment was -2.10 (-7.02 to -0.06), and -0.15 (-1.13 to 0.25) with unsuccessful treatment (P = 0.19). Q fever endocarditis patients had higher IFN-gamma/IL-2 ratios than patients with endovascular infections. CONCLUSION: We propose that the IFN-gamma/IL-2 ratio can be used as an additional biomarker for monitoring chronic Q fever treatment, with declining ratios being indicative of successful treatment.

Contains fulltext : 153819.pdf (Publisher’s version ) (Closed access) BACKGROUND: Infection with Coxiella burnetii can lead to acute and chronic Q fever. Toll-like receptor 1 (TLR1), TLR2, TLR4, TLR6, nucleotide-binding oligomerization domain receptor 1 (NOD1), NOD2, and the mitogen-activated protein kinases are central in the innate immune response against microorganisms, but little is known about their role in the recognition of C. burnetii in humans. METHODS: Human peripheral blood mononuclear cells (PBMCs) were stimulated with C. burnetii Nine Mile and the Dutch outbreak isolate C. burnetii 3262. TLRs were inhibited using specific antibodies or antagonists. Additionally, the influence of human polymorphisms in TLRs and Nod-like receptors (NLRs) on C. burnetii-induced cytokine production was assessed. RESULTS: Inhibition of TLR2, p38, JNK, and ERK led to decreased cytokine responses in C. burnetii-stimulated human PBMCs. Humans with polymorphisms in TLR1 and NOD2 had reduced cytokine production, compared with humans with wild-type genotypes, after stimulation. Interestingly, polymorphisms in TLR6 led to decreased cytokine production after C. burnetii 3262 stimulation but not after C. burnetii Nine Mile stimulation. CONCLUSIONS: The TLR1/TLR2 heterodimer and NOD2 are important recognition receptors for the induction of cytokine responses against C. burnetii in humans. Furthermore, an interesting finding was the divergent recognition of C. burnetii Nine Mile and C. burnetii 3262.

Contains fulltext : 155344.pdf (Publisher’s version ) (Closed access) BACKGROUND: Q fever is an infection caused by Coxiella burnetii. Persistent infection (chronic Q fever) develops in 1%-5% of patients. We hypothesize that inefficient recognition of C. burnetii and/or activation of host-defense in individuals carrying genetic variants in pattern recognition receptors or adaptors would result in an increased likelihood to develop chronic Q fever. METHODS: Twenty-four single-nucleotide polymorphisms in genes encoding Toll-like receptors, nucleotide-binding oligomerization domain-like receptor-2, alphavbeta3 integrin, CR3, and adaptors myeloid differentiation primary response protein 88 (MyD88), and Toll interleukin 1 receptor domain-containing adaptor protein (TIRAP) were genotyped in 139 patients with chronic Q fever and in 220 controls with cardiovascular risk-factors and previous exposure to C. burnetii. Associations between these single-nucleotide polymorphisms and chronic Q fever were assessed by means of univariate logistic regression models. Cytokine production in whole-blood stimulation assays was correlated with relevant genotypes. RESULTS: Polymorphisms in TLR1 (R80T), NOD2 (1007fsX1), and MYD88 (-938C>A) were associated with chronic Q fever. No association was observed for polymorphisms in TLR2, TLR4, TLR6, TLR8, ITGAV, ITGB3, ITGAM, and TIRAP. No correction for multiple testing was performed because only genes with a known role in initial recognition of C. burnetii were included. In the whole-blood assays, individuals carrying the TLR1 80R-allele showed increased interleukin 10 production with C. burnetii exposure. CONCLUSIONS: Polymorphisms in TLR1 (R80T), NOD2 (L1007fsX1), and MYD88 (-938C>A) are associated with predisposition to development of chronic Q fever. For TLR1, increased interleukin 10 responses to C. burnetii in individuals carrying the risk allele may contribute to the increased risk of chronic Q fever.

Contains fulltext : 138727.pdf (Publisher’s version ) (Open Access) In humans, infection with Coxiella burnetii, the causative agent of Q fever, leads to acute or chronic infection, both associated with specific clinical symptoms. In contrast, no symptoms are observed in goats during C. burnetii infection, although infection of the placenta eventually leads to premature delivery, stillbirth and abortion. It is unknown whether these differences in clinical outcome are due to the early immune responses of the goats. Therefore, peripheral blood mononuclear cells (PBMCs) were isolated from pregnant goats. In total, 17 goats were included in the study. Six goats remained naive, while eleven goats were infected with C. burnetii. Toll-like receptor (TLR) and cytokine mRNA expression were measured after in vitro stimulation with heat-killed C. burnetii at different time points (prior infection, day 7, 35 and 56 after infection). In naive goats an increased expression of interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha, IL-10 and interferon (IFN)-gamma mRNA upon C. burnetii stimulation was detected. In addition, TLR2 expression was strongly up-regulated. In goats infected with C. burnetii, PBMCs re-stimulated in vitro with C. burnetii, expressed significantly more TNF-alpha mRNA and IFN-gamma mRNA compared to naive goats. In contrast, IL-10 mRNA production capacity was down-regulated during C. burnetii infection. Interestingly, at day 7 after inoculation a decreased IFN-gamma protein level was observed in stimulated leukocytes in whole blood from infected goats, whereas at other time-points increased production of IFN-gamma protein was seen. Our study shows that goats initiate a robust pro-inflammatory immune response against C. burnetii in vitro. Furthermore, PBMCs from C. burnetii infected goats show augmented pro-inflammatory cytokine responses compared to PBMCs from non-infected goats. However, despite this pro-inflammatory response, goats are not capable of clearing the C. burnetii infection.

Contains fulltext : 137608.pdf (Publisher’s version ) (Closed access) Infection with Coxiella burnetii may lead to life-threatening chronic Q fever endocarditis or vascular infections, which are often difficult to diagnose. The present study aims to investigate whether measurement of in-vitro interferon-gamma (IFN-gamma) production, a key cytokine in the immune response against C. burnetii, differentiates chronic from a past cleared infection, and whether measurement of other cytokines would improve the discriminative power. First, C. burnetii-specific IFN-gamma production was measured in whole blood of 28 definite chronic Q fever patients and compared with 135 individuals with past Q fever (seropositive controls) and 908 seronegative controls. IFN-gamma production was significantly higher in chronic Q fever patients than in controls, but with overlapping values between patients and seropositives. Secondly, the production of a series of other cytokines was measured in a subset of patients and controls, which showed that interleukin (IL)-2 production was significantly lower in patients than in seropositive controls. Subsequently, measuring IL-2 in all patients and all controls with substantial IFN-gamma production showed that an IFN-gamma/IL-2 ratio >11 had a sensitivity and specificity of 79% and 96%, respectively, to diagnose chronic Q fever. This indicates that a high IFN-gamma/IL-2 ratio is highly suggestive for chronic Q fever. In an additional group of 25 individuals with persistent high anti-Coxiella phase I IgG titres without definite chronic infection, all but six showed an IFN-gamma/IL-2 ratio