Papers by Yves Laumonnier
Molecular Immunology, Aug 1, 2011
Journal of Thrombosis and Haemostasis, Oct 1, 2006
We have previously demonstrated that plasmin acts as potent proinflammatory activator of human pe... more We have previously demonstrated that plasmin acts as potent proinflammatory activator of human peripheral monocytes leading to activation of protein kinase cascades and a number of transcription factors. An uncompromised proteolytic activity was essential for the plasmin-mediated monocyte activation pointing to the necessity of a proteolytic activation of the signaling receptor. Despite considerable efforts, little progress has been made so far in the identification of a signal-transducing plasmin receptor. Here we identify the annexin A2 heterotetramer, composed of annexin A2 and S100A10, as receptor for the plasmin-induced signaling in human monocytes. Monocytes express the annexin A2 heterotetramer on the cell surface as shown by i) flow cytometry, ii) fluorescence microscopy, and iii) co-immunoprecipitation of biotinylated cell surface proteins. Binding of plasmin to annexin A2 and S100A10 on monocytes was verified by biotin transfer from plasmin labeled with a trifunctional crosslinker. Antibodies directed against annexin A2 or S100A10 inhibited the chemotaxis elicited by plasmin, but not that induced by the standard chemoattractant fMLP. A control antibody directed against the thrombin receptor PAR-1 had no effect on the plasmin-mediated chemotaxis suggesting that PAR-1 is not involved. Further, downregulation of the annexin A2 or S100A10 expression in monocytes by antisense oligodeoxynucleotides impaired the chemotactic response to plasmin, but not that to fMLP. Antisense oligodeoxynucleotides similarly decreased the TNF-alpha release by plasmin-but not by LPS-stimulated monocytes. At the molecular level, stimulation with plasmin, but not with catalytically inactivated plasmin, induced cleavage of annexin A2 and dissociation of the annexin A2 heterotetramer complex. In contrast to annexin A2, S100A10 is not a substrate for plasmin. Substitution of lysine to alanine in position 27 abolished the cleavage of recombinant annexin A2 in vitro indicating a single cleavage site. These data shed new light on the plasmin-induced signaling; taken together they identify the annexin A2 heterotetramer in human monocytes as a signaling receptor activated by plasmin via proteolysis.
Immunological Reviews, Oct 26, 2016
The activation of the complement system by canonical and non-canonical mechanisms results in the ... more The activation of the complement system by canonical and non-canonical mechanisms results in the generation of multiple C3 and C5 cleavage fragments including anaphylatoxins C3a and C5a as well as opsonizing C3b/iC3b. It is now well appreciated that anaphylatoxins not only act as pro-inflammatory mediators but as immunoregulatory molecules that control the activation status of cells and tissue at several levels. Likewise, C3b/iC3b is more than the opsonizing fragment that facilitates engulfment and destruction of targets by phagocytes. In the circulation, it also facilitates the transport and delivery of bacteria and immune complexes to phagocytes, through a process known as immune adherence, with consequences for adaptive immunity. Here, we will discuss non-classical immunoregulatory properties of C3 and C5 cleavage fragments. We highlight the influence of anaphylatoxins on Th2 and Th17 cell development during allergic asthma with a particular emphasis on their role in the modulation of CD11b(+) conventional dendritic cells and monocyte-derived dendritic cells. Furthermore, we discuss the control of anaphylatoxin-mediated activation of dendritic cells and allergic effector cells by adaptive immune mechanisms that involve allergen-specific IgG1 antibodies and plasma or regulatory T cell-derived IL-10 production. Finally, we take a fresh look at immune adherence with a particular focus on the development of antibacterial cytotoxic T-cell responses.
Frontiers in Immunology, Mar 23, 2016
Allergic asthma is a chronic inflammatory disease of the airways that is driven by maladaptive T ... more Allergic asthma is a chronic inflammatory disease of the airways that is driven by maladaptive T helper 2 (Th2) and Th17 immune responses against harmless, airborne substances. Pulmonary phagocytes represent the first line of defense in the lung where they constantly sense the local environment for potential threats. They comprise two distinct cell types, i.e., macrophages and dendritic cells (DC) that differ in their origins and functions. Alveolar macrophages quickly take up most of the inhaled allergens, yet do not deliver their cargo to naive T cells sampling in draining lymph nodes. In contrast, pulmonary DCs instruct CD4 + T cells develop into Th2 and Th17 effectors, initiating the maladaptive immune responses toward harmless environmental substances observed in allergic individuals. Unraveling the mechanisms underlying this mistaken identity of harmless, airborne substances by innate immune cells is one of the great challenges in asthma research. The identification of different pulmonary DC subsets, their role in antigen uptake, migration to the draining lymph nodes, and their potential to instruct distinct T cell responses has set the stage to unravel this mystery. However, at this point, a detailed understanding of the spatiotemporal resolution of DC subset localization, allergen uptake, processing, autocrine and paracrine cellular crosstalk, and the humoral factors that define the activation status of DCs is still lacking. In addition to DCs, at least two distinct macrophage populations have been identified in the lung that are either located in the airway/alveolar lumen or in the interstitium. Recent data suggest that such populations can exert either pro-or anti-inflammatory functions. Similar to the DC subsets, detailed insights into the individual roles of alveolar and interstitial macrophages during the different phases of asthma development are still missing. Here, we will provide an update on the current understanding of the origin, localization, and function of the diverse pulmonary antigen-presenting cell subsets, in particular with regard to the development and regulation of allergic asthma. While most data are from mouse models of experimental asthma, we have also included available human data to judge the translational value of the findings obtained in experimental asthma models.
Journal of Immunology, Dec 1, 2011
C3a desArg/Acylation stimulating protein (ASP) and C5L2 correlation with expression of in ammator... more C3a desArg/Acylation stimulating protein (ASP) and C5L2 correlation with expression of in ammatory factors and metabolic factors in human adipose tissue (94.25)
Journal of Biological Chemistry, Dec 1, 2000
The endothelial nitric-oxide synthase gene is constitutively expressed in endothelial cells. Seve... more The endothelial nitric-oxide synthase gene is constitutively expressed in endothelial cells. Several transcriptionally active regulatory elements have been identified in the proximal promoter, including a GATA-2 and an Sp-1 binding site. Because they cannot account for the constitutive expression of endothelial nitric-oxide synthase gene in a restricted number of cells, we have searched for other cell-specific regulatory elements. By DNase I hypersensitivity mapping and deletion studies we have identified a 269-base pair activator element located 4.9 kilobases upstream from the transcription start site that acts as an enhancer. DNase I footprinting and linker-scanning experiments showed that several regions within the 269-base pair enhancer are important for transcription factor binding and for full enhancer activity. The endothelial specificity of this activation seems partly due to interaction between this enhancer in its native configuration and the promoter in endothelial cells. EMSA experiments suggested the implication of MZF-like, AP-2, Sp-1-related, and Ets-related factors. Among Ets factors, Erg was the only one able to bind to cognate sites in the enhancer, as found by EMSA and supershift experiments, and to activate the transcriptional activity of the enhancer in cotransfection experiments. Therefore, multiple protein complexes involving Erg, other Ets-related factors, AP-2, Sp-1-related factor, and MZF-like factors are important for the function of this enhancer in endothelial cells.
Journal of Immunology, May 1, 2017
Eosinophils are considered a homogenous cell population, contributing to the allergic asthma phen... more Eosinophils are considered a homogenous cell population, contributing to the allergic asthma phenotype by promoting mucus hypersecretion and airway hyperresponsiveness. Here, we identified a previously unrecognized vacuolated CD11cdim eosinophilic population (vEOS) which accmulates exclusively in the lung tissue and in mediastinal lymph nodes but not in the airways in a house dust mite-mediated allergic asthma model and an IL-33 mediated pulmonary inflammation. In addition to the vEOS, we found classical SiglecF+CD11c− eosinophils (EOS) in the airways and the lung. Both cell types shared typical structural features of eosinophils and expressed CCR3. In contrast to EOS, vEOS expressed higher levels of the CD11a, CD11b and CD18 integrins. Immunofluorescence microscopy revealed that vEOS were located exclusively near arteries and airways, where they were in close contact with T cells. Moreover, vEOS took up antigen and expressed costimulatory molecules more efficiently than EOS. Consequently, in co-culture experiments with OVA-TCR transgenic T cells, vEOS and dendritic cells (DCs) drove T cell proliferation. However, in the presence of vEOS, T cells divided more frequently than in co-culture with DCs. Further, T cell activation in response to co-culture with vEOS together with DCs markedly enhanced IL-17 but not IL-13 production. Collectively, our data demonstrate that vEOS are an important subset of eosinophils, which differ from EOS in their location and their ability to activate T cells. Further, they suggest that cross-talk between vEOS and DCs might be a crucial step for the induction of Th17 cells in allergic asthma. Thus, vEOS may serve as an important target for Th17-associated steroid-resistant asthma.
The FASEB Journal, Mar 1, 2008
Acta pharmacologica Sinica, Oct 3, 2011
Aim: To identify proteins that interact with the C-terminal fragment of annexin A2 (A2IC), genera... more Aim: To identify proteins that interact with the C-terminal fragment of annexin A2 (A2IC), generated by plasmin cleavage of the plasmin receptor, a heterotetramer (AA2t) containing annexin A2. Methods: The gene that encodes the A2IC fragment was obtained from PCR-amplified cDNA isolated from human monocytes, and was ligated into the pBTM116 vector using a DNA ligation kit. The resultant plasmid (pBTM116-A2IC) was sequenced with an ABI PRISM 310 Genetic Analyzer. The expression of an A2IC bait protein fused with a LexA-DNA binding domain (BD) was determined using Western blot analysis. The identification of proteins that interact with A2IC and are encoded in a human monocyte cDNA library was performed using yeast two-hybrid screening. The DNA sequences of the relevant cDNAs were determined using an ABI PRISM BigDye terminator cycle sequencing ready reaction kit. Nucleotide sequence databases were searched for homologous sequences using BLAST search analysis (http://www.ncbi.nlm.nih.gov). Confirmation of the interaction between the protein LexA-A2IC and each of cathepsin S and SNX17 was conducted using a small-scale yeast transformation and X-gal assay. Results: The yeast transformed with plasmids encoding the bait proteins were screened with a human monocyte cDNA library by reconstituting full-length transcription factors containing the GAL4-active domain (GAL4-AD) as the prey in a yeast two-hybrid approach. After screening 1×10 7 clones, 23 independent β-Gal-positive clones were identified. Sequence analysis and a database search revealed that 15 of these positive clones matched eight different proteins (SNX17, ProCathepsin S, RPS2, ZBTB4, OGDH, CCDC32, PAPD4, and actin which was already known to interact with annexin A2). Conclusion: A2IC A2IC interacts with various proteins to form protein complexes, which may contribute to the molecular mechanism of monocyte activation induced by plasmin. The yeast two-hybrid system is an efficient approach for investigating protein interactions.
Cells, Jan 26, 2020
Activation of the C5/C5a/C5a receptor 1 (C5aR1) axis during allergen sensitization protects from ... more Activation of the C5/C5a/C5a receptor 1 (C5aR1) axis during allergen sensitization protects from maladaptive T cell activation. To explore the underlying regulatory mechanisms, we analyzed the impact of C5aR1 activation on pulmonary CD11b + conventional dendritic cells (cDCs) in the context of house-dust-mite (HDM) exposure. BALB/c mice were intratracheally immunized with an HDM/ovalbumin (OVA) mixture. After 24 h, we detected two CD11b + cDC populations that could be distinguished on the basis of C5aR1 expression. C5aR1 − but not C5aR1 + cDCs strongly induced T cell proliferation of OVA-reactive transgenic CD4 + T cells after re-exposure to antigen in vitro. C5aR1 − cDCs expressed higher levels of MHC-II and CD40 than their C5aR1 + counterparts, which correlated directly with a higher frequency of interactions with cognate CD4 + T cells. Priming of OVA-specific T cells by C5aR1 + cDCs could be markedly increased by in vitro blockade of C5aR1 and this was associated with increased CD40 expression. Simultaneous blockade of C5aR1 and CD40L on C5aR1 + cDCs decreased T cell proliferation. Finally, pulsing with OVA-induced C5 production and its cleavage into C5a by both populations of CD11b + cDCs. Thus, we propose a model in which allergen-induced autocrine C5a generation and subsequent C5aR1 activation in pulmonary CD11b + cDCs promotes tolerance towards aeroallergens through downregulation of CD40.
The Journal of Allergy and Clinical Immunology, Jun 1, 2012
Background: Under inflammatory conditions, T cell-dependent (TD) protein antigens induce proinfla... more Background: Under inflammatory conditions, T cell-dependent (TD) protein antigens induce proinflammatory T-and B-cell responses. In contrast, tolerance induction by TD antigens without costimulation triggers the development of regulatory T cells. Under both conditions, IgG antibodies are generated, but whether they have different immunoregulatory functions remains elusive. Objective: It was shown recently that proinflammatory or anti-inflammatory effector functions of IgG molecules are determined by different Fc N-linked glycosylation patterns. We sought to examine the Fc glycosylation and anti-inflammatory quality of IgG molecules formed on TD tolerance induction. Methods: We administered chicken ovalbumin (OVA) with or without costimulus to mice and analyzed OVA-reactive IgG Fc glycosylation. The anti-inflammatory function of differentially glycosylated anti-OVA IgGs was further investigated in studies with dendritic cell cultures and in an in vivo model of allergic airway disease. Additionally, we analyzed the Fc glycosylation pattern of birch pollen-reactive serum IgGs after successful allergen-specific immunotherapy in patients. Results: Stimulation with TD antigens under inflammatory conditions induces plasma cells expressing low levels of a2,6-sialyltransferase and producing desialylated IgGs. In contrast, plasma cells induced on tolerance induction did not downregulate a2,6-sialyltransferase expression and secreted immunosuppressive sialylated IgGs that were sufficient to block antigen-specific T-and B-cell responses, dendritic cell maturation, and allergic airway inflammation. Importantly, successful specific immunotherapy in allergic patients also induced sialylated allergen-specific IgGs. Conclusions: Our data show a novel antigen-specific immunoregulatory mechanism mediated by anti-inflammatory sialylated IgGs that are formed on TD tolerance induction. These findings might help to develop novel antigen-specific therapies for the treatment of allergy and autoimmunity.
Journal of Immunology, May 1, 2017
The anaphylatoxin C3a exerts its biologic functions through the activation of its cognate G-prote... more The anaphylatoxin C3a exerts its biologic functions through the activation of its cognate G-protein-coupled receptor C3a receptor (C3aR). C3aR mRNA expression has been described both in human and mouse lungs. However, no systematic analysis of C3aR expression at steady state and under pro-inflammatory conditions in the different pulmonary cell subsets has been done so far. Using a newly developed Tandem-dye (td)-Tomato-C3aR reporter knock-in mouse, we found C3aR expression in pulmonary eosinophils, resident CD11b + conventional dendritic cells (cDC), and monocyte-derived DCs (moDC) at steady state. In contrast, pulmonary CD103 + cDCs and plasmacytoid DCs stained negative. Surprisingly, lung and bronchoalveolar alveolar macrophages (AMs), as well as neutrophils, lacked td-Tomato-C3aR. C3aR antibody staining revealed that only moDCs express the receptor at the cell surface. House dust mite (HDM) is a potent allergen that drives the development of allergic asthma in humans. Using the td-Tomato-C3aR reporter mouse, we found that HDM exposure resulted in the upregulation of C3aR in AMs, eosinophils, CD11b + cDCs, and moDCs during the early phase of HDM-induced allergic asthma. Further, C3aR translocated to the cell surface in eosinophils and AMs following HDM exposure. Importantly, intra-tracheal administration of IL-33, an alarmin released by epithelial cells upon allergen contact, recapitulated these results. In summary, our findings demonstrate that allergen contact triggers the upregulation of C3aR and its surface expression in previously negative innate immune cells from the lung by an IL-33 dependent mechanism. C3aR-mediated activation of innate immune cells may play a critical role for early asthma development.
Immunobiology, Oct 1, 2016
Immunobiology, Oct 1, 2016
Immunobiology, Oct 1, 2016
Molecular Immunology, Aug 1, 2011
Frontiers in Immunology, May 16, 2018
The division of labor between pulmonary phagocytic subsets [macrophage/monocyte and dendritic cel... more The division of labor between pulmonary phagocytic subsets [macrophage/monocyte and dendritic cell (DC) subpopulations] has been described at the functional level. However, whether these lung phagocytes also display unique spatial distribution remains unclear. Here, to analyze cellular distribution in lung compartments and contacts between phagocyte subpopulations, we established an immunohistochemistry (IHC)-based method to clearly identify murine lung phagocyte subsets in situ based on differential expression of CD11c, CD11b, MHC-II, Langerin and mPDCA-1. Furthermore, we investigated subset-specific functional differences in antigen uptake and spatial changes upon allergic sensitization. Our staining allowed the distinction between alveolar macrophages (AMs), interstitial macrophage (IM) subpopulations, CD11b + DC subpopulations, CD103 + DCs, and plasmacytoid DCs (pDCs). We identified interstitial regions between airways and around airways as regions of IM/CD11b + DC/CD103 + DC clusters, where a subset of IMs (IM2) and CD103 + DCs formed intense contacts that decreased upon allergic sensitization. These data indicate functional interactions between both cell types either in steady state or after antigen encounter affecting the development of allergies or tolerance. Furthermore, we observed major antigen uptake in AMs and IMs rather than DC subpopulations that was not restricted to airways and adjacent areas. This will enable to focus future studies to immunologically relevant cellular interactions and to unravel which cells are tipping the balance between pro-inflammatory immune responses or tolerance.
Journal of Thrombosis and Thrombolysis, Feb 14, 2010
Human cytomegalovirus (HCMV) establishes a life-long persistent infection. HCMV infection could b... more Human cytomegalovirus (HCMV) establishes a life-long persistent infection. HCMV infection could be associated with chronic inflammatory diseases, such as cardiovascular disease and atherosclerosis. Here we observed that in HCMV (AD-169) pre-exposed human umbilical vein endothelial cells (HUVEC), thrombin-induced expression of IL-1alpha and M-CSF is markedly enhanced compared to the un-exposed cells. Study of the expression of thrombin receptor genes in HUVEC showed that HCMV triggered a time- and concentration-dependent expression of the thrombin receptors PAR1, PAR3 and PAR4 at the mRNA level. Induction of PAR1 and PAR3 mRNA expression is due to transcriptional activation of their promoters as shown by gene reporter assay. Furthermore, the virus induced expression of PAR1 and PAR3 but not PAR4 proteins, as analyzed by Western immunoblotting. However, flow cytometric analysis revealed that only PAR3, expressed at very low level in control HUVEC, is induced at the surface during the exposure to the virus. Our data suggest that although exposure to HCMV induces a minor increase of cell-surface receptors expression, it does make endothelial cells more responsive to additional thrombin stimulation.
Genes & Development, Sep 29, 2005
The c-Myb transcription factor coordinates proliferation and differentiation of hematopoietic pre... more The c-Myb transcription factor coordinates proliferation and differentiation of hematopoietic precursor cells. Myb has three consecutive N-terminal SANT-type repeat domains (R1, R2, R3), two of which (R2, R3) form the DNA-binding domain (DBD). Three amino acid substitutions in R2 alter the way Myb regulates genes and determine the leukemogenicity of the retrovirally transduced v-Myb oncogene. The molecular mechanism of how these mutations unleash the leukemogenic potential of Myb is unknown. Here we demonstrate that the c-Myb-DBD binds to the N-terminal histone tails of H3 and H3.3. C-Myb binding facilitates histone tail acetylation, which is mandatory during activation of prevalent differentiation genes in conjunction with CCAAT enhancer-binding proteins (C/EBP). Leukemogenic mutations in v-Myb eliminate the interaction with H3 and acetylation of H3 tails and abolish activation of endogenous differentiation genes. In primary v-myb-transformed myeloblasts, pharmacologic enhancement of H3 acetylation restored activation of differentiation genes and induced cell differentiation. Our data link a novel chromatin function of c-Myb with lineage-specific expression of differentiation genes and relate the loss of this function with the leukemic conversion of Myb.
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Papers by Yves Laumonnier