Publications by authors named "Christophe Combadière"

102 Publications

Chemokine Receptor 2-targeted Molecular Imaging in Pulmonary Fibrosis. A Clinical Trial.

Am J Respir Crit Care Med 2021 01;203(1):78-89

Department of Radiology.

Idiopathic pulmonary fibrosis (IPF) is a progressive inflammatory lung disease without effective molecular markers of disease activity or treatment responses. Monocyte and interstitial macrophages that express the C-C motif CCR2 (chemokine receptor 2) are active in IPF and central to fibrosis. To phenotype patients with IPF for potential targeted therapy, we developed Cu-DOTA-ECL1i, a radiotracer to noninvasively track CCR2 monocytes and macrophages using positron emission tomography (PET). CCR2 cells were investigated in mice with bleomycin- or radiation-induced fibrosis and in human subjects with IPF. The CCR2 cell populations were localized relative to fibrotic regions in lung tissue and characterized using immunolocalization, single-cell mass cytometry, and RNA hybridization and then correlated with parallel quantitation of lung uptake by Cu-DOTA-ECL1i PET. Mouse models established that increased Cu-DOTA-ECL1i PET uptake in the lung correlates with CCR2 cell infiltration associated with fibrosis ( = 72). As therapeutic models, the inhibition of fibrosis by IL-1β blockade ( = 19) or antifibrotic pirfenidone ( = 18) reduced CCR2 macrophage accumulation and uptake of the radiotracer in mouse lungs. In lung tissues from patients with IPF, CCR2 cells concentrated in perifibrotic regions and correlated with radiotracer localization ( = 21). Human imaging revealed little lung uptake in healthy volunteers ( = 7), whereas subjects with IPF ( = 4) exhibited intensive signals in fibrotic zones. These findings support a role for imaging CCR2 cells within the fibrogenic niche in IPF to provide a molecular target for personalized therapy and monitoring.Clinical trial registered with www.clinicaltrials.gov (NCT03492762).
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http://dx.doi.org/10.1164/rccm.202004-1132OCDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7781144PMC
January 2021

Imaging resident and recruited macrophage contribution to Wallerian degeneration.

J Exp Med 2020 11;217(11)

Sorbonne Université, INSERM, CNRS, Centre d'Immunologie et des Maladies Infectieuses Cimi-Paris, Paris, France.

Wallerian degeneration (WD) is a process of autonomous distal degeneration of axons upon injury. Macrophages (MPs) of the peripheral nervous system (PNS) are the main cellular agent controlling this process. Some evidence suggests that resident PNS-MPs along with MPs of hematogenous origin may be involved, but whether these two subsets exert distinct functions is unknown. Combining MP-designed fluorescent reporter mice and coherent anti-Stokes Raman scattering (CARS) imaging of the sciatic nerve, we deciphered the spatiotemporal choreography of resident and recently recruited MPs after injury and unveiled distinct functions of these subsets, with recruited MPs being responsible for efficient myelin stripping and clearance and resident MPs being involved in axonal regrowth. This work provides clues to tackle selectively cellular processes involved in neurodegenerative diseases.
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http://dx.doi.org/10.1084/jem.20200471DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7596821PMC
November 2020

Revising CX3CR1 Expression on Murine Classical and Non-classical Monocytes.

Front Immunol 2020 3;11:1117. Epub 2020 Jun 3.

Sorbonne Université, Inserm, CNRS, Centre d'Immunologie et des Maladies Infectieuses, Cimi-Paris, Paris, France.

In mice, monocytes (Mo) are conventionally described as CX3CR1 classical Mo (CMo) and CX3CR1 non-classical Mo (NCMo) based on the expression of EGFP in Cx3cr1 mice and by analogy with human CX3CR1 expression. Although this terminology is widely used, it may not reflect the expression of CX3CR1 on Mo subsets. Using an unsupervised multiparametric analysis of blood Mo in steady state and after sterile peritonitis, we observed that CX3CR1 expression did not discriminate the CMo from the NCMo subsets. Our results highlight that despite being a reliable reporter to discriminate Mo subpopulations, EGFP level in Cx3cr1 mice does not reflect CX3CR1 expression measured by a fluorescently-labeled CX3CL1 chemokine and a CX3CR1 specific antibody. In conclusion, authors should be cautious not to identify murine classical and non-classical Mo as CX3CR1 and CX3CR1 but rather use alternative markers such as the combination of Ly6C and CD43.
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http://dx.doi.org/10.3389/fimmu.2020.01117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7283740PMC
June 2020

CX3CL1 homo-oligomerization drives cell-to-cell adherence.

Sci Rep 2020 06 3;10(1):9069. Epub 2020 Jun 3.

Sorbonne Université, Inserm, CNRS, Centre d'Immunologie et des Maladies Infectieuses (CIMI-Paris), U1135, 75013, Paris, France.

During inflammatory response, blood leukocytes adhere to the endothelium. This process involves numerous adhesion molecules, including a transmembrane chemokine, CX3CL1, which behaves as a molecular cluster. How this cluster assembles and whether this association has a functional role remain unknown. The analysis of CX3CL1 clusters using native electrophoresis and single molecule fluorescence kinetics shows that CX3CL1 is a homo-oligomer of 3 to 7 monomers. Fluorescence recovery after photobleaching assays reveal that the CX3CL1-transmembrane domain peptide self-associates in both cellular and acellular lipid environments, while its random counterpart (i.e. peptide with the same residues in a different order) does not. This strongly indicates that CX3CL1 oligomerization is driven by its intrinsic properties. According to the molecular modeling, CX3CL1 does not associate in compact bundles but rather with monomers linearly assembled side by side. Finally, the CX3CL1 transmembrane peptide inhibits both the CX3CL1 oligomerization and the adhesive function, while its random counterpart does not. This demonstrates that CX3CL1 oligomerization is mandatory for its adhesive potency. Our results provide a new direction to control CX3CL1-dependent cellular adherence in key immune processes.
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http://dx.doi.org/10.1038/s41598-020-65988-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7271195PMC
June 2020

Sepsis Triggers a Late Expansion of Functionally Impaired Tissue-Vascular Inflammatory Monocytes During Clinical Recovery.

Front Immunol 2020 30;11:675. Epub 2020 Apr 30.

Sorbonne Université, Inserm, CNRS, Centre d'Immunologie et des Maladies Infectieuses, Cimi-Paris, Paris, France.

Sepsis is characterized by a systemic inflammation that can cause an immune dysfunction, for which the underlying mechanisms are unclear. We investigated the impact of cecal ligature and puncture (CLP)-mediated polymicrobial sepsis on monocyte (Mo) mobilization and functions. Our results show that CLP led to two consecutive phases of Mo deployment. The first one occurred within the first 3 days after the induction of the peritonitis, while the second phase was of a larger amplitude and extended up to a month after apparent clinical recovery. The latter was associated with the expansion of Mo in the tissue reservoirs (bone marrow and spleen), their release in the blood and their accumulation in the vasculature of peripheral non-lymphoid tissues. It occurred even after antibiotic treatment but relied on inflammatory-dependent pathways and inversely correlated with increased susceptibility and severity to a secondary infection. The intravascular lung Mo displayed limited activation capacity, impaired phagocytic functions and failed to transfer efficient protection against a secondary infection into monocytopenic CCR2-deficient mice. In conclusion, our work unveiled key dysfunctions of intravascular inflammatory Mo during the recovery phase of sepsis and provided new insights to improve patient protection against secondary infections.
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http://dx.doi.org/10.3389/fimmu.2020.00675DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7212400PMC
April 2020

Phagocytosis of Wnt inhibitor SFRP4 by late wound macrophages drives chronic Wnt activity for fibrotic skin healing.

Sci Adv 2020 03 20;6(12):eaay3704. Epub 2020 Mar 20.

CEA/DRF/IBFJ/iRCM/LRTS, 92265 Fontenay-aux-Roses cedex, France.

Human and murine skin wounding commonly results in fibrotic scarring, but the murine wounding model wound-induced hair neogenesis (WIHN) can frequently result in a regenerative repair response. Here, we show in single-cell RNA sequencing comparisons of semi-regenerative and fibrotic WIHN wounds, increased expression of phagocytic/lysosomal genes in macrophages associated with predominance of fibrotic myofibroblasts in fibrotic wounds. Investigation revealed that macrophages in the late wound drive fibrosis by phagocytizing dermal Wnt inhibitor SFRP4 to establish persistent Wnt activity. In accordance, phagocytosis abrogation resulted in transient Wnt activity and a more regenerative healing. Phagocytosis of SFRP4 was integrin-mediated and dependent on the interaction of SFRP4 with the EDA splice variant of fibronectin. In the human skin condition hidradenitis suppurativa, phagocytosis of SFRP4 by macrophages correlated with fibrotic wound repair. These results reveal that macrophages can modulate a key signaling pathway via phagocytosis to alter the skin wound healing fate.
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http://dx.doi.org/10.1126/sciadv.aay3704DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7083618PMC
March 2020

CD36 Deficiency Inhibits Retinal Inflammation and Retinal Degeneration in Knockout Mice.

Front Immunol 2019 8;10:3032. Epub 2020 Jan 8.

Institut de la Vision, Sorbonne Université, INSERM, CNRS, Paris, France.

CD36, a member of the class B scavenger receptor family, participates in Toll-like receptor signaling on mononuclear phagocytes (MP) and can promote sterile pathogenic inflammation. We here analyzed the effect of CD36 deficiency on retinal inflammation and photoreceptor degeneration, the hallmarks of age-related macular degeneration (AMD), that characterize mice. We analyzed subretinal MP accumulation, and cone- and rod-degeneration in light-challenged and aged, CD36 competent or deficient, hyper-inflammatory mice, using histology and immune-stained retinal flatmounts. Monocytes (Mo) were subretinally adoptively transferred to evaluate their elimination rate from the subretinal space and Interleukin 6 (IL-6) secretion from cultured Mo-derived cells (MdCs) of the different mouse strains were analyzed. CD36 deficient mice were protected against age- and light-induced subretinal inflammation and associated cone and rod degeneration. CD36 deficiency in MPs inhibited their prolonged survival in the immune-suppressive subretinal space and reduced the exaggerated IL-6 secretion observed in MPs that we previously showed leads to increased subretinal MP survival. deficiency significantly protected hyperinflammatory mice against subretinal MP accumulation and associated photoreceptor degeneration. The observed CD36-dependent induction of pro-inflammatory IL-6 might be at least partially responsible for the prolonged MP survival in the immune-suppressive environment and its pathological consequences on photoreceptor homeostasis.
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http://dx.doi.org/10.3389/fimmu.2019.03032DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6960398PMC
November 2020

Tracking Monocytes and Macrophages in Tumors With Live Imaging.

Front Immunol 2019 31;10:1201. Epub 2019 May 31.

Centre d'Immunologie et des Maladies Infectieuses CIMI, CNRS, Sorbonne Université, Inserm, Paris, France.

In most cancers, myeloid cells represent the major component of the immune microenvironment. Deciphering the impact of these cells on tumor growth and in response to various anti-tumor therapies is a key issue. Many studies have elucidated the role of tumor-associated monocytes and tumor-associated macrophages (TAM) in tumor development, angiogenesis, and therapeutic failure. In contrast, tumor dendritic cells (DC) are associated with tumor antigen uptake and T-cell priming. Myeloid subpopulations display differences in ontogeny, state of differentiation and distribution within the neoplastic tissue, making them difficult to study. The development of high-dimensional genomic and cytometric analyses has unveiled the large functional diversity of myeloid cells. Important fundamental insights on the biology of myeloid cells have also been provided by a boom in functional fluorescent imaging techniques, in particular for TAM. These approaches allow the tracking of cell behavior in native physiological environments, incorporating spatio-temporal dimensions in the study of their functional activity. Nevertheless, tracking myeloid cells within the TME remains a challenging process as many markers overlap between monocytes, macrophages, DC, and neutrophils. Therefore, perfect discrimination between myeloid subsets remains impossible to date. Herein we review the specific functions of myeloid cells in tumor development unveiled by image-based tracking, the limits of fluorescent reporters commonly used to accurately track specific myeloid cells, and novel combinations of myeloid-associated fluorescent reporters that better discriminate the relative contributions of these cells to tumor biology according to their origin and tissue localization.
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http://dx.doi.org/10.3389/fimmu.2019.01201DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6555099PMC
August 2020

Ibrutinib induces multiple functional defects in the neutrophil response against .

Haematologica 2020 31;105(2):478-489. Epub 2020 Jan 31.

Sorbonne Université, INSERM, CNRS, Centre d'Immunologie et des Maladies Infectieuses, Cimi-Paris, Paris

The Bruton tyrosine kinase inhibitor ibrutinib has become a leading therapy against chronic lymphoid leukemia. Recently, ibrutinib has been associated with the occurrence of invasive fungal infections, in particular invasive aspergillosis. The mechanisms underlying the increased susceptibility to fungal infections associated with exposure to ibrutinib are currently unknown. Innate immunity, in particular polymer-phonuclear neutrophils, represents the cornerstone of anti- immunity; however, the potential impact of ibrutinib on neutrophils has been little studied. Our study investigated the response to and neutrophil function in patients with chronic lymphoid leukemia or lymphoma, who were undergoing ibrutinib therapy. We studied the consequences of ibrutinib exposure on the functions and anti- responses of neutrophils obtained from healthy donors and 63 blood samples collected at different time points from 32 patients receiving ibrutinib for lymphoid malignancies. We used both flow cytometry and video-microscopy approaches to analyze neutrophils' cell surface molecule expression, cytokine production, oxidative burst, chemotaxis and killing activity against Ibrutinib is associated, both and in patients under treatment, with multiple functional defects in neutrophils, including decreased production of reactive oxygen species, impairment of their capacity to engulf and inability to efficiently kill germinating conidia. Our results demonstrate that ibrutinib-exposed neutrophils develop significant functional defects that impair their response against , providing a plausible explanation for the emergence of invasive aspergillosis in ibrutinib-treated patients.
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http://dx.doi.org/10.3324/haematol.2019.219220DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7012467PMC
January 2020

CCR2-Dependent Recruitment of Tregs and Monocytes Following Radiotherapy Is Associated with TNFα-Mediated Resistance.

Cancer Immunol Res 2019 03 29;7(3):376-387. Epub 2019 Jan 29.

Sorbonne Université, Inserm, CNRS, Centre d'Immunologie et des Maladies Infectieuses Cimi-Paris, Paris, France.

Radiotherapy (RT) represents one of the main anticancer approaches for the treatment of solid tumors. Beyond the expected direct effects of RT on tumor cells, evidence supporting the importance of an immune response to RT is growing. The balance between RT-mediated immunogenic and tolerogenic activity is ill-defined and deserves more attention. Herein, a murine model of head and neck squamous cell carcinoma was used to demonstrate that RT upregulated CCL2 chemokine production in tumor cells, leading to a CCR2-dependent accumulation of tumor necrosis factor alpha (TNFα)-producing monocytes and CCR2 regulatory T cells (Treg). This corecruitment was associated with a TNFα-dependent activation of Tregs, dampening the efficacy of RT. Our results highlight an unexpected cross-talk between innate and adaptive immune system components and indicate CCL2/CCR2 and TNFα as potential clinical candidates to counterbalance the radioprotective action of monocyte-derived cells and Tregs, paving the way for potent combined radioimmunotherapies.
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http://dx.doi.org/10.1158/2326-6066.CIR-18-0633DOI Listing
March 2019

Molecular Imaging Visualizes Recruitment of Inflammatory Monocytes and Macrophages to the Injured Heart.

Circ Res 2019 03;124(6):881-890

Department of Medicine (B.K., G.F., G.B., S.L.B., K.J.L.), Washington University School of Medicine, St. Louis, MO.

Rationale: Paradigm shifting studies have revealed that the heart contains functionally diverse populations of macrophages derived from distinct embryonic and adult hematopoietic progenitors. Under steady-state conditions, the heart is largely populated by CCR2- (C-C chemokine receptor type 2) macrophages of embryonic descent. After tissue injury, a dramatic shift in macrophage composition occurs whereby CCR2+ monocytes are recruited to the heart and differentiate into inflammatory CCR2+ macrophages that contribute to heart failure progression. Currently, there are no techniques to noninvasively detect CCR2+ monocyte recruitment into the heart and thus identify patients who may be candidates for immunomodulatory therapy.

Objective: To develop a noninvasive molecular imaging strategy with high sensitivity and specificity to visualize inflammatory monocyte and macrophage accumulation in the heart.

Methods And Results: We synthesized and tested the performance of a positron emission tomography radiotracer (Ga-DOTA [1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid]-ECL1i [extracellular loop 1 inverso]) that allosterically binds to CCR2. In naive mice, the radiotracer was quickly cleared from the blood and displayed minimal retention in major organs. In contrast, biodistribution and positron emission tomography demonstrated strong myocardial tracer uptake in 2 models of cardiac injury (diphtheria toxin induced cardiomyocyte ablation and reperfused myocardial infarction). Ga-DOTA-ECL1i signal localized to sites of tissue injury and was independent of blood pool activity as assessed by quantitative positron emission tomography and ex vivo autoradiography. Ga-DOTA-ECL1i uptake was associated with CCR2+ monocyte and CCR2+ macrophage infiltration into the heart and was abrogated in CCR2 mice, demonstrating target specificity. Autoradiography demonstrated that Ga-DOTA-ECL1i specifically binds human heart failure specimens and with signal intensity associated with CCR2+ macrophage abundance.

Conclusions: These findings demonstrate the sensitivity and specificity of Ga-DOTA-ECL1i in the mouse heart and highlight the translational potential of this agent to noninvasively visualize CCR2+ monocyte recruitment and inflammatory macrophage accumulation in patients.
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http://dx.doi.org/10.1161/CIRCRESAHA.118.314030DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6435034PMC
March 2019

Macrophages of distinct origins contribute to tumor development in the lung.

J Exp Med 2018 10 10;215(10):2536-2553. Epub 2018 Sep 10.

Sorbonne Universités, Institut National de la Santé et de la Recherche Médicale (Inserm, UMR1135), Centre National de la Recherche Scientifique (CNRS, ERL8255), Centre d'Immunologie et des Maladies Infectieuses CIMI, Paris, France

Tissue-resident macrophages can self-maintain without contribution of adult hematopoiesis. Herein we show that tissue-resident interstitial macrophages (Res-TAMs) in mouse lungs contribute to the pool of tumor-associated macrophages (TAMs) together with CCR2-dependent recruited macrophages (MoD-TAMs). Res-TAMs largely correlated with tumor cell growth in vivo, while MoD-TAMs accumulation was associated with enhanced tumor spreading. Both cell subsets were depleted after chemotherapy, but MoD-TAMs rapidly recovered and performed phagocytosis-mediated tumor clearance. Interestingly, anti-VEGF treatment combined with chemotherapy inhibited both Res and Mod-TAM reconstitution without affecting monocyte infiltration and improved its efficacy. Our results reveal that the developmental origin of TAMs dictates their relative distribution, function, and response to cancer therapies in lung tumors.
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http://dx.doi.org/10.1084/jem.20180534DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6170177PMC
October 2018

Unveiling Cerebral Leishmaniasis: parasites and brain inflammation in Leishmania donovani infected mice.

Sci Rep 2017 08 16;7(1):8454. Epub 2017 Aug 16.

Institut Pasteur, Laboratoire des Processus Infectieux à Trypanosomatidés, Département Infection et Epidémiologie, 25-28 rue du Dr Roux, 75724, Cedex 15, Paris, France.

Visceral leishmaniasis (VL) is a systemic disease with multifaceted clinical manifestations, including neurological signs, however, the involvement of the nervous system during VL is underestimated. Accordingly, we investigated both brain infection and inflammation in a mouse model of VL. Using bioluminescent Leishmania donovani and real-time 2D-3D imaging tools, we strikingly detected live parasites in the brain, where we observed a compartmentalized dual-phased inflammation pattern: an early phase during the first two weeks post-infection, with the prompt arrival of neutrophils and Ly6C macrophages in an environment presenting a variety of pro-inflammatory mediators (IFN-γ, IL-1β, CXCL-10/CXCR-3, CCL-7/CCR-2), but with an intense anti-inflammatory response, led by IL-10; and a re-inflammation phase three months later, extremely pro-inflammatory, with novel upregulation of mediators, including IL-1β, TNF-α and MMP-9. These new data give support and corroborate previous studies connecting human and canine VL with neuroinflammation and blood-brain barrier disruption, and conclusively place the brain among the organs affected by this parasite. Altogether, our results provide convincing evidences that Leishmania donovani indeed infects and inflames the brain.
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http://dx.doi.org/10.1038/s41598-017-09085-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5559479PMC
August 2017

Analysis of monocyte infiltration in MPTP mice reveals that microglial CX3CR1 protects against neurotoxic over-induction of monocyte-attracting CCL2 by astrocytes.

J Neuroinflammation 2017 03 21;14(1):60. Epub 2017 Mar 21.

Inserm, U 1127, F-75013, Paris, France.

Background: Evidence from mice suggests that brain infiltrating immune cells contribute to neurodegeneration, and we previously identified a deleterious lymphocyte infiltration in Parkinson's disease mice. However, this remains controversial for monocytes, due to artifact-prone techniques used to distinguish them from microglia. Our aim was to reassess this open question, by taking advantage of the recent recognition that chemokine receptors CCR2 and CX3CR1 can differentiate between inflammatory monocytes and microglia, enabling to test whether CCR2 monocytes infiltrate the brain during dopaminergic (DA) neurodegeneration and whether they contribute to neuronal death. This revealed unexpected insights into possible regulation of monocyte-attracting CCL2 induction.

Methods: We used acute 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mice and assessed monocyte infiltration by combining laser microdissection-guided chemokine RNA profiling of the substantia nigra (SN) with immunohistochemistry and CCR2-GFP reporter mice. To determine contribution to neuronal loss, we used CCR2-deletion and CCL2-overexpression, to reduce and increase CCR2 monocyte infiltration, and CX3CR1-deletion to assess a potential implication in CCL2 regulation.

Results: Nigral chemokine profiling revealed early CCL2/7/12-CCR2 axis induction, suggesting monocyte infiltration in MPTP mice. CCL2 protein showed early peak induction in nigral astrocytes, while CCR2-GFP mice revealed early but limited nigral monocyte infiltration. However, blocking infiltration by CCR2 deletion did not influence DA neuronal loss. In contrast, transgenic astrocytic CCL2 over-induction increased CCR2 monocyte infiltration and DA neuronal loss in MPTP mice. Surprisingly, CCL2 over-induction was also detected in MPTP intoxicated CX3CR1-deleted mice, which are known to present increased DA neuronal loss. Importantly, CX3CR1/CCL2 double-deletion suggested that increased neurotoxicity was driven by astrocytic CCL2 over-induction.

Conclusions: We show that CCR2 monocytes infiltrate the affected CNS, but at the level observed in acute MPTP mice, this does not contribute to DA neuronal loss. In contrast, the underlying astrocytic CCL2 induction seemed to be tightly controled, as already moderate CCL2 over-induction led to increased neurotoxicity in MPTP mice, likely due to the increased CCR2 monocyte infiltration. Importantly, we found evidence suggesting that during DA neurodegeneration, this control was mediated by microglial CX3CR1 signaling, which protects against such neurotoxic CCL2 over-induction by astrocytes, thus hinting at an endogenous mechanism to limit neurotoxic effects of the CCL2-CCR2 axis.
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http://dx.doi.org/10.1186/s12974-017-0830-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5359822PMC
March 2017

Roles for the CX3CL1/CX3CR1 and CCL2/CCR2 Chemokine Systems in Hypoxic Pulmonary Hypertension.

Am J Respir Cell Mol Biol 2017 05;56(5):597-608

INSERM U955 and Département de Physiologie, Hôpital Henri Mondor, Assistance Publique-Hôpitaux de Paris, Départements Hospitalo Universitaires Ageing Thorax-Vessels-Blood, 94010, Créteil, France; Université Paris-Est Créteil, France; and Sorbonne Universités, Université Pierre et Marie Curie-Université Paris 06, Inserm, UMRS1135, CNRS, Equipes de Recherche Labellisées 8255, Centre d'Immunologie et des Maladies Infectieuses, Paris, France.

Monocytes/macrophages are major effectors of lung inflammation associated with various forms of pulmonary hypertension (PH). Interactions between the CCL2/CCR2 and CX3CL1/CX3CR1 chemokine systems that guide phagocyte infiltration are incompletely understood. Our objective was to explore the individual and combined actions of CCL2/CCR2 and CX3CL1/CX3CR1 in hypoxia-induced PH in mice; particularly their roles in monocyte trafficking, macrophage polarization, and pulmonary vascular remodeling. The development of hypoxia-induced PH was associated with marked increases in lung levels of CX3CR1, CCR2, and their respective ligands, CX3CL1 and CCL2. Flow cytometry revealed that both inflammatory Ly6C and resident Ly6C monocyte subsets exhibited sustained increases in blood and a transient peak in lung tissue, and that lung perivascular and alveolar macrophage counts showed sustained elevations. CX3CR1 mice were protected against hypoxic PH compared with wild-type mice, whereas CCL2 mice and double CX3CR1/CCL2 mice exhibited similar PH severity, as did wild-type mice. The protective effects of CX3CR1 deficiency occurred concomitantly with increases in lung monocyte and macrophage counts and with a change from M2 to M1 macrophage polarization that markedly diminished the ability of conditioned media to induce pulmonary artery smooth muscle cell (PA-SMC) proliferation, which was partly dependent on CX3CL1 secretion. Results in mice given the CX3CR1 inhibitor F1 were similar to those in CX3CR1 mice. In conclusion, CX3CR1 deficiency protects against hypoxia-induced PH by modulating monocyte recruitment, macrophage polarization, and PA-SMC cell proliferation. Targeting CX3CR1 may hold promise for treating PH.
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http://dx.doi.org/10.1165/rcmb.2016-0201OCDOI Listing
May 2017

PET-based Imaging of Chemokine Receptor 2 in Experimental and Disease-related Lung Inflammation.

Radiology 2017 06 3;283(3):758-768. Epub 2017 Jan 3.

From the Mallinckrodt Institute of Radiology (Y.L., D.H.S., H.P.L., Y.Z., R.J.G., S.L.B.) and Departments of Medicine (S.P.G., T.S.B., Z.B.N., J.H.P., D.E.B., J.J.A., M.J.H., R.J.G., S.L.B.), Surgery (D.K.), Pathology and Immunology (D.K.), and Cell Biology (M.J.H.), Washington University School of Medicine, 660 S Euclid Ave, Box 8052, St Louis, MO 63110; and Centre d'Immunologie et des Maladies Infectieuses, CIMI-Paris, Faculté de Médecine Pitié-Salpêtrière, Paris INSERM, Paris, France (C.C.).

Purpose To characterize a chemokine receptor type 2 (CCR2)-binding peptide adapted for use as a positron emission tomography (PET) radiotracer for noninvasive detection of lung inflammation in a mouse model of lung injury and in human tissues from subjects with lung disease. Materials and Methods The study was approved by institutional animal and human studies committees. Informed consent was obtained from patients. A 7-amino acid CCR2 binding peptide (extracellular loop 1 inverso [ECL1i]) was conjugated to tetraazacyclododecane tetraacetic acid (DOTA) and labeled with copper 64 (Cu) or fluorescent dye. Lung inflammation was induced with intratracheal administration of lipopolysaccharide (LPS) in wild-type (n = 19) and CCR2-deficient (n = 4) mice, and these mice were compared with wild-type mice given control saline (n = 5) by using PET performed after intravenous injection of Cu-DOTA-ECL1i. Lung immune cells and those binding fluorescently labeled ECL1i in vivo were detected with flow cytometry. Lung inflammation in tissue from subjects with nondiseased lungs donated for lung transplantation (n = 11) and those with chronic obstructive pulmonary disease (COPD) who were undergoing lung transplantation (n = 16) was evaluated for CCR2 with immunostaining and autoradiography (n = 6, COPD) with Cu-DOTA-ECL1i. Groups were compared with analysis of variance, the Mann-Whitney U test, or the t test. Results Signal on PET images obtained in mouse lungs after injury with LPS was significantly greater than that in the saline control group (mean = 4.43% of injected dose [ID] per gram of tissue vs 0.99% of injected dose per gram of tissue; P < .001). PET signal was significantly diminished with blocking studies using nonradiolabeled ECL1i in excess (mean = 0.63% ID per gram of tissue; P < .001) and in CCR2-deficient mice (mean = 0.39% ID per gram of tissue; P < .001). The ECL1i signal was associated with an elevated level of mouse lung monocytes. COPD lung tissue displayed significantly elevated CCR2 levels compared with nondiseased tissue (median = 12.8% vs 1.2% cells per sample; P = .002), which was detected with Cu-DOTA-ECL1i by using autoradiography. Conclusion Cu-DOTA-ECL1i is a promising tool for PET-based detection of CCR2-directed inflammation in an animal model and in human tissues as a step toward clinical translation. RSNA, 2017 Online supplemental material is available for this article.
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http://dx.doi.org/10.1148/radiol.2016161409DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5452886PMC
June 2017

CX3CR1-dependent endothelial margination modulates Ly6C monocyte systemic deployment upon inflammation in mice.

Blood 2017 03 23;129(10):1296-1307. Epub 2016 Dec 23.

Sorbonne Universités, UPMC Université Paris 06, INSERM UMR 1135, CNRS ERL8255, Centre d'Immunologie et des Maladies Infectieuses, Paris, France.

Two subsets of blood monocytes are commonly described in mice and humans: the classical inflammatory monocytes, which are rapidly mobilized upon inflammation in a CC-chemokine receptor 2-dependent manner, and the nonclassical blood resident monocyte subset that patrols the intraluminal side of the endothelium. Old reports suggest that blood monocytes are distributed into circulating and marginating pools, but no direct evidence of the latter has been obtained so far. Using a combination of in vivo real-time imaging and blood/tissue partitioning by intravascular staining of leukocytes, we showed that both inflammatory and resident monocytes are retained in the bone marrow vasculature, representing an important reservoir of marginated monocytes. Upon lipopolysaccharide or cecal ligation and puncture-induced peritonitis, these marginated cells are rapidly released and recruited to the peritoneum membrane lumen vasculature where they reside through CX3C-chemokine receptor 1 (CX3CR1)-dependent adherence. At a later time point, inflammatory monocytes infiltrate the spleen parenchyma but remain mainly intravascular in the vicinity of the lungs and the peritoneum. Our results show that this monocyte deployment is controlled by a CX3CR1-dependent balance between marginating and circulating monocytes and highlight that tissue infiltration is not a mandatory fate for inflammatory monocytes.
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http://dx.doi.org/10.1182/blood-2016-08-732164DOI Listing
March 2017

CCR2 Influences T Regulatory Cell Migration to Tumors and Serves as a Biomarker of Cyclophosphamide Sensitivity.

Cancer Res 2016 11 28;76(22):6483-6494. Epub 2016 Sep 28.

Sorbonne Universités, UPMC Université Paris 06 UMR_S1135, Institut Universitaire de Cancérologie (IUC), Inserm U1135, CNRS ERL8255, Centre d'Immunologie et des Maladies Infectieuses, Paris, France.

The CCL2 chemokine receptor CCR2 drives cancer by mediating the recruitment of monocytes and myeloid-derived suppressor cells to the tumor microenvironment. In this study, we extend the significance of CCR2 in this setting by identifying a new role for it in mediating recruitment of CD4 T regulatory cells (Treg). Following tumor initiation, an expanded population of CCR2 Tregs required CCR2 expression to traffic between draining lymph nodes (dLN) and the tumor. This Treg subset was enriched in the fraction of tumor antigen-specific cells in the dLN, where they displayed an activated immunosuppressive phenotype. Notably, in mouse models, low-dose cyclophosphamide treatment preferentially depleted CCR2 Treg, enhancing priming of tumor-specific CD8 T cells. In the MMTV-PyMT transgenic mouse model of breast cancer and in oral squamous cell carcinoma patients, tumor development was associated with decreased blood frequency and inversely increased tumor frequency of CCR2 Tregs. Our results define a novel subset of CCR2 Treg involved in tumoral immune escape, and they offer evidence that this Treg subset may be preferentially eradicated by low-dose cyclophosphamide treatment. Cancer Res; 76(22); 6483-94. ©2016 AACR.
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http://dx.doi.org/10.1158/0008-5472.CAN-16-0984DOI Listing
November 2016

Plasmodium falciparum proteins involved in cytoadherence of infected erythrocytes to chemokine CX3CL1.

Sci Rep 2016 Sep 22;6:33786. Epub 2016 Sep 22.

Sorbonne Universités, UPMC Univ Paris 06, Inserm, Centre d'Immunologie et des Maladies Infectieuses (Cimi-Paris), UMR 1135, ERL CNRS 8255, 91 boulevard de l'Hôpital F-75013, Paris, France.

Malaria caused by Plasmodium falciparum is associated with cytoadherence of infected red blood cells (iRBC) to endothelial cells. Numerous host molecules have been involved in cytoadherence, including the adhesive chemokine CX3CL1. Most of the identified parasite ligands are from the multigenic and hypervariable Plasmodium falciparum Erythrocyte Membrane Protein 1 (PfEMP1) family which makes them poor targets for the development of a broadly protective vaccine. Using proteomics, we have identified two 25-kDa parasite proteins with adhesive properties for CX3CL1, called CBP for CX3CL1 Binding Proteins. CBPs are coded by single-copy genes with little polymorphic variation and no homology with other P. falciparum gene products. Specific antibodies raised against epitopes from the predicted extracellular domains of each CBP efficiently stain the surface of RBC infected with trophozoites or schizonts, which is a strong indication of CBP expression at the surface of iRBC. These anti-CBP antibodies partially neutralize iRBC adherence to CX3CL1. This adherence is similarly inhibited in the presence of peptides from the CBP extracellular domains, while irrelevant peptides had no such effect. CBP1 and CBP2 are new P. falciparum ligands for the human chemokine CX3CL1. The identification of this non-polymorphic P. falciparum factors provides a new avenue for innovative vaccination approaches.
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http://dx.doi.org/10.1038/srep33786DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5031962PMC
September 2016

ECL1i, d(LGTFLKC), a novel, small peptide that specifically inhibits CCL2-dependent migration.

FASEB J 2016 06 15;30(6):2370-81. Epub 2016 Mar 15.

*Sorbonne Universités, Université Pierre et Marie Curie (UPMC)/Univ Paris 06, Unité Mixte de Recherche Scientifique (UMRS) 1135, INSERM Unité 1135, Centre National de la Recherche Scientifique, Equipe de Recherche Labellisée (ERL) 8255, Centre d'Immunologie et des Maladies Infectieuses, Paris, France;

CC chemokine receptor type 2 (CCR2) is a key molecule in inflammatory diseases and is an obvious drug target for the treatment of inflammation. A number of nonpeptidic, competitive CCR2 antagonists have been developed, but none has yet been approved for clinical use. Our aim was to identify a short peptide that showed allosteric antagonism against human and mouse CCR2. On the basis of sequence analysis and 3-dimensional modeling, we identified an original 7-d-amino acid peptidic CCR2 inhibitor that we have called extracellular loop 1 inverso (ECL1i), d(LGTFLKC). In vitro, ECL1i selectively and potently inhibits CC chemokine ligand type 2 (CCL2)-triggered chemotaxis (IC50, 2 µM) but no other conventional CCL2-associated events. We used the classic competitive CCR2 antagonist, BMS22 {2-[(isopropylaminocarbonyl)amino]-N-[2-[[cis-2-[[4-(methylthio)benzoyl]amino]cyclohexyl]amino]-2-oxoethyl]-5-(trifluoromethyl)benzamide}, as positive control and inhibited CCL2-dependent chemotaxis with an IC50 of 18 nM. As negative control, we used a peptide with the same composition as ECL1i, but in a different sequence, d(FKLTLCG). In vivo, ECL1i (4 mg/kg) interfered with CCR2-positive cell recruitment and attenuated disease progression in experimental autoimmune encephalomyelitis, a mouse model of multiple sclerosis. This study establishes ECL1i as the first allosteric inhibitor of CCR2 with functional selectivity. ECL1i is a promising new agent in therapeutic development, and it may, by its selective effect, increase our understanding of CCR2 signaling pathways and functions.-Auvynet, C., Baudesson de Chanville, C., Hermand, P., Dorgham, K., Piesse, C., Pouchy, C., Carlier, L., Poupel, L., Barthélémy, S., Felouzis, V., Lacombe, C., Sagan, S., Salomon, B., Deterre, P., Sennlaub, F., Combadière, C. ECL1i, d(LGTFLKC), a novel, small peptide that specifically inhibits CCL2-dependent migration.
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http://dx.doi.org/10.1096/fj.201500116DOI Listing
June 2016

CX3CR1 deficiency promotes muscle repair and regeneration by enhancing macrophage ApoE production.

Nat Commun 2015 Dec 3;6:8972. Epub 2015 Dec 3.

Sorbonne Universités, UPMC Univ Paris 06, Inserm, U1135, CNRS, ERL 8255, Centre d'Immunologie et des Maladies Infectieuses (CIMI-Paris), 91 Boulevard de l'Hôpital, F-75013 Paris, France.

Muscle injury triggers inflammation in which infiltrating mononuclear phagocytes are crucial for tissue regeneration. The interaction of the CCL2/CCR2 and CX3CL1/CX3CR1 chemokine axis that guides phagocyte infiltration is incompletely understood. Here, we show that CX3CR1 deficiency promotes muscle repair and rescues Ccl2(-/-) mice from impaired muscle regeneration as a result of altered macrophage function, not infiltration. Transcriptomic analysis of muscle mononuclear phagocytes reveals that Apolipoprotein E (ApoE) is upregulated in mice with efficient regeneration. ApoE treatment enhances phagocytosis by mononuclear phagocytes in vitro, and restores phagocytic activity and muscle regeneration in Ccl2(-/-) mice. Because CX3CR1 deficiency may compensate for defective CCL2-dependant monocyte recruitment by modulating ApoE-dependent macrophage phagocytic activity, targeting CX3CR1 expressed by macrophages might be a powerful therapeutic approach to improve muscle regeneration.
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http://dx.doi.org/10.1038/ncomms9972DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4686853PMC
December 2015

Comprehensive analysis of chemokine-induced cAMP-inhibitory responses using a real-time luminescent biosensor.

Cell Signal 2016 Jan 26;28(1):120-9. Epub 2015 Oct 26.

Sorbonne Universités, UPMC Université Paris 06, Inserm U 1135, CNRS ERL 8255, Centre d'Immunologie et des Maladies Infectieuses, 91 Boulevard de l'Hôpital, F-75013 Paris, France. Electronic address:

Chemokine receptors are members of the G-protein-coupled receptor (GPCR) family coupled to members of the Gi class, whose primary function is to inhibit the cellular adenylate cyclase. We used a cAMP-related and PKA-based luminescent biosensor (GloSensor™ F-22) to monitor the real-time downstream response of chemokine receptors, especially CX3CR1 and CXCR4, after activation with their cognate ligands CX3CL1 and CXCL12. We found that the amplitudes and kinetic profiles of the chemokine responses were conserved in various cell types and were independent of the nature and concentration of the molecules used for cAMP prestimulation, including either the adenylate cyclase activator forskolin or ligands mediating Gs-mediated responses like prostaglandin E2 or beta-adrenergic agonist. We conclude that the cAMP chemokine response is robustly conserved in various inflammatory conditions. Moreover, the cAMP-related luminescent biosensor appears as a valuable tool to analyze the details of Gi-mediated cAMP-inhibitory cellular responses, even in native conditions and could help to decipher their precise role in cell function.
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http://dx.doi.org/10.1016/j.cellsig.2015.10.011DOI Listing
January 2016

Immune surveillance of the lung by migrating tissue monocytes.

Elife 2015 Jul 13;4:e07847. Epub 2015 Jul 13.

Centre d'Immunologie et des Maladies Infectieuses, University Pierre et Marie Curie, Sorbonne Universities, Paris, France.

Monocytes are phagocytic effector cells in the blood and precursors of resident and inflammatory tissue macrophages. The aim of the current study was to analyse and compare their contribution to innate immune surveillance of the lung in the steady state with macrophage and dendritic cells (DC). ECFP and EGFP transgenic reporters based upon Csf1r and Cx3cr1 distinguish monocytes from resident mononuclear phagocytes. We used these transgenes to study the migratory properties of monocytes and macrophages by functional imaging on explanted lungs. Migratory monocytes were found to be either patrolling within large vessels of the lung or locating at the interface between lung capillaries and alveoli. This spatial organisation gives to monocytes the property to capture fluorescent particles derived from both vascular and airway routes. We conclude that monocytes participate in steady-state surveillance of the lung, in a way that is complementary to resident macrophages and DC, without differentiating into macrophages.
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http://dx.doi.org/10.7554/eLife.07847DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4521583PMC
July 2015

Ly6Chigh Monocytes Protect against Kidney Damage during Sepsis via a CX3CR1-Dependent Adhesion Mechanism.

J Am Soc Nephrol 2016 Mar 9;27(3):792-803. Epub 2015 Jul 9.

Sorbonne Universités, Université Pierre et Marie Curie (UPMC), University of Paris 06, Paris, France; Institut National de la Santé et de la Recherche Médicale (INSERM), U1135, Paris, France; Centre National de la Recherche Scientifique (CNRS), Paris, France;

Monocytes have a crucial role in both proinflammatory and anti-inflammatory phenomena occurring during sepsis. Monocyte recruitment and activation are orchestrated by the chemokine receptors CX3CR1 and CCR2 and their cognate ligands. However, little is known about the roles of these cells and chemokines during the acute phase of inflammation in sepsis. Using intravital microscopy in a murine model of polymicrobial sepsis, we showed that inflammatory Ly6C(high) monocytes infiltrated kidneys, exhibited altered motility, and adhered strongly to the renal vascular wall in a chemokine receptor CX3CR1-dependent manner. Adoptive transfer of Cx3cr1-proficient monocyte-enriched bone marrow cells into septic Cx3cr1-depleted mice prevented kidney damage and promoted mouse survival. Modulation of CX3CR1 activation in septic mice controlled monocyte adhesion, regulated proinflammatory and anti-inflammatory cytokine expression, and was associated with the extent of kidney lesions such that the number of lesions decreased when CX3CR1 activity increased. Consistent with these results, the pro-adhesive I249 CX3CR1 allele in humans was associated with a lower incidence of AKI in patients with sepsis. These data show that inflammatory monocytes have a protective effect during sepsis via a CX3CR1-dependent adhesion mechanism. This receptor might be a new therapeutic target for kidney injury during sepsis.
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http://dx.doi.org/10.1681/ASN.2015010009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4769199PMC
March 2016

Neuronal Cx3cr1 Deficiency Protects against Amyloid β-Induced Neurotoxicity.

PLoS One 2015 3;10(6):e0127730. Epub 2015 Jun 3.

Laboratory of Molecular Immunology, National Institute of Allergy and Infectious Disease, National Institutes of Health, Bethesda, Maryland, United States of America.

Cx3cr1, the receptor for the chemokine Cx3cl1 (fractalkine), has been implicated in the progression and severity of Alzheimer's disease-like pathology in mice, but the underlying mechanisms remain unclear. A complicating factor is that Cx3cr1 has been demonstrated in both neurons and microglia. Here, we have dissected the differences between neuronal and microglial Cx3cr1, specifically by comparing direct amyloid-β-induced toxicity in cultured, mature, microglia-depleted hippocampal neurons from wild-type and Cx3cr1-/- mice. Wild-type neurons expressed both Cx3cl1 and Cx3cr1 and released Cx3cl1 in response to amyloid-β. Knockout of neuronal Cx3cr1 abated amyloid-β-induced lactate dehydrogenase release. Furthermore, amyloid-β differentially induced depression of pre- and postsynaptic components of miniature excitatory postsynaptic currents, in a peptide conformation-dependent manner. Knockout of neuronal Cx3cr1 abated effects of both amyloid-β conformational states, which were differentiable by aggregation kinetics and peptide morphology. We obtained similar results after both acute and chronic treatment of cultured neurons with the Cx3cr1 antagonist F1. Thus, neuronal Cx3cr1 may impact Alzheimer's disease-like pathology by modulating conformational state-dependent amyloid-β-induced synaptotoxicity.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0127730PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4454597PMC
February 2016

Tracking mouse bone marrow monocytes in vivo.

J Vis Exp 2015 Feb 27(96):e52476. Epub 2015 Feb 27.

Centre d'Immunologie et des Maladies Infectieuses (CIMI), INSERM, U1135, CNRS, ERL 8255, Sorbonne Universités, UPMC Univ Paris 06, CR7;

Real time multiphoton imaging provides a great opportunity to study cell trafficking and cell-to-cell interactions in their physiological 3-dimensionnal environment. Biological activities of immune cells mainly rely on their motility capacities. Blood monocytes have short half-life in the bloodstream; they originate in the bone marrow and are constitutively released from it. In inflammatory condition, this process is enhanced, leading to blood monocytosis and subsequent infiltration of the peripheral inflammatory tissues. Identifying the biomechanical events controlling monocyte trafficking from the bone marrow towards the vascular network is an important step to understand monocyte physiopathological relevance. We performed in vivo time-lapse imaging by two-photon microscopy of the skull bone marrow of the Csf1r-Gal4VP16/UAS-ECFP (MacBlue) mouse. The MacBlue mouse expresses the fluorescent reporters enhanced cyan fluorescent protein (ECFP) under the control of a myeloid specific promoter, in combination with vascular network labelling. We describe how this approach enables the tracking of individual medullar monocytes in real time to further quantify the migratory behaviour within the bone marrow parenchyma and the vasculature, as well as cell-to-cell interactions. This approach provides novel insights into the biology of the bone marrow monocyte subsets and allows to further address how these cells can be influenced in specific pathological conditions.
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http://dx.doi.org/10.3791/52476DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4401155PMC
February 2015

NFκB activation by modified vaccinia virus as a novel strategy to enhance neutrophil migration and HIV-specific T-cell responses.

Proc Natl Acad Sci U S A 2015 Mar 4;112(11):E1333-42. Epub 2015 Mar 4.

Departments of Molecular and Cellular Biology and

Neutrophils are antigen-transporting cells that generate vaccinia virus (VACV)-specific T-cell responses, yet how VACV modulates neutrophil recruitment and its significance in the immune response are unknown. We generated an attenuated VACV strain that expresses HIV-1 clade C antigens but lacks three specific viral genes (A52R, K7R, and B15R). We found that these genes act together to inhibit the NFκB signaling pathway. Triple ablation in modified virus restored NFκB function in macrophages. After virus infection of mice, NFκB pathway activation led to expression of several cytokines/chemokines that increased the migration of neutrophil populations (Nα and Nβ) to the infection site. Nβ cells displayed features of antigen-presenting cells and activated virus-specific CD8 T cells. Enhanced neutrophil trafficking to the infection site correlated with an increased T-cell response to HIV vector-delivered antigens. These results identify a mechanism for poxvirus-induced immune response and alternatives for vaccine vector design.
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http://dx.doi.org/10.1073/pnas.1424341112DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4371968PMC
March 2015

Apolipoprotein E promotes subretinal mononuclear phagocyte survival and chronic inflammation in age-related macular degeneration.

EMBO Mol Med 2015 Feb;7(2):211-26

INSERM, Paris, France UPMC Univ Paris 06 UMR_S 968 Institut de la Vision, Paris, France Centre Hospitalier National d'Ophtalmologie des Quinze-Vingts INSERM-DHOS CIC 503, Paris, France

Physiologically, the retinal pigment epithelium (RPE) expresses immunosuppressive signals such as FAS ligand (FASL), which prevents the accumulation of leukocytes in the subretinal space. Age-related macular degeneration (AMD) is associated with a breakdown of the subretinal immunosuppressive environment and chronic accumulation of mononuclear phagocytes (MPs). We show that subretinal MPs in AMD patients accumulate on the RPE and express high levels of APOE. MPs of Cx3cr1(-/-) mice that develop MP accumulation on the RPE, photoreceptor degeneration, and increased choroidal neovascularization similarly express high levels of APOE. ApoE deletion in Cx3cr1(-/-) mice prevents pathogenic age- and stress-induced subretinal MP accumulation. We demonstrate that increased APOE levels induce IL-6 in MPs via the activation of the TLR2-CD14-dependent innate immunity receptor cluster. IL-6 in turn represses RPE FasL expression and prolongs subretinal MP survival. This mechanism may account, in part, for the MP accumulation observed in Cx3cr1(-/-) mice. Our results underline the inflammatory role of APOE in sterile inflammation in the immunosuppressive subretinal space. They provide rationale for the implication of IL-6 in AMD and open avenues toward therapies inhibiting pathogenic chronic inflammation in late AMD.
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http://dx.doi.org/10.15252/emmm.201404524DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4328649PMC
February 2015

Pachymodulin, a new functional formyl peptide receptor 2 peptidic ligand isolated from frog skin has Janus-like immunomodulatory capacities.

J Med Chem 2015 Feb 4;58(3):1089-99. Epub 2015 Feb 4.

Sorbonne Universités, UPMC Univ Paris 06, LBM , 4 place Jussieu, 75005 Paris, France.

Recruitment of leukocytes is essential to fight infections or to heal injuries; however, excessive and/or prolonged responses favor the development of major inflammatory pathologies, such as cardiovascular or neurodegenerative diseases. Thus, it is of great interest to seek novel compounds that can regulate leukocyte recruitment depending on the degree of inflammation. We have isolated and characterized, by different chromatographic techniques, mass spectrometry, and Edman sequencing, a new hexapeptide (SSLSKL) from the Mexican frog Pachymedusa dacnicolor, which we named pachymodulin. In vitro, pachymodulin promotes the migration of leukocytes through the binding and activation of the human and mouse N-formyl peptide receptor 2 (huFPR2). In vivo, it exhibits opposite biological activities: under homeostatic conditions, pachymodulin induces the recruitment of leukocytes, whereas under inflammatory conditions, it inhibits this process. Therefore, pachymodulin represents an interesting template in the quest to design new immunomodulatory drugs in the therapy of immune-related diseases.
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http://dx.doi.org/10.1021/jm501018qDOI Listing
February 2015

Stromal cell-derived CCL2 drives neuropathic pain states through myeloid cell infiltration in injured nerve.

Brain Behav Immun 2015 Mar 4;45:198-210. Epub 2014 Nov 4.

Sorbonne Universités, UPMC Université Paris 06, Institut de la vision, équipe S12, 17 rue Moreau, F-75012, Paris, France; UMR_S 968 INSERM, 17 rue Moreau, F-75012, Paris, France; UMR 7210 CNRS, 17 rue Moreau, F-75012, Paris, France. Electronic address:

Neuropathic pain resulting from peripheral nerve injury involves many persistent neuroinflammatory processes including inflammatory chemokines that control leukocyte trafficking and activate resident cells. Several studies have shown that CCL2 chemokine, a potent attractant of monocytes, and its cognate receptor, CCR2, play a critical role in regulating nociceptive processes during neuropathic pain. However, the role of CCL2 in peripheral leukocyte infiltration-associated neuropathic pain remains poorly understood. In particular, the contribution of individual CCL2-expressing cell populations (i.e. stromal and leukocytes) to immune cell recruitment into the injured nerve has not been established. Here, in preclinical model of peripheral neuropathic pain (i.e. chronic constriction injury of the sciatic nerve), we have demonstrated that, CCL2 content was increased specifically in nerve fibers. This upregulation of CCL2 correlated with local monocyte/macrophage infiltration and pain processing. Furthermore, sciatic intraneural microinjection of CCL2 in naïve animals triggered long-lasting pain behavior associated with local monocyte/macrophage recruitment. Using a specific CCR2 antagonist and mice with a CCL2 genetic deletion, we have also established that the CCL2/CCR2 axis drives monocyte/macrophage infiltration and pain hypersensitivity in the CCI model. Finally, specific deletion of CCL2 in stromal or immune cells respectively using irradiated bone marrow-chimeric CCI mice demonstrated that stromal cell-derived CCL2 (in contrast to CCL2 immune cell-derived) tightly controls monocyte/macrophage recruitment into the lesion and plays a major role in the development of neuropathic pain. These findings demonstrate that in chronic pain states, CCL2 expressed by sciatic nerve cells predominantly drove local neuro-immune interactions and pain-related behavior through CCR2 signaling.
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http://dx.doi.org/10.1016/j.bbi.2014.10.016DOI Listing
March 2015