Deposition of urate and pyrophosphate crystals (MSU and CPP) in the joints are responsible for gout and chondrocalcinosis respectively. The inflammatory reactions induced in these pathologies are due to the activation of resident macrophages involved in the release cytokines including IL1?. The maturation of IL1? is mediated by a multiprotein complex, the NLRP3 inflammasome. We have found that the activation of NLRP3 by crystals depends on cell volume regulation. The LRRC8 family of proteins form the main molecular components of the volume-dependent channel involved in volume regulation. We have found in cultured macrophages that LRRC8 played an essential role in the activation of the inflammasome NRLP3 and the secretion of IL1? following crystal exposure (MSU/CPP). At the same time, we observed in murine inflammatory models that the modulation of osmotic pressure influences the inflammatory processes (IL1?) induced by crystal injection (inhibition by increasing osmotic pressure). We hypothesize that LRRC8 channels control crystal-induced NLRP3 activation through the coincidence of different processes: intracellular Cl- / K + variation, modulation of oxidative stress, and release of extracellular ATP (likely to initiate purinergic action. auto / para-crine, signalling loop). In this project, we will assess i) the role of the LRRC8 / VRAC channel in the activation of NLRP3, IL1? secretion and cell volume in response to crystals, ii) the effect of conditional gene inactivation of LRRC8 in a murine model of crystal inflammation (CR3CR1 / LRRC8A mice) and iii) the involvement of these mechanisms and of LRRC8/VRAC in patients suffering from these pathologies (collection of synovial fluids) in order to identify new therapeutic targets. Altogether, we expect to characterize novel and unexpected functions of the chloride channels LRRC8/VRAC with clear pathophysiological relevance in the context of crystal-induced inflammation.

Brown adipose tissue plays a key role during cold exposure by generating heat, a process named thermogenesis. Thermogenesis is essential to maintain body temperature and brown fat activation improves the host’s metabolism. Various types of immune cells have been identified in brown adipose tissue and their contribution to thermogenesis has been suggested. Macrophages are the most abundant immune cells in brown fat. We recently identified several brown adipose tissue macrophage subsets co-existing within this tissue. The current proposal aims to investigate the functions and relative contribution of brown fat macrophage subsets to homeostatic tissue maintenance and during cold-induced thermogenesis and stress. Molecular biology (RNAseq, lipidomics) and cell biology (spectral cytometry, confocal microscopy) approaches combined to original mouse lines will be used for this project. This consortium gathers 4 research teams with complementary expertise in immunology, metabolism and lipidomics.

Systemic manifestations are a hallmark of chronic inflammatory diseases, as for instance bone destruction in inflammatory bowel disease (IBD). Anti-TNF therapy is the first-line of treatment for IBD but despite its efficacy on gut inflammation and bone destruction, it induces psoriatic skin lesions. In addition of serious consequences for patient care, this side effect raises the question of the tissue specificity of the control of inflammatory responses that remains largely unknown. Our aims are to understand the specific response to anti-TNF in the gut, bone and skin focusing on environmental factors and on immune cells particularly IL-17 secreting cells (Th17 and ILC3). We will combine analysis in a murine IBD model mimicking what observed in patients +/- anti-TNF treatment and in IBD patients treated or not with anti-TNF therapy having or not psoriasis. The results will open new clues for improving therapeutic approaches or prognosis markers for anti-TNF paradoxical effects and may impact other chronic diseases in which such effects are also reported.

Arterial hypertension (HT) is one of the major risk factors for cardiovascular disease and has an important cost for society. Pathogenic mechanisms underlying HT are complex and include genetic and environmental, vascular and hormonal factors. Detection of secondary forms of HT is particularly important since it allows for the targeted management of the underlying disease. Primary aldosteronism (PA) is the most common form of endocrine hypertension. Different adrenal diseases are responsible for PA: i) aldosterone-producing adenoma (APA or Conn's adenoma, ~50% of cases); ii) idiopathic hyperaldosteronism or bilateral adrenal hyperplasia (BAH, 30-40%); iii) unilateral primary adrenal hyperplasia (documented unilateral aldosterone secretion without detectable adenoma, 5-10%) and iv) aldosterone producing adrenal carcinoma in rare cases. Once the diagnosis of PA has been made, it is important to identify its etiology, in order to distinguish between surgically correctable forms (APA and unilateral primary adrenal hyperplasia) and forms to be treated pharmacologically (BAH). Patients with PA exhibit more severe left ventricular hypertrophy and diastolic dysfunction than patients with essential hypertension and a high prevalence of myocardial infarction, stroke and atrial fibrillation. Moreover, among unilateral forms of PA, hypertension is cured after surgery in less than 50% of patients. The understanding of the pathogenic mechanisms underlying PA is then essential to allow for the development of new diagnostic tools and biomarkers and for the identification of new therapeutic targets, concerning up to 10% of the hypertensive population. Increasing evidence shows the relevance of local mechanisms regulating aldosterone production in the adrenal, as opposed to cardiovascular regulation by the renin-angiotensin system. Our research project aims to investigate pivotal aspects of the mechanisms implicated in local control of aldosterone secretion in the adrenal through a strategy that integrates functional genomics, mouse models and clinical studies, with the aim to better understand the physiopathology of PA for improved therapeutic intervention in hypertensive patients. The three French partners and the German partner implicated in the project have a long-lasting and very productive collaboration record in the field of the study of the physiopathology of aldosterone production. In the framework of their previous studies, critical advances have been made in the understanding of the physiopathology of PA, with the characterization of the in vivo role of potassium channels in the regulation of aldosterone secretion by the adrenal cortex and the identification of somatic mutations driving APA phenotype. The first part of our project will focus on the identification of new genes involved in the pathogenesis of APA using the unique model of the Kcnk3 null mice previously characterized by our consortium, which present sex- and sexual hormone-dependent PA, and that allowed to identify a restricted set of genes with a potential role in the regulation of aldosterone secretion in the adrenal. Using a strategy that will integrate genetic, cell and animal studies, we will identify those genes that are relevant for aldosterone secretion in vivo and are associated to PA. In the second part of our project we will tackle the problem of the dissociation of hypersecretion and neoplastic nodule formation in pathological adrenals that overproduce aldosterone. We will produce the first mouse models with an altered function in genes whose homologs are implicated in APA and by genomic analysis we will identify genetic abnormalities in multinodular adrenals whose role in the nodulation process will be validated by producing ad hoc mouse models. We are confident that the results of our project will shed new light on the pathogenetic mechanisms of PA and allow to design new targeted therapies for this disease.

Hematopoietic stem and progenitor cells are responsible for replenishing immune cells and reside in bone marrow (BM) endosteal and vascular niches. The divergence of lymphoid and myeloid lineages occurs at the multipotent progenitor (MPP) stage. Although numerous studies focus on molecular regulation of MPP lineage specification, the influence of extracellular factors or localization of MPPs in distinct niches remain to be explored. Distinct subpopulations of lineage-biased MPPs operate together to control blood leukocyte production. Both the localization and niches of these MPP subsets in BM are unknown. It is also unclear whether regional BM localization of MPP subpopulations impacts their lineage specification and commitment through specific adhesion cues or chemokine signaling. Reciprocally, it is unknown whether MPPs can regulate their own niche through interactions with BM stroma, and if so, by which mechanisms. The general objective of this project that gathers three partners with complementary expertise in osteo-immunology, hematology and leuko-stroma interaction biology aims at delineating how CXCL12 and its two receptors, namely CXCR4 and CXCR7/ACKR3, contribute to the integrity and maintenance of the BM osteo-vascular niches. We will also determine whether the CXCL12/CXCR4-CXCR7 trio endows these niche elements with the capacity to support MPP localization and differentiation during hematopoiesis in mice and humans. Reciprocally, we will ask whether MPPs use this axis to influence their niches. The global strategy will consist in characterizing the functional expression of the CXCL12/CXCR4-CXCR7 trio in MPP subsets as well as in the osteo-vascular niche elements. Microarray-based trans-interactome analyses of these cell populations will be used to identify and validate ligand/receptor pairs involved in MPP/stroma crosstalks that govern MPP positioning and specification. This will be addressed in the context of gain-of-CXCR4-function mutation found in WHIM Syndrome (WS), which has been reproduced in mouse model and that causes defects in MPPs and deregulated expression of CXCL12 in BM. Access to BM and blood samples from healthy and WS subjects to translate results to human is an asset for the project.