BackgroundPulmonary arterial hypertension (PAH) is a vasculopathy characterized by enhanced pulmonary artery smooth muscle cell (PASMC) proliferation and suppressed apoptosis. This results in both increase in pulmonary arterial pressure and pulmonary vascular resistance. Recent studies have shown the implication of the signal transducer and activator of transcription 3 (STAT3)/bone morphogenetic protein receptor 2 (BMPR2)/peroxisome proliferator‐activated receptor gamma (PPARγ) in PAH. STAT3 activation induces BMPR2 downregulation, decreasing PPARγ, which both contribute to the proproliferative and antiapoptotic phenotype seen in PAH. In chondrocytes, activation of this axis has been attributed to the advanced glycation end‐products receptor (RAGE). As RAGE is one of the most upregulated proteins in PAH patients' lungs and a strong STAT3 activator, we hypothesized that by activating STAT3, RAGE induces BMPR2 and PPARγ downregulation, promoting PAH‐PASMC proliferation and resistance to apoptosis.Methods and ResultsIn vitro, using PASMCs isolated from PAH and healthy patients, we demonstrated that RAGE is overexpressed in PAH‐PASMC (6‐fold increase), thus inducing STAT3 activation (from 10% to 40% positive cells) and decrease in BMPR2 and PPARγ levels (>50% decrease). Pharmacological activation of RAGE in control cells by S100A4 recapitulates the PAH phenotype (increasing RAGE by 6‐fold, thus activating STAT3 and decreasing BMPR2 and PPARγ). In both conditions, this phenotype is totally reversed on RAGE inhibition. In vivo, RAGE inhibition in monocrotaline‐ and Sugen‐induced PAH demonstrates therapeutic effects characterized by PA pressure and right ventricular hypertrophy decrease (control rats have an mPAP around 15 mm Hg, PAH rats have an mPAP >40 mm Hg, and with RAGE inhibition, mPAP decreases to 20 and 28 mm Hg, respectively, in MCT and Sugen models). This was associated with significant improvement in lung perfusion and vascular remodeling due to decrease in proliferation (>50% decrease) and BMPR2/PPARγ axis restoration (increased by ≥60%).ConclusionWe have demonstrated the implications of RAGE in PAH etiology. Thus, RAGE constitutes a new attractive therapeutic target for PAH.

Non-pathogenic environmental microbial exposures during pregnancy can be transplacentally transcribed into beneficial immune training signals for the developing fetus.These signals equip offspring for more rapid adaptation to the microbe-rich postnatal environment by optimising immunoregulatory innate cell function. We have previously identified that maternal treatment with a microbial-derived therapeutic (OM-85) can protect offspring against allergic airways inflammation. Here, we show that oral treatment of pregnant mice with OM-85 induces transplacental signals that manifest in fetal bone marrow as an enriched population of conventional dendric cells (cDC) displaying enhanced functional maturation. Moreover, the myeloid progenitor populations directly upstream of this cDC pool were significantly boosted in response to maternal treatment. Transcriptomic analysis of fetal bone marrow identified maternal OM-85-induced activation of X-box binding protein 1 (XBP1), with upregulation of active XBP1 restricted to cDC precursors. These data provide direct evidence that transplacental immune training with a microbial-derived therapeutic can accelerate functional immune competence of the fetal bone marrow myeloid compartment. KTM, PGH and DHS designed the study. KTM, MB , NMS and JFLJ performed the experiments. KTM, ACJ, MB and DHS analysed the data. PAS and AB contributed to the project design and discussions on data interpretation. KTM, PAS, PGH and DHS wrote the manuscript. All authors reviewed the final version of the manuscript.

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Background Hypersensitivity pneumonitis (HP) is an interstitial lung disease characterized by antigen-triggered neutrophilic exacerbations. Although CD4+ T cells are sufficient for HP pathogenesis, this never translated into efficient T cell-specific therapies. Increasing evidence shows that B cells also play decisive roles in HP. Here, we aimed to further define the respective contributions of B and T cells in subacute experimental HP. Methods Mice were subjected to a protocol of subacute exposure to the archaeon Methanosphaera stadmanae to induce experimental HP. Using models of adoptive transfers of B cells and T cells in Rag1-deficient mice and of B cell-specific S1P1 deletion, we assessed the importance of B cells in the development of HP by evaluating inflammation in bronchoalveolar lavage fluid. We also aimed to determine if injected antibodies targeting B and/or T cells could alleviate HP exacerbations using a therapeutic course of intervention. Results Even though B cells are not sufficient to induce HP, they strongly potentiate CD4+ T cell-induced HP‑associated neutrophilic inflammation in the airways. However, the reduction of 85% of lung B cells in mice with a CD19-driven S1P1 deletion does not dampen HP inflammation, suggesting that lung B cells are not necessary in large numbers to sustain local inflammation. Finally, we found that injecting antibodies targeting B cells after experimental HP was induced does not dampen neutrophilic exacerbation. Yet, injection of antibodies directed against B cells and T cells yielded a potent 76% inhibition of neutrophilic accumulation in the lungs. This inhibition occurred despite partial, sometimes mild, depletion of B cells and T cells subsets. Conclusions Although B cells are required for maximal inflammation in subacute experimental HP, partial reduction of B cells fails to reduce HP-associated inflammation by itself. However, co-modulation of T cells and B cells yields enhanced inhibition of HP exacerbation caused by an antigenic rechallenge.

IntroductionAt lung mucosal surfaces, immune cells must initiate inflammatory response against pathogen without inducing tissue damage. Loss of this equilibrium can lead to acute respiratory distress syndrome (ARDS), a severe lung inflammatory disease characterized by excessive inflammation and dysregulation of anti-inflammatory pathways.MethodsTo investigate the role of anti-inflammatory pathway CD200/CD200R in lung inflammatory response, we administered LPS intratracheally in CD200 KO and wild type (WT) rats. Inflammation was evaluated using bronchoalveolar lavage (BAL) cellularity. Lung injury was measured by total protein level in BAL fluid, and levels of proinflammatory cytokines (TNF, IL-6) and chemokines (CXCL2, CCL2) were determined in BAL supernatants. In a second series of experiments, recombinant CD200Fc was administered to KO rats to restore the anti-inflammatory response.ResultsAt baseline, CD200 KO rats did not show sign of inflammation, however KO rats had lower number of alveolar macrophages. In addition, LPS administration induced greater pulmonary edema in CD200 KO rats. This was accompanied with a higher recruitment of neutrophils as well as levels of TNF, IL-6, CXCL2, and CCL2 in BAL compared to WT rats. CD200Fc administration in KO rats reduced neutrophil accumulation and TNF and CXCL2 levels in BAL. Interestingly, the increased inflammatory response of CD200 KO rats could be attributed to greater activation potential of alveolar macrophages with higher levels of ERK and P-ERK MAPK.ConclusionThis study shows that lung inflammatory response is exacerbated in absence of CD200 in an experimental model of ARDS in rats. In addition, CD200/CD200R pathway shows selective regulation of acute lung inflammation and cannot completely abrogate the complex LPS-induced inflammatory response. However, addition of CD200 agonist in a multi-target therapy strategy could have beneficial impacts.

Airway inflammation is a defense mechanism against inhaled agents characterized by infiltration of circulating immune cells. Given the inconsistent cellular identification across pre-clinical rat model, we have developed a flow cytometry panel of six colors to characterize macrophages subsets, lymphocytes and granulocytes in bronchoalveolar lavage fluid (BAL). Rats were challenged with intratracheal instillation of lipopolysaccharide (LPS). BAL were harvested 24 h after one LPS exposure in rats.This flow cytometry panel involve the description of macrophage subsets, T and B lymphocytes and neutrophils, which are central to airway immune responses, as based on scientific literature. By using a relatively small number of parameters to identify multiple cell types, additional parameters can be used for project/diseasespecific activation markers.

Alveolar Macrophages are central regulators of pulmonary immune responses, yet their regulatory functions are misunderstood. CD200, a transmembrane protein, and its receptor (CD200R) play a critical role in the resolution of inflammation, but their role in asthma is still unclear. CD200 is expressed on many cell types such as leukocytes, T cells, B cells, epithelial cells, and endothelial cells, while CD200R is expressed exclusively on myeloid cells including alveolar macrophages (AM) and mast cells (MC), which play a key role in asthma. Recent study showed that leukocytes from asthmatic patients express lower level of CD200 during exacerbations, suggesting a dysregulation of CD200 pathway. Thus, we investigated the modulation of CD200 expression on AM following allergen challenge and the inhibition of MC functions by AM CD200. Expression of CD200 and CD200R were measured on AM from naive and sensitized rats before and after allergen challenge using flow cytometry. AM modulation of antigen‐stimulated MC was investigated in vitro using co‐culture and MC degranulation and cytokine production. AM from naive and sensitized rats express similar level of CD200. However, allergen exposure increased CD200 expression on AM of naïve rats (that do not develop signs of asthma), but not on AM of sensitized rats (that develop experimental asthma). No difference between groups was observed for CD200R expression. Naive AM inhibited antigen‐stimulated MC degranulation and cytokine production. The addition of neutralizing anti‐CD200R antibody abrogated AM inhibitory effects. These results suggest that modulation of CD200 expression on AM could have an important role in asthma regulation.

INTRODUCTION Acute respiratory distress syndrome (ARDS) is a severe lung inflammatory disease caused by a variety of precipitants, including SARS-CoV-2 (COVID-19). In addition to excessive inflammation, ARDS is characterised by the dysregulation of anti-inflammatory pathways, including CD200/CD200R pathway. OBJECTIVE To investigate the role of CD200/CD200R pathway in lung ARDS inflammatory response. METHODS LPS was administered intratracheally to induce ARDS in Sprague-Dawley CD200 KO and wild type (WT) rats. Inflammation was evaluated using bronchoalveolar lavage (BAL) cellularity. Lung injury was measured by total protein level in BAL fluid, and levels of proinflammatory cytokines (TNF, IL-6) and chemokines (CXCL2, CCL2) were determined in BAL supernatants. In a second experiment, recombinant CD200Fc was administered to KO rats to restore the anti-inflammatory response. RESULTS Although there was no difference in total BAL cell counts at 3 h, cell recruitment was greater in CD200 KO rats given the low cell number in naïve KO rats. BAL of KO rats had higher levels of TNF, IL-6, CXCL2, and CCL2 compared to WT rats. Total protein level in BAL was higher in CD200 KO rats, implying more pronounced pulmonary edema. CD200Fc administration in KO rats significantly decreased levels of TNF and CCL2 in BAL, suggesting an attenuation of the inflammatory response. CONCLUSION This study shows that ARDS inflammatory response is exacerbated in absence of CD200 in an experimental model of ARDS in rats and that CD200 supplementation alleviates this phenotype. Further analyses will be needed to better understand the contribution of different cell types expressing CD200 to control lung inflammatory response resulting from ARDS. Supported by grant by CIHR and IUCPQ fondation.