Effects of different amosite preparations on macrophages, lung damages, and autoimmunity.

The underlying mechanisms of asbestos-related autoimmunity are poorly understood. As the size, surface reactivity, and free radical activity of asbestos particles are considered crucial regarding the health effects, this study aims to compare the effects of exposure to pristine amosite (pAmo) or milled amosite (mAmo) particles on lung damage, autoimmunity, and macrophage phenotype. Four months after lung exposure to 0.1 mg of amosite, BAL levels of lactate dehydrogenase, protein, free DNA, CCL2, TGF-ß1, TIMP-1, and immunoglobulin A of pAmo-exposed C57Bl/6 mice were increased when compared to fluids from control- and mAmo-exposed mice. Effects in pAmo-exposed mice were associated with lung fibrosis and autoimmunity including anti-double-strand DNA autoantibody production. mAmo or pAmo at 20 µg/cm2 induced a pro-inflammatory phenotype characterized by a significant increase in TNFα and IL-6 secretion on human monocyte-derived macrophages (MDMs). mAmo and pAmo exposure induced a decrease in the efferocytosis capacities of MDMs, whereas macrophage abilities to phagocyte fluorescent beads were unchanged when compared to control MDMs. mAmo induced IL-6 secretion and reduced the percentage of MDMs expressing MHCII and CD86 markers involved in antigen and T-lymphocyte stimulation. By contrast, pAmo but not mAmo activated the NLRP3 inflammasome, as evaluated through quantification of caspase-1 activity and IL-1ß secretion. Our results demonstrated that long-term exposure to pAmo may induce significant lung damage and autoimmune effects, probably through an alteration of macrophage phenotype, supporting in vivo the higher toxicity of entire amosite (pAmo) with respect to grinded amosite. However, considering their impact on efferocytosis and co-stimulation markers, mAmo effects should not be neglected. KEY MESSAGES: Lung fibrosis and autoimmunity induced by amosite particles depend on their physicochemical characteristics (size and surface) Inhalation exposure of mice to pristine amosite fibers is associated with lung fibrosis and autoimmunity Anti-dsDNA antibody is a marker of autoimmunity in mice exposed to pristine amosite fibers Activation of lung mucosa-associated lymphoid tissue, characterized by IgA production, after exposure to pristine amosite fibers Pristine and milled amosite particle exposure reduced the efferocytosis capacity of human-derived macrophages.

Distinct Pro-Inflammatory Mechanisms Elicited by Short and Long Amosite Asbestos Fibers in Macrophages.

While exposure to long amphibolic asbestos fibers (L > 10 µm) results in the development of severe diseases including inflammation, fibrosis, and mesothelioma, the pathogenic activity associated with short fibers (L < 5 µm) is less clear. By exposing murine macrophages to short (SFA) or long (LFA) fibers of amosite asbestos different in size and surface chemistry, we observed that SFA internalization resulted in pyroptotic-related immunogenic cell death (ICD) characterized by the release of the pro-inflammatory damage signal (DAMP) IL-1α after inflammasome activation and gasdermin D (GSDMD)-pore formation. In contrast, macrophage responses to non-internalizable LFA were associated with tumor necrosis factor alpha (TNF-α) release, caspase-3 and -7 activation, and apoptosis. SFA effects exclusively resulted from Toll-like receptor 4 (TLR4), a pattern-recognition receptor (PRR) recognized for its ability to sense particles, while the response to LFA was elicited by a multifactorial ignition system involving the macrophage receptor with collagenous structure (SR-A6 or MARCO), reactive oxygen species (ROS) cascade, and TLR4. Our findings indicate that asbestos fiber size and surface features play major roles in modulating ICD and inflammatory pathways. They also suggest that SFA are biologically reactive in vitro and, therefore, their inflammatory and toxic effects in vivo should not be underestimated.

Usefulness of FAPα assessment in bronchoalveolar lavage as a marker of fibrogenesis: results of a preclinical study and first report in patients with idiopathic pulmonary fibrosis.

BACKGROUND: Fibroblast activation protein-α (FAPα) is a marker of activated fibroblasts that can be selectively targeted by an inhibitor (FAPI) and visualised by PET/CT imaging. We evaluated whether the measurement of FAPα in bronchoalveolar lavage fluids (BALF) and the uptake of FAPI by PET/CT could be used as biomarkers of fibrogenesis. METHODS: The dynamics of lung uptake of 18F-labeled FAPI ([18F]FAPI-74) was assessed in the bleomycin mouse model at various time points and using different concentrations of bleomycin by PET/CT. FAPα was measured in BALFs from these bleomycin-treated and control mice. FAPα levels were also assessed in BALFs from controls and patients with idiopathic pulmonary fibrosis (IPF). RESULTS: Bleomycin-treated mice presented a significantly higher uptake of [18F]FAPI-74 during lung fibrinogenesis (days 10 and 16 after instillation) compared to control mice. No significant difference was observed at initial inflammatory phase (3 days) and when fibrosis was already established (28 days). [18F]FAPI-74 tracer was unable to show a dose-response to bleomycin treatment. On the other hand, BALF FAPα levels were steeply higher in bleomycin-treated mice at day 10 and a significant dose-response effect was observed. Moreover, FAPα levels were strongly correlated with lung fibrosis as measured by the modified Aschroft histological analysis, hydroxyproline and the percentage of weight loss. Importantly, higher levels of FAPα were observed in IPF patients where the disease was progressing as compared to stable patients and controls. Moreover, patients with FAPα BALF levels higher than 192.5 pg/mL presented a higher risk of progression, transplantation or death compared to patients with lower levels. CONCLUSIONS: Our preclinical data highlight a specific increase of [18F]FAPI-74 lung uptake during the fibrotic phase of the bleomycin murine model. The measurement of FAPα in BALF appears to be a promising marker of the fibrotic activity in preclinical models of lung fibrosis and in IPF patients. Further studies are required to confirm the role of FAPα in BALF as biomarker of IPF activity and assess the relationship between FAPα levels in BALF and [18F]FAPI-74 uptake on PET/CT in patients with fibrotic lung disease.

B-1 cells in immunotoxicology: Mechanisms underlying their response to chemicals and particles.

Since their discovery nearly 40 years ago, B-1 cells have continued to challenge the boundaries between innate and adaptive immunity, as well as myeloid and lymphoid functions. This B-cell subset ensures early immunity in neonates before the development of conventional B (B-2) cells and respond to immune injuries throughout life. B-1 cells are multifaceted and serve as natural- and induced-antibody-producing cells, phagocytic cells, antigen-presenting cells, and anti-/pro-inflammatory cytokine-releasing cells. This review retraces the origin of B-1 cells and their different roles in homeostatic and infectious conditions before focusing on pollutants comprising contact-sensitivity-inducing chemicals, endocrine disruptors, aryl hydrocarbon receptor (AHR) ligands, and reactive particles.

Gasdermin D membrane pores orchestrate IL-1α secretion from necrotic macrophages after NFS-rich silica exposure.

IL-1α is an intracellular danger signal (DAMP) released by macrophages contributing to the development of silica-induced lung inflammation. The exact molecular mechanism orchestrating IL-1α extracellular release from particle-exposed macrophages is still unclear. To delineate this process, murine J774 and bone-marrow derived macrophages were exposed to increasing concentrations (1-40 cm2/ml) of a set of amorphous and crystalline silica particles with different surface chemical features. In particular, these characteristics include the content of nearly free silanols (NFS), a silanol population responsible for silica cytotoxicity recently identified. We first observed de novo stocks of IL-1α in macrophages after silica internalization regardless of particle physico-chemical characteristics and cell stress. IL-1α intracellular production and accumulation were observed by exposing macrophages to biologically-inert or cytotoxic crystalline and amorphous silicas. In contrast, only NFS-rich reactive silica particles triggered IL-1α release into the extracellular milieu from necrotic macrophages. We demonstrate that IL-1α is actively secreted through the formation of gasdermin D (GSDMD) pores in the plasma membrane and not passively released after macrophage plasma membrane lysis. Our findings indicate that the GSDMD pore-dependent secretion of IL-1α stock from macrophages solely depends on cytotoxicity induced by NFS-rich silica. This new regulated process represents a key first event in the mechanism of silica toxicity, suitable to refine the existing adverse outcome pathway (AOP) for predicting the inflammatory activity of silicas.

Bleomycin-induced lung injury: Revisiting an old tool to model group III PH associated with pulmonary fibrosis.

Pulmonary hypertension (PH) is a chronic disorder of the pulmonary circulation that often associates with other respiratory diseases (i.e., group III PH), leading to worsened symptoms and prognosis, notably when combined with interstitial lung diseases such as pulmonary fibrosis (PF). PH may lead to right ventricular (RV) failure, which accounts for a substantial part of the mortality in chronic lung disease patients. The disappointing results of pulmonary arterial hypertension (PAH)-related therapies in patients with PF emphasize the need to better understand the pathophysiologic mechanisms that drive PH development and progression in this specific setting. In this work, we validated an animal model of group III PH associated with PF (PH-PF), by using bleomycin (BM) intratracheal instillation and characterizing the nature of induced lung and vascular remodeling, including the influence on RV structure and function. To our knowledge, this is the first work describing this dose of BM in Sprague Dawley rats and the effects upon the heart and lungs, using different techniques such as echocardiography, heart catheterization, and histology. Our data shows the successful implementation of a rat model that mimics combined PF-PH, with most features seen in the equivalent human disease, such as lung and arterial remodeling, increased mPAP and RV dysfunction.

Disruption of GCN2 Pathway Aggravates Vascular and Parenchymal Remodeling during Pulmonary Fibrosis.

Pulmonary fibrosis (PF) and pulmonary hypertension (PH) are chronic diseases of the pulmonary parenchyma and circulation, respectively, which may coexist, but underlying mechanisms remain elusive. Mutations in the GCN2 (general control nonderepressible 2) gene (EIF2AK4 [eukaryotic translation initiation factor 2 alpha kinase 4]) were recently associated with pulmonary veno-occlusive disease. The aim of this study is to explore the involvement of the GCN2/eIF2α (eukaryotic initiation factor 2α) pathway in the development of PH during PF, in both human disease and in a laboratory animal model. Lung tissue from patients with PF with or without PH was collected at the time of lung transplantation, and control tissue was obtained from tumor resection surgery. Experimental lung disease was induced in either male wild-type or EIF2AK4-mutated Sprague-Dawley rats, randomly receiving a single intratracheal instillation of bleomycin or saline. Hemodynamic studies and organ collection were performed 3 weeks after instillation. Only significant results (P < 0.05) are presented. In PF lung tissue, GCN2 protein expression was decreased compared with control tissue. GCN2 expression was reduced in CD31+ endothelial cells. In line with human data, GCN2 protein expression was decreased in the lung of bleomycin rats compared with saline. EIF2AK4-mutated rats treated with bleomycin showed increased parenchymal fibrosis (hydroxyproline concentrations) and vascular remodeling (media wall thickness) as well as increased right ventricular systolic pressure compared with wild-type animals. Our data show that GCN2 is dysregulated in both humans and in an animal model of combined PF and PH. The possibility of a causative implication of GCN2 dysregulation in PF and/or PH development should be further studied.

BRAFV600E Expression in Thyrocytes Causes Recruitment of Immunosuppressive STABILIN-1 Macrophages.

Papillary thyroid carcinoma (PTC) is the most frequent histological subtype of thyroid cancers (TC), and BRAFV600E genetic alteration is found in 60% of this endocrine cancer. This oncogene is associated with poor prognosis, resistance to radioiodine therapy, and tumor progression. Histological follow-up by anatomo-pathologists revealed that two-thirds of surgically-removed thyroids do not present malignant lesions. Thus, continued fundamental research into the molecular mechanisms of TC downstream of BRAFV600E remains central to better understanding the clinical behavior of these tumors. To study PTC, we used a mouse model in which expression of BRAFV600E was specifically switched on in thyrocytes by doxycycline administration. Upon daily intraperitoneal doxycycline injection, thyroid tissue rapidly acquired histological features mimicking human PTC. Transcriptomic analysis revealed major changes in immune signaling pathways upon BRAFV600E induction. Multiplex immunofluorescence confirmed the abundant recruitment of macrophages, among which a population of LYVE-1+/CD206+/STABILIN-1+ was dramatically increased. By genetically inactivating the gene coding for the scavenger receptor STABILIN-1, we showed an increase of CD8+ T cells in this in situ BRAFV600E-dependent TC. Lastly, we demonstrated the presence of CD206+/STABILIN-1+ macrophages in human thyroid pathologies. Altogether, we revealed the recruitment of immunosuppressive STABILIN-1 macrophages in a PTC mouse model and the interest to further study this macrophage subpopulation in human thyroid tissues.

Think Beyond Particle Cytotoxicity: When Self-Cellular Components Released After Immunogenic Cell Death Explain Chronic Disease Development.

The prolonged perturbation of the immune system following the release of a plethora of self-molecules (known as damage-associated molecular patterns, DAMPs) by stressed or dying cells triggers acute and chronic pathological responses. DAMPs are commonly released after plasma membrane damage or complete rupture due to immunogenic cell death (ICD), upon numerous stressors including infectious and toxic agents. The set of DAMPs released after ICD include mature proinflammatory cytokines and alarmins, but also polymeric macromolecules. These self-intracellular components are recognized by injured and healthy surrounding cells via innate receptors, and induce upregulation of stress-response mechanisms, including inflammation. In this review, by overstepping the simple toxicological evaluation, we apply ICD and DAMP concepts to silica cytotoxicity, providing new insights on the mechanisms driving the progress and/or the exacerbation of certain SiO2-related pathologies. Finally, by proposing self-DNA as new crucial DAMP, we aim to pave the way for the development of innovative and easy-to-perform predictive tests to better identify the hazard of fine and ultrafine silica particles. Importantly, such mechanisms could be extended to nano/micro plastics and diesel particles, providing strategic advice and reports on their health issues.

The EXIMIOUS project-Mapping exposure-induced immune effects: connecting the exposome and the immunome.

Immune-mediated, noncommunicable diseases-such as autoimmune and inflammatory diseases-are chronic disorders, in which the interaction between environmental exposures and the immune system plays an important role. The prevalence and societal costs of these diseases are rising in the European Union. The EXIMIOUS consortium-gathering experts in immunology, toxicology, occupational health, clinical medicine, exposure science, epidemiology, bioinformatics, and sensor development-will study eleven European study populations, covering the entire lifespan, including prenatal life. Innovative ways of characterizing and quantifying the exposome will be combined with high-dimensional immunophenotyping and -profiling platforms to map the immune effects (immunome) induced by the exposome. We will use two main approaches that "meet in the middle"-one starting from the exposome, the other starting from health effects. Novel bioinformatics tools, based on systems immunology and machine learning, will be used to integrate and analyze these large datasets to identify immune fingerprints that reflect a person's lifetime exposome or that are early predictors of disease. This will allow researchers, policymakers, and clinicians to grasp the impact of the exposome on the immune system at the level of individuals and populations.

Mouse innate-like B-1 lymphocytes promote inhaled particle-induced in vitro granuloma formation and inflammation in conjunction with macrophages.

The current paradigm for explaining lung granulomatous diseases induced by inhaled particles is mainly based on macrophages. This mechanism is now challenging because B lymphocytes also infiltrate injured tissue, and the deficiency in B lymphocytes is associated with limited lung granulomas in silica-treated mice. Here, we investigated how B lymphocytes respond to micro- and nanoparticles by combining in vivo and in vitro mouse models. We first demonstrated that innate-like B-1 lymphocytes (not conventional B-2 lymphocytes or plasma cells) specifically accumulated during granuloma formation in mice instilled with crystalline silica (DQ12, 2.5 mg/mouse) and carbon nanotubes (CNT Mitsui, 0.2 mg/mouse). In comparison to macrophages, peritoneal B-1 lymphocytes purified from naïve mice were resistant to the pyroptotic activity of reactive particles (up to 1 mg/mL) but clustered to establish in vitro cell/particle aggregates. Mouse B-1 lymphocytes (not B-2 lymphocytes) in coculture with macrophages and CNT (0.1 µg/mL) organized three-dimensional spheroid structures in Matrigel and stimulated the release of TIMP-1. Furthermore, purified B-1 lymphocytes are sensitive to nanosilica toxicity through radical generation in culture. Nanosilica-exposed B-1 lymphocytes released proinflammatory cytokines and alarmins. In conclusion, our data indicate that in addition to macrophages, B-1 lymphocytes participate in micrometric particle-induced granuloma formation and display inflammatory functions in response to nanoparticles.

Interpreting Immunoregulation in Lung Fibrosis: A New Branch of the Immune Model.

Immunostimulation is recognized as an important contribution in lung fibrosis in some animal models and patient subsets. With this review, we illustrate an additional scenario covering the possible implication of immunoregulation during fibrogenesis. Available animal and human data indicate that pulmonary fibrosis also includes diverse and discrete immunoregulating populations comprising regulatory lymphocytes (T and B regs) and myeloid cells (immunosuppressive macrophages and myeloid-derived suppressive cells; MDSC). They are initially recruited to limit the establishment of deleterious inflammation but participate in the development of lung fibrosis by producing immunoregulatory mediators (mainly TGF-ß1 and IL-10) that directly or indirectly stimulate fibroblasts and matrix protein deposition. The existence of this silent immunoregulatory environment sustains an alternative mechanism of fibrosis that explains why in some conditions neither pro-inflammatory cytokine deficiency nor steroid and immunosuppressive therapies limit lung fibrosis. Therefore, the persistent presence of immunoregulation is an important parameter to consider for refining therapeutical strategies in lung fibrotic disorders under non-immunostimulatory conditions.

Monocytic Ontogeny of Regenerated Macrophages Characterizes the Mesotheliomagenic Responses to Carbon Nanotubes.

Macrophages are not only derived from circulating blood monocytes or embryonic precursors but also expand by proliferation. The origin determines macrophage fate and functions in steady state and pathological conditions. Macrophages predominantly infiltrate fibre-induced mesothelioma tumors and contribute to cancer development. Here, we revealed their ontogeny by comparing the response to needle-like mesotheliomagenic carbon nanotubes (CNT-7) with tangled-like non-mesotheliomagenic CNT-T. In a rat peritoneal cavity model of mesothelioma, both CNT induced a rapid macrophage disappearance reaction (MDR) of MHCIIlow resident macrophages generating an empty niche available for macrophage repopulation. Macrophage depletion after mesotheliomagenic CNT-7 was followed by a substantial inflammatory reaction, and macrophage replenishment completed after 7 days. Thirty days after non-mesotheliomagenic CNT-T, macrophage repopulation was still incomplete and accompanied by a limited inflammatory reaction. Cell depletion experiments, flow cytometry and RNA-seq analysis demonstrated that, after mesotheliomagenic CNT-7 exposure, resident macrophages were mainly replaced by an influx of monocytes, which differentiated locally into MHCIIhigh inflammatory macrophages. In contrast, the low inflammatory response induced by CNT-T was associated by the accumulation of self-renewing MHCIIlow macrophages that initially derive from monocytes. In conclusion, the mesotheliomagenic response to CNT specifically relies on macrophage niche recolonization by monocyte-derived inflammatory macrophages. In contrast, the apparent homeostasis after non-mesotheliomagenic CNT treatment involves a macrophage regeneration by proliferation. Macrophage depletion and repopulation are thus decisive events characterizing the carcinogenic activity of particles and fibres.

Chronic Pseudomonas aeruginosa Lung Infection Is IL-1R Independent, but Relies on MyD88 Signaling.

Cystic fibrosis is associated with chronic Pseudomonas aeruginosa colonization and inflammation. The role of MyD88, the shared adapter protein of the proinflammatory TLR and IL-1R families, in chronic P. aeruginosa biofilm lung infection is unknown. We report that chronic lung infection with the clinical P. aeruginosa RP73 strain is associated with uncontrolled lung infection in complete MyD88-deficient mice with epithelial damage, inflammation, and rapid death. Then, we investigated whether alveolar or myeloid cells contribute to heightened sensitivity to infection. Using cell-specific, MyD88-deficient mice, we uncover that the MyD88 pathway in myeloid or alveolar epithelial cells is dispensable, suggesting that other cell types may control the high sensitivity of MyD88-deficient mice. By contrast, IL-1R1-deficient mice control chronic P. aeruginosa RP73 infection and IL-1ß Ab blockade did not reduce host resistance. Therefore, the IL-1R1/MyD88 pathway is not involved, but other IL-1R or TLR family members need to be investigated. Our data strongly suggest that IL-1 targeted neutralizing therapies used to treat inflammatory diseases in patients unlikely reduce host resistance to chronic P. aeruginosa infection.

Digital Image Analysis of Picrosirius Red Staining: A Robust Method for Multi-Organ Fibrosis Quantification and Characterization.

Current understanding of fibrosis remains incomplete despite the increasing burden of related diseases. Preclinical models are used to dissect the pathogenesis and dynamics of fibrosis, and to evaluate anti-fibrotic therapies. These studies require objective and accurate measurements of fibrosis. Existing histological quantification methods are operator-dependent, organ-specific, and/or need advanced equipment. Therefore, we developed a robust, minimally operator-dependent, and tissue-transposable digital method for fibrosis quantification. The proposed method involves a novel algorithm for more specific and more sensitive detection of collagen fibers stained by picrosirius red (PSR), a computer-assisted segmentation of histological structures, and a new automated morphological classification of fibers according to their compactness. The new algorithm proved more accurate than classical filtering using principal color component (red-green-blue; RGB) for PSR detection. We applied this new method on established mouse models of liver, lung, and kidney fibrosis and demonstrated its validity by evidencing topological collagen accumulation in relevant histological compartments. Our data also showed an overall accumulation of compact fibers concomitant with worsening fibrosis and evidenced topological changes in fiber compactness proper to each model. In conclusion, we describe here a robust digital method for fibrosis analysis allowing accurate quantification, pattern recognition, and multi-organ comparisons useful to understand fibrosis dynamics.

HIF-1α is a key mediator of the lung inflammatory potential of lithium-ion battery particles.

BACKGROUND: Li-ion batteries (LIB) are increasingly used worldwide. They are made of low solubility micrometric particles, implying a potential for inhalation toxicity in occupational settings and possibly for consumers. LiCoO2 (LCO), one of the most used cathode material, induces inflammatory and fibrotic lung responses in mice. LCO also stabilizes hypoxia-inducible factor (HIF) -1α, a factor implicated in inflammation, fibrosis and carcinogenicity. Here, we investigated the role of cobalt, nickel and HIF-1α as determinants of toxicity, and evaluated their predictive value for the lung toxicity of LIB particles in in vitro assays. RESULTS: By testing a set of 5 selected LIB particles (LCO, LiNiMnCoO2, LiNiCoAlO2) with different cobalt and nickel contents, we found a positive correlation between their in vivo lung inflammatory activity, and (i) Co and Ni particle content and their bioaccessibility and (ii) the stabilization of HIF-1α in the lung. Inhibition of HIF-1α with chetomin or PX-478 blunted the lung inflammatory response to LCO in mice. In IL-1ß deficient mice, HIF-1α was the upstream signal of the inflammatory lung response to LCO. In vitro, the level of HIF-1α stabilization induced by LIB particles in BEAS-2B cells correlated with the intensity of lung inflammation induced by the same particles in vivo. CONCLUSIONS: We conclude that HIF-1α, stabilized in lung cells by released Co and Ni ions, is a mechanism-based biomarker of lung inflammatory responses induced by LIB particles containing Co/Ni. Documenting the Co/Ni content of LIB particles, their bioaccessibility and their capacity to stabilize HIF-1α in vitro can be used to predict the lung inflammatory potential of LIB particles.

The puzzling issue of silica toxicity: are silanols bridging the gaps between surface states and pathogenicity?

BACKGROUND: Silica continues to represent an intriguing topic of fundamental and applied research across various scientific fields, from geology to physics, chemistry, cell biology, and particle toxicology. The pathogenic activity of silica is variable, depending on the physico-chemical features of the particles. In the last 50 years, crystallinity and capacity to generate free radicals have been recognized as relevant features for silica toxicity. The 'surface' also plays an important role in silica toxicity, but this term has often been used in a very general way, without defining which properties of the surface are actually driving toxicity. How the chemical features (e.g., silanols and siloxanes) and configuration of the silica surface can trigger toxic responses remains incompletely understood. MAIN BODY: Recent developments in surface chemistry, cell biology and toxicology provide new avenues to improve our understanding of the molecular mechanisms of the adverse responses to silica particles. New physico-chemical methods can finely characterize and quantify silanols at the surface of silica particles. Advanced computational modelling and atomic force microscopy offer unique opportunities to explore the intimate interactions between silica surface and membrane models or cells. In recent years, interdisciplinary research, using these tools, has built increasing evidence that surface silanols are critical determinants of the interaction between silica particles and biomolecules, membranes, cell systems, or animal models. It also has become clear that silanol configuration, and eventually biological responses, can be affected by impurities within the crystal structure, or coatings covering the particle surface. The discovery of new molecular targets of crystalline as well as amorphous silica particles in the immune system and in epithelial lung cells represents new possible toxicity pathways. Cellular recognition systems that detect specific features of the surface of silica particles have been identified. CONCLUSIONS: Interdisciplinary research bridging surface chemistry to toxicology is progressively solving the puzzling issue of the variable toxicity of silica. Further interdisciplinary research is ongoing to elucidate the intimate mechanisms of silica pathogenicity, to possibly mitigate or reduce surface reactivity.

Cationic Nanoliposomes Are Efficiently Taken up by Alveolar Macrophages but Have Little Access to Dendritic Cells and Interstitial Macrophages in the Normal and CpG-Stimulated Lungs.

The purpose of this study was to assess whether cationic nanoliposomes could address tumor vaccines to dendritic cells in the lungs in vivo. Nanoliposomes were prepared using a cationic lipid, dimethylaminoethanecarbamoyl-cholesterol (DC-cholesterol) or dioleoyltrimethylammoniumpropane (DOTAP), and dipalmitoylphosphatidylcholine (DPPC), the most abundant phospholipid in lung surfactant. The liposomes presented a size below 175 nm and they effectively entrapped tumor antigens, an oligodeoxynucletotide containing CpG motifs (CpG) and the fluorescent dye calcein used as a tracer. Although the liposomes could permanently entrap a large fraction of the actives, they could not sustain their release in vitro. Liposomes made of DOTAP were safe to respiratory cells in vitro, while liposomes composed of DC-cholesterol were cytotoxic. DOTAP nanoliposomes were mainly taken up by alveolar macrophages following delivery to the lungs in mice. Few dendritic cells took up the liposomes, and interstitial macrophages did not take up liposomal calcein more than they took up soluble calcein. Stimulation of the innate immune system using liposomal CpG strongly enhanced uptake of calcein liposomes by all phagocytes in the lungs. Although a small percentage of dendritic cells took up the nanoliposomes, alveolar macrophages represented a major barrier to dendritic cell access in the lungs.

Aggregated Tau activates NLRP3-ASC inflammasome exacerbating exogenously seeded and non-exogenously seeded Tau pathology in vivo.

Brains of Alzheimer's disease patients are characterized by the presence of amyloid plaques and neurofibrillary tangles, both invariably associated with neuroinflammation. A crucial role for NLRP3-ASC inflammasome [NACHT, LRR and PYD domains-containing protein 3 (NLRP3)-Apoptosis-associated speck-like protein containing a CARD (ASC)] in amyloid-beta (Aß)-induced microgliosis and Aß pathology has been unequivocally identified. Aß aggregates activate NLRP3-ASC inflammasome (Halle et al. in Nat Immunol 9:857-865, 2008) and conversely NLRP3-ASC inflammasome activation exacerbates amyloid pathology in vivo (Heneka et al. in Nature 493:674-678, 2013), including by prion-like ASC-speck cross-seeding (Venegas et al. in Nature 552:355-361, 2017). However, the link between inflammasome activation, as crucial sensor of innate immunity, and Tau remains unexplored. Here, we analyzed whether Tau aggregates acting as prion-like Tau seeds can activate NLRP3-ASC inflammasome. We demonstrate that Tau seeds activate NLRP3-ASC-dependent inflammasome in primary microglia, following microglial uptake and lysosomal sorting of Tau seeds. Next, we analyzed the role of inflammasome activation in prion-like or templated seeding of Tau pathology and found significant inhibition of exogenously seeded Tau pathology by ASC deficiency in Tau transgenic mice. We furthermore demonstrate that chronic intracerebral administration of the NLRP3 inhibitor, MCC950, inhibits exogenously seeded Tau pathology. Finally, ASC deficiency also decreased non-exogenously seeded Tau pathology in Tau transgenic mice. Overall our findings demonstrate that Tau-seeding competent, aggregated Tau activates the ASC inflammasome through the NLRP3-ASC axis, and we demonstrate an exacerbating role of the NLRP3-ASC axis on exogenously and non-exogenously seeded Tau pathology in Tau mice in vivo. The NLRP3-ASC inflammasome, which is an important sensor of innate immunity and intensively explored for its role in health and disease, hence presents as an interesting therapeutic approach to target three crucial pathogenetic processes in AD, including prion-like seeding of Tau pathology, Aß pathology and neuroinflammation.

Absence of early metabolic response assessed by 18F-FDG PET/CT after initiation of antifibrotic drugs in IPF patients.

BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is characterized by a progressive and irreversible respiratory failure. Non-invasive markers of disease activity are essential for prognosis and evaluation of early response to anti-fibrotic treatments. OBJECTIVES: The aims of this study were to determine whether fluorodeoxyglucose ([18F]-FDG) lung uptake is reduced after initiation of pirfenidone or nintedanib and to assess its possible use as a prognostic factor. METHODS: [18F]-FDG PET/CT was performed in IPF patients and in a murine model of pulmonary fibrosis. PET/CTs were performed at day 8 and day 15 post-instillation of bleomycin in pirfenidone- or vehicule-treated mice. In IPF patients, PET-CT was performed before and 3 months after the initiation of pirfenidone or nintedanib. RESULTS: In bleomycin-treated mice, pirfenidone significantly reduced the [18F]-FDG uptake compared to vehicule-treated mice at day 15 (p < 0.001), whereas no difference was observed at day 8 after bleomycin administration. In IPF patients, [18F]-FDG lung uptake before and after 3 months of treatment by nintedanib (n = 11) or pirfenidone (n = 14) showed no significant difference regardless the antifibrotic treatment. Moreover, no difference was noticed between patients with progressive or non-progressive disease at one year of follow up. CONCLUSIONS: Pirfenidone significantly reduces the lung [18F]-FDG uptake during the fibrotic phase in a mouse model of IPF. However, these preclinical data were not confirmed in IPF patients 3 months after the initiation of antifibrotic therapy. [18F]-FDG seems therefore not useful in clinical practice to assess the early response of IPF patients to nintedanib or pirfenidone.