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.

mTOR Activation Underlies Enhanced B Cell Proliferation and Autoimmunity in PrkcdG510S/G510S Mice.

Autosomal recessive PRKCD deficiency has previously been associated with the development of systemic lupus erythematosus in human patients, but the mechanisms underlying autoimmunity remain poorly understood. We introduced the Prkcd G510S mutation that we previously associated to a Mendelian cause of systemic lupus erythematosus in the mouse genome, using CRISPR-Cas9 gene editing. PrkcdG510S/G510S mice recapitulated the human phenotype and had reduced lifespan. We demonstrate that this phenotype is linked to a B cell-autonomous role of Prkcd. A detailed analysis of B cell activation in PrkcdG510S/G510S mice shows an upregulation of the PI3K/mTOR pathway after the engagement of the BCR in these cells, leading to lymphoproliferation. Treatment of mice with rapamycin, an mTORC1 inhibitor, significantly improves autoimmune symptoms, demonstrating in vivo the deleterious effect of mTOR pathway activation in PrkcdG510S/G510S mice. Additional defects in PrkcdG510S/G510S mice include a decrease in peripheral mature NK cells that might contribute to the known susceptibility to viral infections of patients with PRKCD mutations.

Combinatorial Expression of NK Cell Receptors Governs Cell Subset Reactivity and Effector Functions but Not Tumor Specificity.

NK cell receptors allow NK cells to recognize targets such as tumor cells. Many of them are expressed on a subset of NK cells, independently of each other, which creates a vast diversity of receptor combinations. Whether these combinations influence NK cell antitumor responses is not well understood. We addressed this question in the C57BL/6 mouse model and analyzed the individual effector response of 444 mouse NK cell subsets, defined by combinations of 12 receptors, against tumor cell lines originating from different tissues and mouse strains. We found a wide range of reactivity among NK subsets, but the same hierarchy of responses was observed for the different tumor types, showing that the repertoire of NK cell receptors does not encode for different tumor specificities but for different intrinsic reactivities. The coexpression of CD27, NKG2A, and DNAM-1 identified subsets with relative cytotoxic specialization, whereas reciprocally, CD11b and KLRG1 defined the best IFN-γ producers. The expression of educating receptors Ly49C, Ly49I, and NKG2A was also strongly correlated with IFN-γ production, but this effect was suppressed by unengaged receptors Ly49A, Ly49F, and Ly49G2. Finally, IL-15 coordinated NK cell effector functions, but education and unbound inhibitory receptors retained some influence on their response. Collectively, these data refine our understanding of the mechanisms governing NK cell reactivity, which could help design new NK cell therapy protocols.

Sequential actions of EOMES and T-BET promote stepwise maturation of natural killer cells.

EOMES and T-BET are related T-box transcription factors that control natural killer (NK) cell development. Here we demonstrate that EOMES and T-BET regulate largely distinct gene sets during this process. EOMES is dominantly expressed in immature NK cells and drives early lineage specification by inducing hallmark receptors and functions. By contrast, T-BET is dominant in mature NK cells, where it induces responsiveness to IL-12 and represses the cell cycle, likely through transcriptional repressors. Regardless, many genes with distinct functions are co-regulated by the two transcription factors. By generating two gene-modified mice facilitating chromatin immunoprecipitation of endogenous EOMES and T-BET, we show a strong overlap in their DNA binding targets, as well as extensive epigenetic changes during NK cell differentiation. Our data thus suggest that EOMES and T-BET may distinctly govern, via differential expression and co-factors recruitment, NK cell maturation by inserting partially overlapping epigenetic regulations.