Macrophages are considered to contribute to chronic inflammatory diseases such as rheumatoid arthritis
. However, both the exact origin and the role of macrophages in inflammatory joint disease ...remain unclear. Here we use fate-mapping approaches in conjunction with three-dimensional light-sheet fluorescence microscopy and single-cell RNA sequencing to perform a comprehensive spatiotemporal analysis of the composition, origin and differentiation of subsets of macrophages within healthy and inflamed joints, and study the roles of these macrophages during arthritis. We find that dynamic membrane-like structures, consisting of a distinct population of CX
CR1
tissue-resident macrophages, form an internal immunological barrier at the synovial lining and physically seclude the joint. These barrier-forming macrophages display features that are otherwise typical of epithelial cells, and maintain their numbers through a pool of locally proliferating CX
CR1
mononuclear cells that are embedded into the synovial tissue. Unlike recruited monocyte-derived macrophages, which actively contribute to joint inflammation, these epithelial-like CX
CR1
lining macrophages restrict the inflammatory reaction by providing a tight-junction-mediated shield for intra-articular structures. Our data reveal an unexpected functional diversification among synovial macrophages and have important implications for the general role of macrophages in health and disease.
12/15-Lipoxygenase (12/15-LOX) mediates the enzymatic oxidation of polyunsaturated fatty acids, thereby contributing to the generation of various bioactive lipid mediators. Although 12/15-LOX has ...been implicated in the pathogenesis of multiple chronic inflammatory diseases, its physiologic functions seem to include potent immune modulatory properties that physiologically contribute to the resolution of inflammation and the clearance of inflammation-associated tissue damage. This review aims to give a comprehensive overview about our current knowledge on the role of this enzyme during the regulation of inflammation and immunity. This article is part of a Special Issue entitled: Lipid modification and lipid peroxidation products in innate immunity and inflammation edited by Christoph J. Binder.
•In mammals 12/15-LOX is expressed either in steady state or after induction in a variety of cell types.•12/15-LOX generates lipid mediators with immune modulatory properties by enzymatic oxidation of polyunsaturated fatty acids.•Controlled and timed 12/15-LOX expression can facilitate the resolution of inflammation.•Its deregulated activity however, contributes to tissue damage, cell death and chronic inflammation.•As an important regulator of inflammation, 12/15-LOX might be a relevant future target for clinical purposes.
Blood coagulation is essential for physiological hemostasis but simultaneously contributes to thrombotic disease. However, molecular and cellular events controlling initiation and propagation of ...coagulation are still incompletely understood. In this study, we demonstrate an unexpected role of eosinophils during plasmatic coagulation, hemostasis, and thrombosis. Using a large-scale epidemiological approach, we identified eosinophil cationic protein as an independent and predictive risk factor for thrombotic events in humans. Concurrent experiments showed that eosinophils contributed to intravascular thrombosis by exhibiting a strong endogenous thrombin-generation capacity that relied on the enzymatic generation and active provision of a procoagulant phospholipid surface enriched in 12/15-lipoxygenase-derived hydroxyeicosatetraenoic acid-phosphatidylethanolamines. Our findings reveal a previously unrecognized role of eosinophils and enzymatic lipid oxidation as regulatory elements that facilitate both hemostasis and thrombosis in response to vascular injury, thus identifying promising new targets for the treatment of thrombotic disease.
The checkpoints and mechanisms that contribute to autoantibody-driven disease are as yet incompletely understood. Here we identified the axis of interleukin 23 (IL-23) and the T
17 subset of helper T ...cells as a decisive factor that controlled the intrinsic inflammatory activity of autoantibodies and triggered the clinical onset of autoimmune arthritis. By instructing B cells in an IL-22- and IL-21-dependent manner, T
17 cells regulated the expression of β-galactoside α2,6-sialyltransferase 1 in newly differentiating antibody-producing cells and determined the glycosylation profile and activity of immunoglobulin G (IgG) produced by the plasma cells that subsequently emerged. Asymptomatic humans with rheumatoid arthritis (RA)-specific autoantibodies showed identical changes in the activity and glycosylation of autoreactive IgG antibodies before shifting to the inflammatory phase of RA; thus, our results identify an IL-23-T
17 cell-dependent pathway that controls autoantibody activity and unmasks a preexisting breach in immunotolerance.
Eosinophils were reported to serve as an essential component of the plasma cell niche within the bone marrow. As the potential contribution of eosinophils to humoral immunity has remained ...incompletely understood, we aimed to further characterize their role during antibody responses and to additionally investigate their role in autoimmune disease. Contrary to our expectations and the currently prevailing paradigm, we found that eosinophils are fully dispensable for the survival of murine bone marrow plasma cells and accordingly do not contribute to antibody production and autoantibody‐mediated disease. Littermate wild type and eosinophil‐deficient ΔdblGATA‐1 animals showed similar numbers and frequencies of plasma cells and did not differ in steady state levels of immunoglobulins or their ability to raise antigen‐specific antibody responses. Eosinophils were likewise dispensable for autoantibody production or autoantibody‐induced disease in a mouse model of systemic lupus erythematosus. Our findings thus argue against a role of eosinophils during the maintenance of the plasma cell pool and challenge the hitherto postulated concept of an eosinophil‐sustained bone marrow niche.
Eosinophils are nonessential for the production of plasma cell survival factors and for the maintenance of the plasma cell survival niche. Other cell types produce these survival factors and compensate for a lack of eosinophils in this context by enabling the survival of long lived plasma cells in the bone marrow in the absence of eosinophils.
Structural remodeling or damage as a result of disease or injury is often not evenly distributed throughout a tissue but strongly depends on localization and extent of damaging stimuli. Skeletal ...muscle as a mechanically active organ can express signs of local or even systemic myopathic damage, necrosis, or repair. Conventionally, muscle biopsies (patients) or whole muscles (animal models) are mechanically sliced and stained to assess structural alterations histologically. Three-dimensional tissue information can be obtained by applying deep imaging modalities,
multiphoton or light-sheet microscopy. Chemical clearing approaches reduce scattering,
through matching refractive tissue indices, to overcome optical penetration depth limits in thick tissues.
Here, we optimized a range of different clearing protocols. We find aqueous solution-based protocols employing (20-80%) 2,2'-thiodiethanol (TDE) to be advantageous over organic solvents (dibenzyl ether, cinnamate) regarding the preservation of muscle morphology, ease-of-use, hazard level, and costs.
Applying TDE clearing to a mouse model of local cardiotoxin (CTX)-induced muscle necrosis, a complete loss of myosin-II signals was observed in necrotic areas with little change in fibrous collagen or autofluorescence (AF) signals. The 3D aspect of myofiber integrity could be assessed, and muscle necrosis in whole muscle was quantified locally
the ratios of detected AF, forward- and backward-scattered Second Harmonic Generation (fSHG, bSHG) signals.
TDE optical clearing is a versatile tool to study muscle architecture in conjunction with label-free multiphoton imaging in 3D in injury/myopathy models and might also be useful in studying larger biofabricated constructs in regenerative medicine.
Signals from the BCR are required for Ag-specific B cell recruitment into the immune response. Binding of Ag to the BCR induces phosphorylation of immune receptor tyrosine-based activation motifs in ...the cytoplasmic domains of the CD79a and CD79b signaling subunits, which subsequently bind and activate the Syk protein tyrosine kinase. Earlier work with the DT40 chicken B cell leukemia cell line showed that Syk was required to transduce BCR signals to proximal activation events, suggesting that Syk also plays an important role in the activation and differentiation of primary B cells during an immune response. In this study, we show that Syk-deficient primary mouse B cells have a severe defect in BCR-induced activation, proliferation, and survival. Furthermore, we demonstrate that Syk is required for both T-dependent and T-independent Ab responses, and that this requirement is B cell intrinsic. In the absence of Syk, Ag fails to induce differentiation of naive B cells into germinal center B cells and plasma cells. Finally, we show that the survival of existing memory B cells is dependent on Syk. These experiments demonstrate that Syk plays a critical role in multiple aspects of B cell Ab responses.
Oxidation of low-density lipoprotein (LDL) and oxidized LDL-mediated activation of the innate immune system have been recognized as early key events during the pathogenesis of atherosclerosis. Recent ...evidence identified eosinophils as a major source of enzymatic lipid oxidation and suggested a potential role of type 2 immunity in atherogenesis. However, the involvement of individual type 2 immune cell subsets involved in this process has been incompletely defined. We therefore sought to determine the role of eosinophils during LDL oxidation and the pathogenesis of this disease.
Using eosinophil-deficient dblGATA1 mice, we studied the role of eosinophils in two established mouse models of atherosclerosis.
These experiments revealed that the presence of eosinophils did neither affect biomarkers of LDL oxidation nor atherosclerotic lesion development.
The obtained results show that LDL oxidation and development of atherosclerosis are largely independent of eosinophils or eosinophil-mediated LDL oxidation.
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•Eosinophils do not alter oxidation of low-density lipoprotein (LDL) in vivo.•Absence of eosinophils does not influence development of atherosclerotic burden in two different mouse models.•Other sources than eosinophils contribute to enzymatic oxidation of LDL in vivo.
Grb2 is a ubiquitously expressed adaptor protein, which activates Ras and MAP kinases in growth factor receptor signalling, while in B‐cell receptor (BCR) signalling this role is controversial. In B ...cell lines it was shown that Grb2 can inhibit BCR‐induced Ca2+ signalling. Nonetheless, the physiological role of Grb2 in primary B cells is still unknown. We generated a B‐cell‐specific Grb2‐deficient mouse line, which had a severe reduction of mature follicular B cells in the periphery due to a differentiation block and decreased B‐cell survival. Moreover, we found several changes in important signalling pathways: enhanced BCR‐induced Ca2+ signalling, alterations in mitogen‐activated protein kinase activation patterns and strongly impaired Akt activation, the latter pointing towards a defect in PI3K signalling. Interestingly, B‐cell‐specific Grb2‐deficient mice showed impaired IgG and B‐cell memory responses, and impaired germinal centre formation. Thus, Grb2‐dependent signalling pathways are crucial for lymphocyte differentiation processes, as well as for control of secondary humoral immune responses.
Grb2 regulates B‐cell maturation, B‐cell memory responses and inhibits B‐cell Ca2+ signalling. The adaptor protein Grb2 regulates different signalling pathways, but its role in primary B cells is not known. Using B‐cell‐specific Grb2‐deficient mice, the findings show that Grb2 is important for the development of mature B cells and memory B‐cell function.
Platelets are anuclear cell fragments derived from bone marrow megakaryocytes (MKs) that safeguard vascular integrity but may also cause pathological vessel occlusion. One major pathway of platelet ...activation is triggered by 2 receptors that signal through an (hem)immunoreceptor tyrosine-based activation motif (ITAM), the activating collagen receptor glycoprotein (GP) VI and the C-type lectin-like receptor 2 (CLEC-2). Growth factor receptor-bound protein 2 (Grb2) is a ubiquitously expressed adapter molecule involved in signaling processes of numerous receptors in different cell types, but its function in platelets and MKs is unknown.
We tested the hypothesis that Grb2 is a crucial adapter protein in (hem)immunoreceptor tyrosine-based activation motif signaling in platelets.
Here, we show that genetic ablation of Grb2 in MKs and platelets did not interfere with MK differentiation or platelet production. However, Grb2-deficiency severely impaired glycoprotein VI-mediated platelet activation because of defective stabilization of the linker of activated T-cell (LAT) signalosome and activation of downstream signaling proteins that resulted in reduced adhesion, aggregation, and coagulant activity on collagen in vitro. Similarly, CLEC-2-mediated signaling was impaired in Grb2-deficient platelets, whereas the cells responded normally to stimulation of G protein-coupled receptors. In vivo, this selective (hem)immunoreceptor tyrosine-based activation motif signaling defect resulted in prolonged bleeding times but affected arterial thrombus formation only after concomitant treatment with acetylsalicylic acid, indicating that defective glycoprotein VI signaling in the absence of Grb2 can be compensated through thromboxane A2-induced G protein-coupled receptor signaling pathways.
These results reveal an important contribution of Grb2 in (hem)immunoreceptor tyrosine-based activation motif signaling in platelets in hemostasis and thrombosis by stabilizing the LAT signalosome.
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