Immune complex-induced haptokinesis in human non-classical monocytes.

Formation and deposition of immune complexes (ICs) are hallmarks of various autoimmune diseases. Detection of ICs by IC receptors on leukocytes induces downstream signaling and shapes the local immune response. In many cases the pathological relevance of ICs is not well understood. We here show that ICs induce a distinct migratory response, i.e. haptokinesis in 6-sulfo LacNAc+ monocytes (slanMo) and in non-classical monocytes (ncMo) but not in intermediate (imMo) and classical monocytes (cMo). Using live imaging combined with automated cell tracking, we show that the main features of IC-dependent haptokinesis are elongation of the cell body, actin polarization at the leading edge, and highly directional migration. We find that CD16-dependent signaling mediates haptokinesis as blocking of CD16 or blocking SYK-signaling inhibited the migratory response. The activity of the metalloproteinase ADAM17 also modifies IC-dependent haptokinesis, likely at least partially via cleavage of CD16. Furthermore, using matrices with defined ligand spacing, we show that ligand density impacts the magnitude of the migratory response. Taken together, we have demonstrated that ICs induce a specific migratory response in ncMo but not in other monocyte subsets. Therefore, our work lays the groundwork for the investigation of IC-dependent haptokinesis in ncMo as a potential pathomechanism in IC-mediated autoimmune diseases.

IFNγ Causes Keratinocyte Necroptosis in Acute Graft-Versus-Host Disease.

Epidermal keratinocytes form the first-line cellular barrier of the skin for protection against external injuries and maintenance of local tissue homeostasis. Expression of ZBP1 was shown to cause necroptotic keratinocyte cell death and skin inflammation in mice. We sought to characterize the relevance of ZBP1 and necroptosis in human keratinocytes and type 1-driven cutaneous acute graft-versus-host disease. in this study, we identify ZBP1 expression, necroptosis, and interface dermatitis as being the hallmarks of acute graft-versus-host disease. ZBP1 expression was dependent on leukocyte-derived IFNγ, and interference with IFNγ signaling by Jak inhibition prevented cell death. In predominantly IL-17-driven psoriasis, both ZBP1 expression and necroptosis could not be detected. Of note, in contrast to the signaling in mice, ZBP1 signaling in human keratinocytes was not affected by RIPK1's presence. These findings show that ZBP1 drives inflammation in IFNγ-dominant type 1 immune responses in human skin and may further point to a general role of ZBP1-mediated necroptosis.

Current Concepts on 6-sulfo LacNAc Expressing Monocytes (slanMo).

The human mononuclear phagocytes system consists of dendritic cells (DCs), monocytes, and macrophages having different functions in bridging innate and adaptive immunity. Among the heterogeneous population of monocytes the cell surface marker slan (6-sulfo LacNAc) identifies a specific subset of human CD14- CD16+ non-classical monocytes, called slan+ monocytes (slanMo). In this review we discuss the identity and functions of slanMo, their contributions to immune surveillance by pro-inflammatory cytokine production, and cross talk with T cells and NK cells. We also consider the role of slanMo in the regulation of chronic inflammatory diseases and cancer. Finally, we highlight unresolved questions that should be the focus of future research.

Phenotype, Function, and Mobilization of 6-Sulfo LacNAc-Expressing Monocytes in Atopic Dermatitis.

Mononuclear phagocytes (MPs) are important immune regulatory cells in atopic dermatitis (AD). We previously identified 6-sulfo LacNAc-expressing monocytes (slanMo) as TNF-α- and IL-23-producing cells in psoriatic skin lesions and as inducers of IFN-γ-, IL-17-, and IL-22-producing T cells. These cytokines are also upregulated in AD and normalize with treatment, as recently shown for dupilumab-treated patients. We here asked for the role of slanMo in AD. Increased numbers of slanMo were found in AD skin lesions. In difference to other MPs in AD, slanMo lacked expression of FcɛRI, CD1a, CD14, and CD163. slanMo from blood of patients with AD expressed increased levels of CD86 and produced IL-12 and TNF-α at higher amounts than CD14+ monocytes and myeloid dendritic cells. While CD14+ monocytes from patients with AD revealed a reduced IL-12 production, we observed no difference in the cytokine production comparing slanMo in AD and healthy controls. Interestingly, experimentally induced mental stress, a common trigger of flares in patients with AD, rapidly mobilized slanMo which retained their high TNF-α-producing capacity. This study identifies slanMo as a distinct population of inflammatory cells in skin lesions and as proinflammatory blood cells in patients with AD. slanMo may, therefore, represent a potent future target for treatment of AD.

Intracapillary immune complexes recruit and activate slan-expressing CD16+ monocytes in human lupus nephritis.

Lupus nephritis is a major cause of morbidity in patients with systemic lupus erythematosus. Among the different types of lupus nephritis, intracapillary immune complex (IC) deposition and accumulation of monocytes are hallmarks of lupus nephritis class III and IV. The relevance of intracapillary ICs in terms of monocyte recruitment and activation, as well as the nature and function of these monocytes are not well understood. For the early focal form of lupus nephritis (class III) we demonstrate a selective accumulation of the proinflammatory population of 6-sulfo LacNAc+ (slan) monocytes (slanMo), which locally expressed TNF-α. Immobilized ICs induced a direct recruitment of slanMo from the microcirculation via interaction with Fc γ receptor IIIA (CD16). Interestingly, intravenous immunoglobulins blocked CD16 and prevented cell recruitment. Engagement of immobilized ICs by slanMo induced the production of neutrophil-attracting chemokine CXCL2 as well as TNF-α, which in a forward feedback loop stimulated endothelial cells to produce the slanMo-recruiting chemokine CX3CL1 (fractalkine). In conclusion, we observed that expression of CD16 equips slanMo with a unique capacity to orchestrate early IC-induced inflammatory responses in glomeruli and identified slanMo as a pathogenic proinflammatory cell type in lupus nephritis.

The phosphodiesterase 4 inhibitor apremilast inhibits Th1 but promotes Th17 responses induced by 6-sulfo LacNAc (slan) dendritic cells.

BACKGROUND: The phosphodiesterase 4 (PDE4) inhibitor apremilast increases cellular cAMP levels and has proven effective in the treatment of psoriasis and psoriasis arthritis. We recently described 6-sulfo LacNAc dendritic cells (slanDCs) as immature DCs in blood and as a subset of inflammatory dermal DCs in psoriasis with a pronounced capacity to produce proinflammatory cytokines and to program Th17/Th1 T cell responses. OBJECTIVE: The aim of this study was to investigate possible immune regulatory effects of the PDE4 inhibitor apremilast on slanDCs. METHODS: In vitro studies were performed analyzing the effects of apremilast on the proinflammatory function of slanDCs and their capacity to induce Th1/Th17-biased T cell responses. RESULTS: Increasing cAMP levels in slanDCs by PDE4 inhibition strongly reduced production of IL-12 and TNF-α. In line with these findings, co-culture experiments with apremilast-pulsed slanDCs and allogeneic T cells either from psoriasis patients or healthy controls, revealed a significant reduction of IFN-γ production and expression of the transcription factor T-bet. In parallel, production of IL-23 and IL-1ß by slanDCs was increased and co-cultured T cells revealed a largely augmented IL-17 production and an upregulated RORyt expression. CONCLUSIONS: We here demonstrate anti-inflammatory as well as Th17-promoting effects of apremilast when studying blood precursors of human inflammatory dermal dendritic cells. In the concert of the broad anti-inflammatory effects of apremilast on keratinocytes, fibroblasts and endothelial cells, the dual effect on slan+ inflammatory dermal DCs should be taken into account and may constrain therapeutic responses.

Inter-SINE Amplified Polymorphism (ISAP) for rapid and robust plant genotyping.

The unambiguous differentiation of crop genotypes is often laborious or expensive. A rapid, robust, and cost-efficient marker system is required for routine genotyping in plant breeding and marker-assisted selection. We describe the Inter-SINE Amplified Polymorphism (ISAP) system that is based on standard molecular methods resulting in genotype-specific fingerprints at high resolution. These markers are derived from Short Interspersed Nuclear Elements (SINEs) which are dispersed repetitive sequences present in most if not all plant genomes and can be efficiently extracted from plant genome sequences. The ISAP method was developed on potato as model plant but is also transferable to other plant species.

[The role of human 6-sulfo LacNAc dendritic cells (slanDCs) in autoimmunity and tumor diseases]. / Die Bedeutung von humanen 6-sulfo LacNAc dendritischen Zellen (slanDCs) bei Autoimmunität und Tumorerkrankungen.

Dendritic cells play a central role in the regulation of immunological reactivity. The existence of functionally specialized populations of skin dendritic cells is a consequence of qualitatively different attacks on our organism. slanDCs are human inflammatory dendritic cells that are characterized by the specific expression of the carbohydrate 6-sulfo LacNAc (slan). After phenotypic maturation slanDCs are capable of producing very high amounts of proinflammatory mediators like IL-12, TNF-α, IL-1ß â and IL-23. Recent data describe a potential role of slanDCs in a number of different diseases like psoriasis, lupus erythematosus but also tumor diseases and therefore open up new areas of research on their respective pathogenesis. Furthermore, as a basis of a directed therapeutic manipulation,a slanDC-specific targeting system has been developed. Future challenges of slanDC research include the elaboration of a deeper understanding of the significance of slanDCs for the regulation of adaptive and innate immune responses as well as a translation of this knowledge into therapeutic options.

Future treatment options for atopic dermatitis - small molecules and beyond.

Atopic dermatitis (AD) is a common eczematous skin disease with a chronic and relapsing course. Current therapeutic options for moderate to severe AD in children and adults are unsatisfactory. Along with the success of basic research to define pathogenesis-related targets, novel small molecule inhibitors and biologics for the treatment of AD have been developed. These compounds focus on the specific reduction of pruritus, interfere with the pro-allergic Th2-deviation of the immune system or inhibit inflammatory pathways in the skin. Based on studies registered at ClinicalTrials.gov we present novel treatment strategies of AD, their molecular mechanisms of action, and discuss the current status of the clinical results. As many of the new compounds target pathogenesis-related traits of the disease, we face a new era in the treatment and understanding of AD.

FcγRIII (CD16) equips immature 6-sulfo LacNAc-expressing dendritic cells (slanDCs) with a unique capacity to handle IgG-complexed antigens.

Binding and uptake of immune complexes (ICs) via low-affinity Fcγ receptors (FcγRs) on dendritic cells (DCs) is well known as a booster of immune responses. It can be helpful when stimulating immunity against pathogenic microbes but may be harmful when antibodies form complexes with autologous antigens. To date, no human DC subtype specialized in handling ICs has been identified. By incubating human blood mononuclear cells with ICs and studying their cellular binding, we identified 6-sulfo LacNAc-expressing DCs (slanDCs) as having an outstanding capacity to bind ICs compared with other myeloid DCs, plasmacytoid DCs, or monocytes. Using selective blocking of different (FcγRs), we identified CD16 (FcγRIII) as the major IC-binding structure on slanDCs. In addition, CD16 proved critical for phagocytosis of IgG-coated erythrocytes, and CD16-targeted antigen led to a more efficient proliferation of CD4(+) T cells than CD32 (FcγRII)-targeted antigen. Interestingly, these CD16-mediated functions are short-lived and restricted to the immature stage of slanDCs in blood. We show that CD16 is rapidly shed from the surface of maturing slanDCs, resulting from the combined action of the metalloproteinases ADAM10 and ADAM17. In conclusion, these data provide strong evidence that slanDCs play an important role in IC-driven immune responses.

Human 6-sulfo LacNAc (slan) dendritic cells have molecular and functional features of an important pro-inflammatory cell type in lupus erythematosus.

Lupus erythematosus (LE) is an autoimmune disease with evidence for an IL-23- and IL-17-induced immunopathology. Little is known about the type of dendritic cells supporting this immune response. We recently demonstrated the strong Th1- and Th17-T-cell inducing capacity of human 6-sulfo LacNAc-dendritic cells (slanDCs), and identified slanDCs as inflammatory dermal dendritic cells in psoriasis locally expressing IL-23, TNF-α and inducible nitric oxide synthase (iNOS). In this study, we investigated the role of slanDCs in LE. Using immunohistochemistry, we identified slanDCs at increased frequency in affected skin lesions of cutaneous and systemic LE. slanDCs were found scattered in the dermal compartment and also clustered in lymph follicle-like structures. Here, they colocalized with T cells in the periphery but not with B cells in the center. The positive staining of dermal slanDCs for TNF-α indicated their pro-inflammatory status. In vitro the production of TNF-α was induced when slanDCs were cultured in the presence of serum from patients with LE. Stimulatory components of LE serum were previously identified as autoimmune complexes with ssRNA binding to TLR7 and TLR8. We found that slanDCs express mRNA for TLR7 and TLR8. slanDCs stimulated with ssRNA, selective TLR7 or TLR8 ligands responded with high-level TNF-α and IL-12 production. In contrast to slanDCs, the population of CD1c(+) DCs and plasmacytoid DCs (pDCs) expressed either TLR7 or TLR8, and their production of TNF-α and IL-12 to respective ligands was far less pronounced. We conclude that slanDCs have molecular and functional features of a pro-inflammatory myeloid DC type relevant for the immunopathogenesis of LE.

Targeted identification of short interspersed nuclear element families shows their widespread existence and extreme heterogeneity in plant genomes.

Short interspersed nuclear elements (SINEs) are non-long terminal repeat retrotransposons that are highly abundant, heterogeneous, and mostly not annotated in eukaryotic genomes. We developed a tool designated SINE-Finder for the targeted discovery of tRNA-derived SINEs. We analyzed sequence data of 16 plant genomes, including 13 angiosperms and three gymnosperms and identified 17,829 full-length and truncated SINEs falling into 31 families showing the widespread occurrence of SINEs in higher plants. The investigation focused on potato (Solanum tuberosum), resulting in the detection of seven different SolS SINE families consisting of 1489 full-length and 870 5' truncated copies. Consensus sequences of full-length members range in size from 106 to 244 bp depending on the SINE family. SolS SINEs populated related species and evolved separately, which led to some distinct subfamilies. Solanaceae SINEs are dispersed along chromosomes and distributed without clustering but with preferred integration into short A-rich motifs. They emerged more than 23 million years ago and were species specifically amplified during the radiation of potato, tomato (Solanum lycopersicum), and tobacco (Nicotiana tabacum). We show that tobacco TS retrotransposons are composite SINEs consisting of the 3' end of a long interspersed nuclear element integrated downstream of a nonhomologous SINE family followed by successfully colonization of the genome. We propose an evolutionary scenario for the formation of TS as a spontaneous event, which could be typical for the emergence of SINE families.