Skap2 is required for β2 integrin-mediated neutrophil recruitment and functions.

Integrin activation is required for neutrophil functions. Impaired integrin activation on neutrophils is the hallmark of leukocyte adhesion deficiency (LAD) syndrome in humans, characterized by impaired leukocyte recruitment and recurrent infections. The Src kinase-associated phosphoprotein 2 (Skap2) is involved in integrin functions in different leukocyte subtypes. However, the role of Skap2 in β2 integrin activation and neutrophil recruitment is unknown. In this study, we demonstrate the crucial role of Skap2 in regulating actin polymerization and binding of talin-1 and kindlin-3 to the β2 integrin cytoplasmic domain, thereby being indispensable for β2 integrin activation and neutrophil recruitment. The direct interaction of Skap2 with the Wiskott-Aldrich syndrome protein via its SH3 domain is critical for integrin activation and neutrophil recruitment in vivo. Furthermore, Skap2 regulates integrin-mediated outside-in signaling events and neutrophil functions. Thus, Skap2 is essential to activate the β2 integrins, and loss of Skap2 function is sufficient to cause a LAD-like phenotype in mice

Filamin A Phosphorylation at Serine 2152 by the Serine/Threonine Kinase Ndr2 Controls TCR-Induced LFA-1 Activation in T Cells

The integrin LFA-1 (CD11a/CD18) plays a critical role in the interaction of T cells with antigen presenting cells (APCs) to promote lymphocyte differentiation and proliferation. This integrin can be present either in a closed or in an open active conformation and its activation upon T-cell receptor (TCR) stimulation is a critical step to allow interaction with APCs. In this study we demonstrate that the serine/threonine kinase Ndr2 is critically involved in the initiation of TCR-mediated LFA-1 activation (open conformation) in T cells. Ndr2 itself becomes activated upon TCR stimulation and phosphorylates the intracellular integrin binding partner Filamin A (FLNa) at serine 2152. This phosphorylation promotes the dissociation of FLNa from LFA-1, allowing for a subsequent association of Talin and Kindlin-3 which both stabilize the open conformation of LFA-1. Our data suggest that Ndr2 activation is a crucial step to initiate TCR-mediated LFA-1 activation in T cells

The adapter protein ADAP is required for selected dendritic cell functions

<p>Abstract</p> <p>Background</p> <p>The cytosolic adaptor protein ADAP (adhesion and degranulation promoting adapter protein) is expressed by T cells, natural killer cells, myeloid cells and platelets. ADAP is involved in T-cell-receptor-mediated inside-out signaling, which leads to integrin activation, adhesion and reorganization of the actin cytoskeleton. However, little is known about the role of ADAP in myeloid cells. In the present study, we analyzed the function of ADAP in bone-marrow-derived dendritic cells (BMDCs) from ADAP-deficient mice.</p> <p>Results</p> <p>ADAP-deficient BMDCs showed almost normal levels of antigen uptake, adhesion, maturation, migration from the periphery to the draining lymph nodes, antigen-specific T-cell activation, and production of the proinflammatory cytokines IL-6 and TNF-∝. Furthermore, we provide evidence that the activation of signaling pathways after lipopolysaccharide (LPS) stimulation are not affected by the loss of ADAP. In contrast, ADAP-deficient BMDCs showed defects in CD11c-mediated cellular responses, with significantly diminished production of IL-6, TNF-∝ and IL-10. Actin polymerization was enhanced after CD11c integrin stimulation.</p> <p>Conclusions</p> <p>In summary, we propose that the adapter molecule ADAP is critical for selected CD11c integrin-mediated functions of dendritic cells.</p

Homologous and Heterologous Anti-COVID-19 Vaccination Does Not Induce New-Onset Formation of Autoantibodies Typically Accompanying Lupus Erythematodes, Rheumatoid Arthritis, Celiac Disease and Antiphospholipid Syndrome

The COVID-19 pandemics has caused the death of almost six million people worldwide. In order to establish collective immunity, the first vaccines that were approved in Germany were the vector virus-based vaccine Vaxzevria and the mRNA vaccines Comirnaty and Spikevax, respectively. As it was reported that SARS-CoV-2 can trigger autoimmunity, it is of significant interest to investigate whether COVID-19 vaccines evoke the formation of autoantibodies and subsequent autoimmunity. Here, we analyzed immune responses after different vaccination regimens (mRNA/mRNA, Vector/Vector or Vector/mRNA) with respect to anti-SARS-CoV-2-specific immunity and the development of autoantibodies well known for their appearance in distinct autoimmune diseases. We found that anti-SARS-CoV-2 antibody levels were 90% lower after Vector/Vector vaccination compared to the other vaccinations and that Vector/mRNA vaccination was more effective than mRNA/mRNA vaccination in terms of IgM and IgA responses. However, until 4 months after booster vaccination we only detected increases in autoantibodies in participants with already pre-existing autoantibodies whereas vaccinees showing no autoantibody formation before vaccination did not respond with sustained autoantibody production. Taken together, our study suggests that all used COVID-19 vaccines do not significantly foster the appearance of autoantibodies commonly associated with lupus erythematodes, rheumatoid arthritis, Celiac disease and antiphospholipid-syndrome but provide immunity to SARS-CoV-2

Pitavastatin Is a Highly Potent Inhibitor of T-Cell Proliferation

Repositioning of approved drugs is an alternative time- and cost-saving strategy to classical drug development. Statins are 3-hydroxy-3-methylglutaryl-CoA (HMG CoA) reductase inhibitors that are usually used as cholesterol-lowering medication, and they also exhibit anti-inflammatory effects. In the present study, we observed that the addition of Pitavastatin at nanomolar concentrations inhibits the proliferation of CD3/CD28 antibody-stimulated human T cells of healthy donors in a dose-dependent fashion. The 50% inhibition of proliferation (IC50) were 3.6 and 48.5 nM for freshly stimulated and pre-activated T cells, respectively. In addition, Pitavastatin suppressed the IL-10 and IL-17 production of stimulated T cells. Mechanistically, we found that treatment of T cells with doses &lt;1 µM of Pitavastatin induced hyperphosphorylation of ERK1/2, and activation of caspase-9, -3 and -7, thus leading to apoptosis. Mevalonic acid, cholesterol and the MEK1/2 inhibitor U0126 reversed this Pitavastatin-mediated ERK1/2 activation and apoptosis of T cells. In summary, our results suggest that Pitavastatin is a highly potent inhibitor of T-cell proliferation, which induces apoptosis via pro-apoptotic ERK1/2 activation, thus representing a potential repositioning candidate for the treatment of T-cell-mediated autoimmune diseases

Characterization of mice with a platelet-specific deletion of the adapter molecule ADAP.

The adhesion and degranulation-promoting adapter protein (ADAP) is expressed in T cells, NK cells, myeloid cells, and platelets. The involvement of ADAP in the regulation of receptor-mediated inside-out signaling leading to integrin activation is well characterized, especially in T cells and in platelets. Due to the fact that animal studies using conventional knock-out mice are limited by the overlapping effects of the different ADAP-expressing cells, we generated conditional ADAP knock-out mice (ADAPfl/fl PF4-Cretg). We observed that loss of ADAP restricted to the megakaryocytic lineage has no impact on other hematopoietic cells even after stimulation conditions. ADAPfl/fl PF4-Cretg mice showed thrombocytopenia in combination with reduced plasma levels of PF4 and TGF-β1. In vitro, platelets from these mice revealed reduced P-selectin expression, lower TGF-β1 release, diminished integrin αIIbβ3 activation and decreased fibrinogen binding after stimulation with podoplanin, the ligand of the C-type lectin-like receptor-2 (CLEC-2). Furthermore, loss of ADAP was associated with impaired CLEC-2-mediated activation of PLCγ2 and Erk1/2. Induction of experimental autoimmune encephalomyelitis (EAE) in mice lacking ADAP expression in platelets caused a more severe disease. In vivo administration of TGF-β1 early after T cell transfer improved EAE severity in mice with loss of ADAP restricted to platelets. Our results reveal a regulatory function of ADAP in platelets in vitro and during autoimmune disease EAE in vivo

Zinc Aspartate Induces IL-16 Secretion and Apoptosis in Human T Cells

T cell activation mediates immunity to pathogens. On the flipside, T cells are also involved in pathological immune responses during chronic autoimmune diseases. We recently reported that zinc aspartate, a registered drug with high bioavailability, dose-dependently inhibits T cell activation and Th1/Th2/Th17 cytokine production of stimulated human and mouse T cells. To understand the suppressive effect of zinc on T cell function, we here investigated the influence of zinc aspartate on human T cells focusing on the secretion of immunosuppressive cytokines, induction of apoptosis, and caspase 3/7 activity. To this end, we monitored either freshly stimulated or pre-activated human T cells in the presence of zinc aspartate from 40–140 µM over a period of 72 h. Under both experimental conditions, we observed a dose-dependent suppression of human T cell proliferation. While IL-1ra, latent TGF-β1, and IL-10 were dose-dependently reduced, we, unexpectedly, detected elevated levels of IL-16 upon zinc supplementation. In addition, the number of cells with active caspase 3/7 and, consecutively, the amount of cells undergoing apoptosis, steadily increased at zinc aspartate concentrations exceeding 100 µM. Taken together, our findings suggest that zinc aspartate impairs T cell fitness and might be beneficial for the treatment of T cell-mediated autoimmune diseases