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Various autoimmune diseases are characterized by distinct cell subset distributions and activation profiles of peripheral blood mononuclear cells (PBMCs). PBMCs can therefore serve as an ideal biomarker material, which is easily accessible and allows for screening of multiple cell types. A detailed understanding of the immune landscape is critical for the diagnosis of patients with autoimmune diseases, as well as for a personalized treatment approach. In our study, we investigate the potential of multi-parameter spectral flow cytometry for the identification of patients suffering from autoimmune diseases and its power as an evaluation tool for in vitro drug screening approaches (advanced immunophenotyping). We designed a combination of two 22-color immunophenotyping panels for profiling cell subset distribution and cell activation. Downstream bioinformatics analyses included percentages of individual cell populations and median fluorescent intensity of defined markers which were then visualized as heatmaps and in dimensionality reduction approaches. In vitro testing of epigenetic immunomodulatory drugs revealed an altered activation status upon treatment, which supports the use of spectral flow cytometry as a high-throughput drug screening tool. Advanced immunophenotyping might support the exploration of novel therapeutic drugs and contribute to future personalized treatment approaches in autoimmune diseases and beyond., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Preglej, Brinkmann, Steiner, Aletaha, Göschl and Bonelli.)

Background: Patients with inborn errors of immunity (IEI) are at increased risk for severe courses of SARS-CoV-2 infection. COVID-19 vaccination provides effective protection in healthy individuals. However, it remains unclear whether vaccination is efficient and safe in patients with constitutional dysfunctions of the immune system. Thus, we analyzed the humoral response, adverse reactions and assessed the disease activity of the underlying disease after COVID-19 vaccination in a cohort of patients suffering from IEIs or mannan-binding lectin deficiency (MBLdef)., Methods: Vaccination response was assessed after basic immunization using the Elecsys anti-SARS-CoV-2 S immunoassay and via Vero E6 cell based assay to detect neutralization capabilities. Phenotyping of lymphocytes was performed by flow cytometry. Patient charts were reviewed for disease activity, autoimmune phenomena as well as immunization status and reactogenicity of the vaccination. Activity of the underlying disease was assessed using a patient global numeric rating scale (NRS)., Results: Our cohort included 11 individuals with common variable immunodeficiency (CVID), one patient with warts hypogammaglobulinemia immunodeficiency myelokathexis (WHIM) syndrome, two patients with X-linked agammaglobulinemia (XLA), one patient with Muckle Wells syndrome, two patients with cryopyrin-associated periodic syndrome, one patient with Interferon-gamma (IFN-gamma) receptor defect, one patient with selective deficiency in pneumococcal antibody response combined with a low MBL level and seven patients with severe MBL deficiency. COVID-19 vaccination was generally well tolerated with little to no triggering of autoimmune phenomena. 20 out of 26 patients developed an adequate humoral vaccine response. 9 out of 11 patients developed a T cell response comparable to healthy control subjects. Tested immunoglobulin replacement therapy (IgRT) preparations contained Anti-SARS-CoV-2 S antibodies implicating additional protection through IgRT., Summary: In summary the data support the efficacy and safety of a COVID-19 vaccination in patients with IEIs/MBLdef. We recommend evaluation of the humoral immune response and testing for virus neutralization after vaccination in this cohort., Competing Interests: DA is a member of the editorial board of “Annals of the Rheumatic diseases”. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Göschl, Mrak, Grabmeier-Pfistershammer, Stiasny, Haslacher, Schneider, Deimel, Kartnig, Tobudic, Aletaha, Burgmann, Bonelli, Pickl, Förster-Waldl, Scheinecker and Vossen.)

Impaired response to COVID-19 vaccination is of particular concern in immunosuppressed patients. To determine the best vaccination strategy for this vulnerable group we performed a single center, 1:1 randomized blinded clinical trial. Patients who failed to seroconvert upon two mRNA vaccinations (BNT162b2 or mRNA-1273) are randomized to receive either a third dose of the same mRNA or the vector vaccine ChAdOx1 nCoV-19. Primary endpoint is the difference in SARS-CoV-2 spike antibody seroconversion rate between vector and mRNA vaccinated patients four weeks after the third dose. Secondary outcomes include cellular immune responses. Seroconversion rates at week four are significantly higher in the mRNA (homologous vaccination, 15/24, 63%) as compared to the vector vaccine group (heterologous vaccination, 4/22, 18%). SARS-CoV-2-specific T-cell responses are reduced but could be increased after a third dose of either vector or mRNA vaccine. In a multivariable logistic regression analysis, patient age and vaccine type are associated with seroconversion. No serious adverse event is attributed to COVID-19 booster vaccination. Efficacy and safety data underline the importance of a booster vaccination and support the use of a homologous mRNA booster vaccination in immunosuppressed patients.Trial registration: EudraCT No.: 2021-002693-10., (© 2022. The Author(s).)

A 53-year old female patient with history of hypocomplementaemic urticarial vasculitis syndrome (HUVS) and polyarteritis nodosa presented with progressive dyspnoea on exertion due to emphysema. Lung function revealed a severe obstructive ventilator disorder with a forced expiratory volume in 1 second of 22% of predicted, and a significant hyperinflation with a residual volume of 321% of predicted. Multi-detector computed tomography (MDCT) scan and quantitative CT analysis (StratX software) confirmed a lower lobe predominant emphysema. Considering the young age, the very severely impaired lung function, the relatively low nicotine abuse, the exclusion of alpha-1 antitrypsin deficiency, together with the known diagnosis of HUVS, the emphysema was more likely due to the vasculitis than to a typical chronic obstructive lung disease. MDCT scan showed that particularly the segment 8 of the right lower lobe was severely emphysematous destroyed and hyperinflated. Invasive Chartis® measurement revealed no significant collateral ventilation of the isolated segment 8 of the right lower lobe, so that an endobronchial valve placement was performed. Three months following intervention, the MDCT scan revealed a complete collapse of the segment 8 on the right, which was associated with a significant clinical benefit and a mild reduction of the hyperinflation in the lung function test., (© 2022 The Authors. Published by Elsevier Ltd.)

Some effector CD4+ T cell subsets display cytotoxic activity, thus breaking the functional dichotomy of CD4+ helper and CD8+ cytotoxic T lymphocytes. However, molecular mechanisms regulating CD4+ cytotoxic T lymphocyte (CD4+ CTL) differentiation are poorly understood. Here we show that levels of histone deacetylases 1 and 2 (HDAC1-HDAC2) are key determinants of CD4+ CTL differentiation. Deletions of both Hdac1 and 1 Hdac2 alleles (HDAC1cKO-HDAC2HET) in CD4+ T cells induced a T helper cytotoxic program that was controlled by IFN-γ-JAK1/2-STAT1 signaling. In vitro, activated HDAC1cKO-HDAC2HET CD4+ T cells acquired cytolytic activity and displayed enrichment of gene signatures characteristic of effector CD8+ T cells and human CD4+ CTLs. In vivo, murine cytomegalovirus-infected HDAC1cKO-HDAC2HET mice displayed a stronger induction of CD4+ CTL features compared with infected WT mice. Finally, murine and human CD4+ T cells treated with short-chain fatty acids, which are commensal-produced metabolites acting as HDAC inhibitors, upregulated CTL genes. Our data demonstrate that HDAC1-HDAC2 restrain CD4+ CTL differentiation. Thus, HDAC1-HDAC2 might be targets for the therapeutic induction of CD4+ CTLs.

Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease, characterized by synovial infiltration of various inflammatory cells. Chemokines are involved in controlling the recruitment of different cell types into the synovial membrane. The role of CCR6 in the development of arthritis so far remains unclear. In this study, we investigated the role of CCR6 in the pathogenesis of arthritis using three different murine arthritis models. Compared to WT animals, CCR6 -/- mice developed less clinical signs of arthritis in the collagen-induced arthritis model but not in the K/BxN serum transfer arthritis model and in the human tumour necrosis factor transgenic arthritis model, suggesting a defect in adaptive effector functions but intact innate effector functions in the development of arthritis in CCR6 -/- animals. In line with this, anti-collagen antibody levels were significantly reduced in CCR6 -/- mice compared with WT mice. Moreover, we demonstrate enhanced osteoclastogenesis in vitro in CCR6 -/- mice compared with WT mice. However, we did not detect differences in bone mass under steady state conditions in vivo between WT and CCR6-deficient mice. These data suggest that CCR6 is crucially involved in adaptive but not in innate immunity-driven arthritis. CCR6 or its chemokine ligand CCL20 might represent a possible new target for the treatment of RA., (© 2018 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

Objective: Abatacept (CTLA-4Ig) blocks CD28-mediated T cell activation by binding to the costimulatory B7 ligands CD80/CD86 on antigen presenting cells. Costimulatory molecules, however, can also be expressed on T cells upon activation. Therefore, the aim of our study was to investigate direct effects of CTLA-4Ig on distinct T cell subsets in RA patients., Methods: Phenotypic and functional analyses of CD4(+) T cells, including CD4(+) FoxP3(+) CD25(+) regulatory T cells (Treg), from RA patients were performed before and during CTLA-4Ig therapy. In addition T cells from healthy volunteers were analysed on in vitro culture with CTLA-4Ig or anti-CD80 and anti-CD86 antibodies. Apoptotic DNA fragmentation in CD4(+) and CD4(+) FoxP3(+) T cells was measured by TUNEL staining., Results: We observed an increase in T cells, including Treg cells, after initiation of CTLA-4Ig therapy, which was linked to a downregulation of activation-associated marker molecules and CD95 on CD4(+) T cells and Treg cells. CTLA-4Ig decreased CD95-mediated cell death in vitro in a dose-dependent manner. Functional analysis of isolated Treg cells from RA patients further revealed a diminished suppression of responder T cell proliferation. This was found to be due to CTLA-4Ig-mediated blocking of CD80 and CD86 on responder T cells that led to a diminished susceptibility for Treg cell suppression., Conclusion: CTLA-4Ig therapy in RA patients exerts effects beyond the suppression of T cell activation, which has to be taken into account as an additional mechanism of CTLA-4Ig treatment., (© The Author 2015. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Reversible lysine acetylation plays an important role in the regulation of T cell responses. HDAC1 has been shown to control peripheral T helper cells, however the role of HDAC1 in CD8+ T cell function remains elusive. By using conditional gene targeting approaches, we show that LckCre-mediated deletion of HDAC1 led to reduced numbers of thymocytes as well as peripheral T cells, and to an increased fraction of CD8+CD4- cells within the CD3/TCRβlo population, indicating that HDAC1 is essential for the efficient progression of immature CD8+CD4- cells to the DP stage. Moreover, CD44hi effector CD8+ T cells were enhanced in mice with a T cell-specific deletion of HDAC1 under homeostatic conditions and HDAC1-deficient CD44hi CD8+ T cells produced more IFNγ upon ex vivo PMA/ionomycin stimulation in comparison to wild-type cells. Naïve (CD44l°CD62L+) HDAC1-null CD8+ T cells displayed a normal proliferative response, produced similar amounts of IL-2 and TNFα, slightly enhanced amounts of IFNγ, and their in vivo cytotoxicity was normal in the absence of HDAC1. However, T cell-specific loss of HDAC1 led to a reduced anti-viral CD8+ T cell response upon LCMV infection and impaired expansion of virus-specific CD8+ T cells. Taken together, our data indicate that HDAC1 is required for the efficient generation of thymocytes and peripheral T cells, for proper CD8+ T cell homeostasis and for an efficient in vivo expansion and activation of CD8+ T cells in response to LCMV infection.

Introduction: Systemic lupus erythematosus (SLE) is a heterogenous autoimmune disease, which can affect different organs. Increased proportions of CD4⁺CD25-Foxp3⁺ T cells have been described in SLE patients. The exact role of this cell population in SLE patients still remains unclear. We therefore analyzed this T cell subset in a large cohort of SLE patients with different organ manifestations., Methods: Phenotypic analyses, proportions and absolute cell numbers of CD4⁺CD25-Foxp3⁺ T cells were determined by flow cytometry (FACS) in healthy controls (HC) (n = 36) and SLE patients (n = 61) with different organ manifestations. CD4⁺CD25⁻Foxp3⁺ T cells were correlated with clinical data, the immunosuppressive therapy and different disease activity indices. In patients with active glomerulonephritis, CD4⁺CD25⁻Foxp3⁺ T cells were analyzed in urine sediment samples. Time course analyses of CD4⁺CD25⁻Foxp3⁺ T cells were performed in patients with active disease activity before and after treatment with cyclophosphamide and prednisone., Results: CD4⁺CD25⁻Foxp3⁺ T cells were significantly increased in active SLE patients and the majority expressed Helios. Detailed analysis of this patient cohort revealed increased proportions of CD4⁺CD25⁻Foxp3⁺ T cells in SLE patients with renal involvement. CD4⁺CD25⁻Foxp3⁺ T cells were also detected in urine sediment samples of patients with active glomerulonephritis and correlated with the extent of proteinuria., Conclusion: CD4⁺CD25⁻Foxp3⁺ T cells resemble regulatory rather than activated T cells. Comparative analysis of CD4⁺CD25⁻Foxp3⁺ T cells in SLE patients revealed a significant association of this newly described cell population with active nephritis. Therefore CD4⁺CD25⁻Foxp3⁺ T cells might serve as an important tool to recognize and monitor SLE patients with renal involvement.