The effector T-cell lineage shows great plasticity. Th17 cells are acknowledged to be instrumental in the response against microbial infection, but are also associated with autoimmune inflammatory processes. Here, we report that human regulatory T cells ( IntroductionIn mice, IL-17A (IL-17)-producing T cells have been established as an important T-helper (Th) effector lineage, clearly distinct from the Th1 or Th2 lineage. 1,2 Infectious disease mouse models indicate that IL-17-producing cells (Th17) mediate protection against extracellular pathogens. 3-5 However, on the downside, Th17 cells also appear to be the driving force in the pathogenesis of several autoimmune diseases. 2,6 Furthermore, mouse studies showed that differentiation of Th17 cells from naive CD4 T cells requires the concomitant activity of IL-6 and transforming growth factor-␤ (TGF␤). [7][8][9] The key transcription factor driving Th17 cell differentiation is the orphan nuclear receptor ROR␥t, which is needed for constitutive expression of In in vivo studies in mice, IL-17-and ROR␥t-expressing cells were shown to be present in the lung and digestive mucosal compartments, 11 and especially throughout the intestinal lamina propria. 10 In humans, IL-17 is associated with many inflammatory disorders such as rheumatoid arthritis, asthma, multiple sclerosis, lupus, Crohn disease, and allograft rejection. 3,[12][13][14][15] Recently, human ROR␥t-positive IL-17-producing T cells were identified under physiologic conditions in peripheral blood and tonsils, [15][16][17] being contained within the CCR6 ϩ (CCR4 ϩ )CD4 ϩ memory T-cell population. 15,16 Most recent data show that human Th17 differentiation, distinct from mouse Th17 development, is under control of IL-1␤, 19 In mice, the development of Th17 cells was described to be linked to that of regulatory T cells (Tregs) in a reciprocal fashion, whereby under the influence of TGF␤, IL-6 levels determine the outcome. 8,9 In vitro, activated Tregs promoted Th17 cell differentiation from CD4 T cells, 9,20 likely through their production of TGF␤.In addition, in vivo the transfer of Treg enhanced IL-17 production in a mouse model of systemic autoimmune disease. 21 Next, to this reciprocal relationship, it was recently demonstrated that IL-17-producing cells directly develop from mouse Tregs. 20 This is a remarkable finding, because Tregs are typically associated with T-cell tolerance and immune homeostasis. 22 Different Treg subsets have been described, and even within the naturally occurring CD4 pos CD25 high Foxp3 pos population heterogeneity is evident. This heterogeneity reflects differences in differentiation or developmental stage, trafficking properties, and suppressor capacity. [23][24][25] Although the lineage differentiation of helper T cells is very well accepted, that of Tregs is only scarcely recognized. In humans, naive and memory-like CD4 pos CD25 pos Foxp3 pos Treg have been defined in peripheral blood, discriminated by the expression of CD45 isoforms CD45RA and CD45RO. [25][26][27][28] Al...

Psoriasis is an autoimmune-related chronic inflammatory skin disease that is strongly associated with IL-23 and T helper-17 (Th17) effector cytokines. In addition, CD4+CD25(high) regulatory T-cell (Treg) function appeared to be impaired in psoriasis. CD4+CD25(high)Foxp3+ Tregs are typically considered inhibitors of autoimmune responses. However, under proinflammatory conditions, Tregs can differentiate into inflammation-associated Th17 cells--a paradigm shift, with as yet largely unknown consequences for human disease initiation or progression. Th17 cells are highly proinflammatory T cells that are characterized by IL-17A and IL-22 production and expression of the transcription factor retinoic acid-related orphan receptor γt (RORγt). We here show that Tregs of patients with severe psoriasis, as compared with those of healthy controls, have an enhanced propensity to differentiate into IL-17A-producing cells on ex vivo stimulation. This enhanced Treg differentiation was linked to unexpectedly high RORγt levels and enhanced loss of Foxp3. Notably, IL-23 boosted this Treg differentiation process particularly in patients with psoriasis but less so in controls. IL-23 further reduced Foxp3 expression while leaving the high RORγt levels unaffected. The histone/protein deacetylase inhibitor, Trichostatin-A, prevented Th17 differentiation of Tregs in psoriasis patients. Importantly, IL-17A+/Foxp3+/CD4+ triple-positive cells were present in skin lesions of patients with severe psoriasis. These data stress the clinical relevance of Treg differentiation for the perpetuation of chronic inflammatory disease and may pave novel ways for immunotherapy.

BCG vaccination in children protects against heterologous infections and improves survival independently of tuberculosis prevention. The phase III ACTIVATE trial assessed whether BCG has similar effects in the elderly. In this double-blind, randomized trial, elderly patients (n = 198) received BCG or placebo vaccine at hospital discharge and were followed for 12 months for new infections. At interim analysis, BCG vaccination significantly increased the time to first infection (median 16 weeks compared to 11 weeks after placebo). The incidence of new infections was 42.3% (95% CIs 31.9%–53.4%) after placebo vaccination and 25.0% (95% CIs 16.4%–36.1%) after BCG vaccination; most of the protection was against respiratory tract infections of probable viral origin (hazard ratio 0.21, p = 0.013). No difference in the frequency of adverse effects was found. Data show that BCG vaccination is safe and can protect the elderly against infections. Larger studies are needed to assess protection against respiratory infections, including COVID-19 (ClinicalTrials.gov NCT03296423).

Highlights d BCG vaccination of humans induces long-term immunophenotypic changes in neutrophils d BCG increases antimicrobial activity of neutrophils against unrelated pathogens d BCG-induced functional changes associate with modifications in histone methylation d Trained immunity may be a therapeutic target in neutrophilmediated diseases

The V-ATPase is the main regulator of intra-organellar acidification. Assembly of this complex has extensively been studied in yeast, while limited knowledge exists for man. We identified 11 male patients with hemizygous missense mutations in ATP6AP1, encoding accessory protein Ac45 of the V-ATPase. Homology detection at the level of sequence profiles indicated Ac45 as the long-sought human homologue of yeast V-ATPase assembly factor Voa1. Processed wild-type Ac45, but not its disease mutants, restored V-ATPase-dependent growth in Voa1 mutant yeast. Patients display an immunodeficiency phenotype associated with hypogammaglobulinemia, hepatopathy and a spectrum of neurocognitive abnormalities. Ac45 in human brain is present as the common, processed ∼40-kDa form, while liver shows a 62-kDa intact protein, and B-cells a 50-kDa isoform. Our work unmasks Ac45 as the functional ortholog of yeast V-ATPase assembly factor Voa1 and reveals a novel link of tissue-specific V-ATPase assembly with immunoglobulin production and cognitive function.

Regulatory T cell (Treg)-mediated immunosuppression is considered a major obstacle for successful cancer immunotherapy. The association between clinical outcome and Tregs is being studied extensively in clinical trials, but unfortunately, no consensus has been reached about (a) the markers and (b) the gating strategy required to define human Tregs in this context, making it difficult to draw final conclusions. Therefore, we have organized an international workshop on the detection and functional testing of Tregs with leading experts in the field, and 40 participants discussing different analyses and the importance of different markers and context in which Tregs were analyzed. This resulted in a rationally composed ranking list of “Treg markers”. Subsequently, the proposed Treg markers were tested to get insight into the overlap/differences between the most frequently used Treg definitions and their utility for Treg detection in various human tissues. Here, we conclude that the CD3, CD4, CD25, CD127, and FoxP3 markers are the minimally required markers to define human Treg cells. Staining for Ki67 and CD45RA showed to provide additional information on the activation status of Tregs. The use of markers was validated in a series of PBMC from healthy donors and cancer patients, as well as in tumor-draining lymph nodes and freshly isolated tumors. In conclusion, we propose an essential marker set comprising antibodies to CD3, CD4, CD25, CD127, Foxp3, Ki67, and CD45RA and a corresponding robust gating strategy for the context-dependent analysis of Tregs by flow cytometry.Electronic supplementary materialThe online version of this article (doi:10.1007/s00262-015-1729-x) contains supplementary material, which is available to authorized users.

The immune response to pathogens varies substantially among people. While both genetic and non-genetic factors contribute to inter-person variation, their relative contributions and potential predictive power have remained largely unknown. By systematically correlating host factors in 534 healthy volunteers, including baseline immunological parameters and molecular profiles (genome, metabolome and gut microbiome), with cytokine-production capacity after stimulation with 20 pathogens, we identified distinct patterns of co-regulation. Among the 91 different cytokine–stimulus pairs, 11 categories of host factors together explained up to 67% of inter-individual variation in cytokine production induced by stimulation. A computational model based on genetic data predicted the genetic component of stimulus-induced cytokine-production (correlation 0.28-0.89), while non-genetic factors influenced cytokine production as well.

BackgroundTreg based immunotherapy is of great interest to facilitate tolerance in autoimmunity and transplantation. For clinical trials, it is essential to have a clinical grade Treg isolation protocol in accordance with Good Manufacturing Practice (GMP) guidelines. To obtain sufficient Treg for immunotherapy, subsequent ex vivo expansion might be needed.Methodology/Principal FindingsTreg were isolated from leukapheresis products by CliniMACS based GMP isolation strategies, using anti-CD25, anti-CD8 and anti-CD19 coated microbeads. CliniMACS isolation procedures led to 40–60% pure CD4posCD25highFoxP3pos Treg populations that were anergic and had moderate suppressive activity. Such CliniMACS isolated Treg populations could be expanded with maintenance of suppressive function. Alloantigen stimulated expansion caused an enrichment of alloantigen-specific Treg. Depletion of unwanted CD19pos cells during CliniMACS Treg isolation proved necessary to prevent B-cell outgrowth during expansion. CD4posCD127pos conventional T cells were the major contaminating cell type in CliniMACS isolated Treg populations. Depletion of CD127pos cells improved the purity of CD4posCD25highFoxP3pos Treg in CliniMACS isolated cell populations to approximately 90%. Expanded CD127neg CliniMACS isolated Treg populations showed very potent suppressive capacity and high FoxP3 expression. Furthermore, our data show that cryopreservation of CliniMACS isolated Treg is feasible, but that activation after thawing is necessary to restore suppressive potential.Conclusions/SignificanceThe feasibility of Treg based therapy is widely accepted, provided that tailor-made clinical grade procedures for isolation and ex vivo cell handling are available. We here provide further support for this approach by showing that a high Treg purity can be reached, and that isolated cells can be cryopreserved and expanded successfully.