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Infliximab (IFX) is a frequent therapeutic option for Crohn disease (CD) patients. Early detection of responders to IFX is critical for the management of CD in order to avoid long-term exposure to the drug without benefit. This retrospective study aimed at analysing which early parameters recorded during the induction period are able to predict response to IFX during the maintenance period in pediatric CD.Medical records of all CD patients ages from 2 to 18 years who received IFX at a tertiary IBD center were retrospectively analyzed. Children were classified in 3 groups according to their response at week 14 (W14) remission, clinical response or , no response. The factors recorded at W0, W2, and W6, which were associated with remission at W14 were analyzed using a logistic regression.Among the 111 patients included, 74.8% patients were responders to IFX at W14, including 38.7% in clinical remission and 36% with partial clinical response. Clinical remission at W14 was associated with normal growth (P 0.01), and normal albuminemia (P = 0.01) at baseline, It was also associated with trough levels to IFX8.3 μg/ml at week 6 (P 0.01).Trough levels to IFX8.3 μg/ml at week 6 are predictive of remission at W14 for luminal disease.

There is a Blood Commentary on this article in this issue., Key Points As a proof of concept, COVID-19 convalescent plasma represents an interesting approach in B-cell–depleted patients with protracted COVID-19.COVID-19 convalescent plasma induces a decrease in temperature and inflammatory parameters within 1 week associated with oxygen weaning., Anti-CD20 monoclonal antibodies are widely used for the treatment of hematological malignancies or autoimmune disease but may be responsible for a secondary humoral deficiency. In the context of COVID-19 infection, this may prevent the elicitation of a specific SARS-CoV-2 antibody response. We report a series of 17 consecutive patients with profound B-cell lymphopenia and prolonged COVID-19 symptoms, negative immunoglobulin G (IgG)-IgM SARS-CoV-2 serology, and positive RNAemia measured by digital polymerase chain reaction who were treated with 4 units of COVID-19 convalescent plasma. Within 48 hours of transfusion, all but 1 patient experienced an improvement of clinical symptoms. The inflammatory syndrome abated within a week. Only 1 patient who needed mechanical ventilation for severe COVID-19 disease died of bacterial pneumonia. SARS-CoV-2 RNAemia decreased to below the sensitivity threshold in all 9 evaluated patients. In 3 patients, virus-specific T-cell responses were analyzed using T-cell enzyme-linked immunospot assay before convalescent plasma transfusion. All showed a maintained SARS-CoV-2 T-cell response and poor cross-response to other coronaviruses. No adverse event was reported. Convalescent plasma with anti–SARS-CoV-2 antibodies appears to be a very promising approach in the context of protracted COVID-19 symptoms in patients unable to mount a specific humoral response to SARS-CoV-2., Visual Abstract

ObjectivesThe emergence of strains of SARS-CoV-2 exhibiting increase viral fitness and immune escape potential, such as the Delta variant (B.1.617.2), raises concerns in immunocompromised patients. We aimed to evaluate seroconversion, cross-neutralisation and T-cell responses induced by BNT162b2 in immunocompromised patients with systemic inflammatory diseases.MethodsProspective monocentric study including patients with systemic inflammatory diseases and healthcare immunocompetent workers as controls. Primary endpoints were anti-spike antibodies levels and cross-neutralisation of Alpha and Delta variants after BNT162b2 vaccine. Secondary endpoints were T-cell responses, breakthrough infections and safety.ResultsSixty-four cases and 21 controls not previously infected with SARS-CoV-2 were analysed. Kinetics of anti-spike IgG after BNT162b2 vaccine showed lower and delayed induction in cases, more pronounced with rituximab. Administration of two doses of BNT162b2 generated a neutralising response against Alpha and Delta in 100% of controls, while sera from only one of rituximab-treated patients neutralised Alpha (5%) and none Delta. Other therapeutic regimens induced a partial neutralising activity against Alpha, even lower against Delta. All controls and cases except those treated with methotrexate mounted a SARS-CoV-2 specific T-cell response. Methotrexate abrogated T-cell responses after one dose and dramatically impaired T-cell responses after two doses of BNT162b2. Third dose of vaccine improved immunogenicity in patients with low responses.ConclusionRituximab and methotrexate differentially impact the immunogenicity of BNT162b2, by impairing B-cell and T-cell responses, respectively. Delta fully escapes the humoral response of individuals treated with rituximab. These findings support efforts to improve BNT162b2 immunogenicity in immunocompromised individuals (ClinicalTrials.gov number, NCT04870411).

International audience; Significance: The vast majority of autoimmune diseases are polygenic, and causal loci uncovered by genetic-mapping studies explain only a minority of the heritable contribution to trait variation. Multiple explanations for this missing heritability include rare meaningful variants, rare copy number variations or deletions, epistasis, epigenetics, disease heterogeneity, and rare or infrequent variants that segregate within individual families (even within monozygotic twins). Here we demonstrate that experimental models of spontaneous autoimmune diseases may be invaluable tools to map rare germline variants impacting disease susceptibility traits. We identified a variant of the dual-specificity phosphatase 10 encoding gene that accelerates disease in an autoimmune type 1 diabetes model, the nonobese diabetic mouse.Abstract: Insulin-dependent or type 1 diabetes (T1D) is a polygenic autoimmune disease. In humans, more than 60 loci carrying common variants that confer disease susceptibility have been identified by genome-wide association studies, with a low individual risk contribution for most variants excepting those of the major histocompatibility complex (MHC) region (40 to 50% of risk); hence the importance of missing heritability due in part to rare variants. Nonobese diabetic (NOD) mice recapitulate major features of the human disease including genetic aspects with a key role for the MHC haplotype and a series of Idd loci. Here we mapped in NOD mice rare variants arising from genetic drift and significantly impacting disease risk. To that aim we established by selective breeding two sublines of NOD mice from our inbred NOD/Nck colony exhibiting a significant difference in T1D incidence. Whole-genome sequencing of high (H)- and low (L)-incidence sublines (NOD/NckH and NOD/NckL) revealed a limited number of subline-specific variants. Treating age of diabetes onset as a quantitative trait in automated meiotic mapping (AMM), enhanced susceptibility in NOD/NckH mice was unambiguously attributed to a recessive missense mutation of Dusp10, which encodes a dual specificity phosphatase. The causative effect of the mutation was verified by targeting Dusp10 with CRISPR-Cas9 in NOD/NckL mice, a manipulation that significantly increased disease incidence. The Dusp10 mutation resulted in islet cell down-regulation of type I interferon signature genes, which may exert protective effects against autoimmune aggression. De novo mutations akin to rare human susceptibility variants can alter the T1D phenotype.

Objectives In adult inflammatory bowel disease (IBD) treated by anti-TNF antibodies, paradoxical psoriasis has an estimated prevalence of 1.6 to 22%, especially in infliximab (IFX)-treated patients. Little is known in the pediatric IBD (PIBD) populations. Methods All patients ages from 2 to 18 years with Crohn disease (CD) or ulcerative colitis (UC) and treated for the first time by IFX between January 2002 and March 2014, were considered for inclusion in this retrospective study performed in a tertiary PIBD centre. Paradoxical psoriasis events together with clinical and biological data were collected in all patients. Comparisons between psoriasis and control groups were performed using univariate statistical analyses. Results One hundred and twenty-three CD patients and 24 UC patients were treated with IFX. Twenty patients (13.6%) experienced a paradoxical psoriasis. All of them were affected by CD. Perianal CD was more frequent in the psoriasis group (P = 0.033). Fourteen patients (70%) were in remission when skin lesions occurred. Paradoxical psoriasis was diagnosed 355 days (median, interquartile range [IQR] 239; 532) after the initiation of IFX corresponding to the eighth injection (median, IQR: 6; 15). Psoriasis lesions were controlled by local steroids in all cases and no patients discontinued IFX therapy. Conclusions 13.6% of our IBD patients treated with IFX developed psoriasis during a median follow-up of 23.9 months (IQR: 11.6; 36.5). Crohn disease patients with perianal disease were at a higher risk to develop this common side effect.

All authors: Andrea Cossarizza Hyun‐Dong Chang Andreas Radbruch Andreas Acs Dieter Adam Sabine Adam‐Klages William W. Agace Nima Aghaeepour Mübeccel Akdis Matthieu Allez Larissa Nogueira Almeida Giorgia Alvisi Graham Anderson Immanuel Andrä Francesco Annunziato Achille Anselmo Petra Bacher Cosima T. Baldari Sudipto Bari Vincenzo Barnaba Joana Barros‐Martins Luca Battistini Wolfgang Bauer Sabine Baumgart Nicole Baumgarth Dirk Baumjohann Bianka Baying Mary Bebawy Burkhard Becher Wolfgang Beisker Vladimir Benes Rudi Beyaert Alfonso Blanco Dominic A. Boardman Christian Bogdan Jessica G. Borger Giovanna Borsellino Philip E. Boulais Jolene A. Bradford Dirk Brenner Ryan R. Brinkman Anna E. S. Brooks Dirk H. Busch Martin Büscher Timothy P. Bushnell Federica Calzetti Garth Cameron Ilenia Cammarata Xuetao Cao Susanna L. Cardell Stefano Casola Marco A. Cassatella Andrea Cavani Antonio Celada Lucienne Chatenoud Pratip K. Chattopadhyay Sue Chow Eleni Christakou Luka Čičin‐Šain Mario Clerici Federico S. Colombo Laura Cook Anne Cooke Andrea M. Cooper Alexandra J. Corbett Antonio Cosma Lorenzo Cosmi Pierre G. Coulie Ana Cumano Ljiljana Cvetkovic Van Duc Dang Chantip Dang‐Heine Martin S. Davey Derek Davies Sara De Biasi Genny Del Zotto Gelo Victoriano Dela Cruz Michael Delacher Silvia Della Bella Paolo Dellabona Günnur Deniz Mark Dessing James P. Di Santo Andreas Diefenbach Francesco Dieli Andreas Dolf Thomas Dörner Regine J. Dress Diana Dudziak Michael Dustin Charles‐Antoine Dutertre Friederike Ebner Sidonia B. G. Eckle Matthias Edinger Pascale Eede Götz R.A. Ehrhardt Marcus Eich Pablo Engel Britta Engelhardt Anna Erdei Charlotte Esser Bart Everts Maximilien Evrard Christine S. Falk Todd A. Fehniger Mar Felipo‐Benavent Helen Ferry Markus Feuerer Andrew Filby Kata Filkor Simon Fillatreau Marie Follo Irmgard Förster John Foster Gemma A. Foulds Britta Frehse Paul S. Frenette Stefan Frischbutter Wolfgang Fritzsche David W. Galbraith Anastasia Gangaev Natalio Garbi Brice Gaudilliere Ricardo T. Gazzinelli Jens Geginat Wilhelm Gerner Nicholas A. Gherardin Kamran Ghoreschi Lara Gibellini Florent Ginhoux Keisuke Goda Dale I. Godfrey Christoph Goettlinger Jose M. González‐Navajas Carl S. Goodyear Andrea Gori Jane L. Grogan Daryl Grummitt Andreas Grützkau Claudia Haftmann Jonas Hahn Hamida Hammad Günter Hämmerling Leo Hansmann Goran Hansson Christopher M. Harpur Susanne Hartmann Andrea Hauser Anja E. Hauser David L. Haviland David Hedley Daniela C. Hernández Guadalupe Herrera Martin Herrmann Christoph Hess Thomas Höfer Petra Hoffmann Kristin Hogquist Tristan Holland Thomas Höllt Rikard Holmdahl Pleun Hombrink Jessica P. Houston Bimba F. Hoyer Bo Huang Fang‐Ping Huang Johanna E. Huber Jochen Huehn Michael Hundemer Christopher A. Hunter William Y. K. Hwang Anna Iannone Florian Ingelfinger Sabine M Ivison Hans‐Martin Jäck Peter K. Jani Beatriz Jávega Stipan Jonjic Toralf Kaiser Tomas Kalina Thomas Kamradt Stefan H. E. Kaufmann Baerbel Keller Steven L. C. Ketelaars Ahad Khalilnezhad Srijit Khan Jan Kisielow Paul Klenerman Jasmin Knopf Hui‐Fern Koay Katja Kobow Jay K. Kolls Wan Ting Kong Manfred Kopf Thomas Korn Katharina Kriegsmann Hendy Kristyanto Thomas Kroneis Andreas Krueger Jenny Kühne Christian Kukat Désirée Kunkel Heike Kunze‐Schumacher Tomohiro Kurosaki Christian Kurts Pia Kvistborg Immanuel Kwok Jonathan Landry Olivier Lantz Paola Lanuti Francesca LaRosa Agnès Lehuen Salomé LeibundGut‐Landmann Michael D. Leipold Leslie Y.T. Leung Megan K. Levings Andreia C. Lino Francesco Liotta Virginia Litwin Yanling Liu Hans‐Gustaf Ljunggren Michael Lohoff Giovanna Lombardi Lilly Lopez Miguel López‐Botet Amy E. Lovett‐Racke Erik Lubberts Herve Luche Burkhard Ludewig Enrico Lugli Sebastian Lunemann Holden T. Maecker Laura Maggi Orla Maguire Florian Mair Kerstin H. Mair Alberto Mantovani Rudolf A. Manz Aaron J. Marshall Alicia Martínez‐Romero Glòria Martrus Ivana Marventano Wlodzimierz Maslinski Giuseppe Matarese Anna Vittoria Mattioli Christian Maueröder Alessio Mazzoni James McCluskey Mairi McGrath Helen M. McGuire Iain B. McInnes Henrik E. Mei Fritz Melchers Susanne Melzer Dirk Mielenz Stephen D. Miller Kingston H.G. Mills Hans Minderman Jenny Mjösberg Jonni Moore Barry Moran Lorenzo Moretta Tim R. Mosmann Susann Müller Gabriele Multhoff Luis Enrique Muñoz Christian Münz Toshinori Nakayama Milena Nasi Katrin Neumann Lai Guan Ng Antonia Niedobitek Sussan Nourshargh Gabriel Núñez José‐Enrique O'Connor Aaron Ochel Anna Oja Diana Ordonez Alberto Orfao Eva Orlowski‐Oliver Wenjun Ouyang Annette Oxenius Raghavendra Palankar Isabel Panse Kovit Pattanapanyasat Malte Paulsen Dinko Pavlinic Livius Penter Pärt Peterson Christian Peth Jordi Petriz Federica Piancone Winfried F. Pickl Silvia Piconese Marcello Pinti A. Graham Pockley Malgorzata Justyna Podolska Zhiyong Poon Katharina Pracht Immo Prinz Carlo E. M. Pucillo Sally A. Quataert Linda Quatrini Kylie M. Quinn Helena Radbruch Tim R. D. J. Radstake Susann Rahmig Hans‐Peter Rahn Bartek Rajwa Gevitha Ravichandran Yotam Raz Jonathan A. Rebhahn Diether Recktenwald Dorothea Reimer Caetano Reis e Sousa Ester B.M. Remmerswaal Lisa Richter Laura G. Rico Andy Riddell Aja M. Rieger J. Paul Robinson Chiara Romagnani Anna Rubartelli Jürgen Ruland Armin Saalmüller Yvan Saeys Takashi Saito Shimon Sakaguchi Francisco Sala‐de‐Oyanguren Yvonne Samstag Sharon Sanderson Inga Sandrock Angela Santoni Ramon Bellmàs Sanz Marina Saresella Catherine Sautes‐Fridman Birgit Sawitzki Linda Schadt Alexander Scheffold Hans U. Scherer Matthias Schiemann Frank A. Schildberg Esther Schimisky Andreas Schlitzer Josephine Schlosser Stephan Schmid Steffen Schmitt Kilian Schober Daniel Schraivogel Wolfgang Schuh Thomas Schüler Reiner Schulte Axel Ronald Schulz Sebastian R. Schulz Cristiano Scottá Daniel Scott‐Algara David P. Sester T. Vincent Shankey Bruno Silva‐Santos Anna Katharina Simon Katarzyna M. Sitnik Silvano Sozzani Daniel E. Speiser Josef Spidlen Anders Stahlberg Alan M. Stall Natalie Stanley Regina Stark Christina Stehle Tobit Steinmetz Hannes Stockinger Yousuke Takahama Kiyoshi Takeda Leonard Tan Attila Tárnok Gisa Tiegs Gergely Toldi Julia Tornack Elisabetta Traggiai Mohamed Trebak Timothy I.M. Tree Joe Trotter John Trowsdale Maria Tsoumakidou Henning Ulrich Sophia Urbanczyk Willem van de Veen Maries van den Broek Edwin van der Pol Sofie Van Gassen Gert Van Isterdael René A.W. van Lier Marc Veldhoen Salvador Vento‐Asturias Paulo Vieira David Voehringer Hans‐Dieter Volk Anouk von Borstel Konrad von Volkmann Ari Waisman Rachael V. Walker Paul K. Wallace Sa A. Wang Xin M. Wang Michael D. Ward Kirsten A Ward‐Hartstonge Klaus Warnatz Gary Warnes Sarah Warth Claudia Waskow James V. Watson Carsten Watzl Leonie Wegener Thomas Weisenburger Annika Wiedemann Jürgen Wienands Anneke Wilharm Robert John Wilkinson Gerald Willimsky James B. Wing Rieke Winkelmann Thomas H. Winkler Oliver F. Wirz Alicia Wong Peter Wurst Jennie H. M. Yang Juhao Yang Maria Yazdanbakhsh Liping Yu Alice Yue Hanlin Zhang Yi Zhao Susanne Maria Ziegler Christina Zielinski Jakob Zimmermann Arturo Zychlinsky., These guidelines are a consensus work of a considerable number of members of the immunology and flow cytometry community. They provide the theory and key practical aspects of flow cytometry enabling immunologists to avoid the common errors that often undermine immunological data. Notably, there are comprehensive sections of all major immune cell types with helpful Tables detailing phenotypes in murine and human cells. The latest flow cytometry techniques and applications are also described, featuring examples of the data that can be generated and, importantly, how the data can be analysed. Furthermore, there are sections detailing tips, tricks and pitfalls to avoid, all written and peer‐reviewed by leading experts in the field, making this an essential research companion., This work was supported by the Netherlands Organisation for Scientific Research – Domain Applied and Engineering Sciences (NWO-TTW), research program VENI 15924. This work was funded by the Deutsche Forschungsgemeinschaft. European Union Innovative Medicines Initiative - Joint Undertaking - RTCure Grant Agreement 777357 and innovation program (Grant Agreement 695551).

Immunity to the SARS-CoV-2 virus ensures protection against reinfection by this virus thanks to the combined action of neutralizing antibodies and T lymphocytes specific to viral proteins, in particular the Spike protein. It must be distinguished from the immune response that ensures healing of the infection following contamination that involves innate immunity, particularly type 1 interferons, and which is followed by adaptive cellular and humoral immunity. The importance of the effect of interferons is highlighted by the occurrence of severe forms of the disease in genetically deficient subjects or in patients with antibodies neutralizing type 1 interferon. Herd immunity is not an individual biological property. It is a mathematical property that qualifies the fact that when the proportion of subjects with individual immunity is high enough, there is little chance that an epidemic can occur. The level of that proportion-the herd immunity of the population can be computed under theoretical, often unrealistic, hypotheses, and is difficult to assess in natural conditions.L’immunité individuelle contre le virus SARS-CoV-2 assure la protection contre la réinfection par ce virus grâce à l’action conjuguée des anticorps neutralisants et des lymphocytes T spécifiques des protéines virales, notamment la protéine Spike (spicule). Il faut la distinguer de la réponse immunitaire qui assure la guérison de l’infection dans les jours suivant la contamination. Celle-ci fait intervenir l’immunité innée et tout particulièrement les interférons de type 1 puis l’immunité adaptative cellulaire et humorale. L’importance de l’effet des interférons est soulignée par la survenue de formes graves chez des sujets génétiquement déficients dans leur synthèse ou encore des patients présentant des anticorps neutralisant l’interféron de type 1. L’immunité collective caractérise la faible probabilité de développement d’une épidémie dans une population ayant un pourcentage élevé de sujets présentant une immunité individuelle. Le taux d’immunité collective nécessaire pour faire disparaître l’épidémie a été calculé dans des modèles mathématiques supposant la panmixie ; il est difficile à évaluer dans les populations réelles.

International audience; The link between SARS‐CoV‐2 and the reported cutaneous manifestations has not been established. We assessed a possible correlation between the paediatric dermatological manifestations and the biological investigations, using for the first time 3 different SARS‐CoV‐2 tests.

International audience; BackgroundMast cells are key players in innate immunity and the TH2 adaptive immune response. The latter counterbalances the TH1 response, which is critical for antiviral immunity. Clonal mast cell activation disorders (cMCADs, such as mastocytosis and clonal mast cell activation syndrome) are characterized by abnormal mast cell accumulation and/or activation. No data on the antiviral immune response in patients with MCADs have been published.ObjectiveTo study a comprehensive range of outcomes in patients with cMCAD with PCR- or serologically confirmed coronavirus disease 2019 and to characterize the specific anti–severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) immune response in this setting.MethodsClinical follow-up and outcome data were collected prospectively over a 12-month period by members of the French Centre de Référence des Mastocytoses rare disease network. Anti–SARS-CoV-2–specific T-cell activity was measured with an ELISA, and humoral responses were evaluated by assaying circulating levels of specific IgG, IgA, and neutralizing antibodies.ResultsOverall, 32 patients with cMCAD were evaluated. None required noninvasive or mechanical ventilation. Two patients were admitted to hospital for oxygen and steroid therapy. The SARS-CoV-2–specific immune response was characterized in 21 of the 32 patients. Most had high counts of circulating SARS-CoV-2–specific, IFN-γ–producing T cells and high titers of neutralizing antispike IgGs. The patients frequently showed spontaneous T-cell IFN-γ production in the absence of stimulation; this production was correlated with basal circulating tryptase levels (a marker of the mast cell burden).ConclusionsPatients with cMCADs might not be at risk of severe coronavirus disease 2019, perhaps due to their spontaneous production of IFN-γ.

Background Chronic disseminated candidiasis (CDC) is a rare disease that mostly occurs after chemotherapy-induced prolonged neutropenia in patients with hematological malignancies. It is believed to ensue from Candida colonization, breach of the intestinal epithelial barrier, and venous translocation to organs. Fungal blood or liver biopsy cultures are generally negative, suggesting the absence of an ongoing invasive fungal disease. Methods To unravel the contribution of the immune system to CDC pathogenesis, we undertook a prospective multicentric exploratory study in 44 CDC patients at diagnosis and 44 matched controls. Results Analysis of Candida-specific T-cell responses using enzyme-linked immunospot assays revealed higher numbers of interferon (IFN)γ-producing T cells reactive to mp65 or candidin in 27 CDC cases compared with 33 controls. Increased plasma levels of soluble CD25, interleukin (IL)-6, IL-1β, tumor necrosis factor-α, and IL-10 and lower levels of IL-2 were observed in CDC patients versus controls. Neutrophilia and higher levels of CD4 and CD8 T-cell activation were found in CDC patients as well as increased proportions of CXCR3-expressing TCRγδ +Vδ2+ cells. Conclusions The expansion of Candida-specific IFNγ-producing T cells together with features of T-cell activation and systemic inflammation identified here support the view that CDC belongs to the broad spectrum of fungal-associated immune reconstitution inflammatory syndromes.

IL-17 and mucosal-associated invariant T (MAIT) cells have been involved in asthma pathogenesis. However, IL-17-producing MAIT cells (MAIT-17) were not evidenced. We aimed to determine whether circulating MAIT-17 were detectable in children with asthma, and whether they correlated with asthma symptoms or lung function. Children from the SPASM cohort of preschoolers with severe wheeze were reassessed for asthma at school age, and categorized as exacerbators (1 or more severe exacerbations in the previous 12months) or non-exacerbators. Nineteen children (10.9years) were included (9 non-exacerbators, 10 exacerbators). Circulating MAIT-17 were detected by flow cytometry. Their frequency was higher in exacerbators than in non-exacerbators (1.9 [1.01-3.55] vs 0.58 [0.46-1.15], p0.01). MAIT-17 correlated with the number of severe exacerbations (r=0.68, p0.001), and correlated negatively with the ACT score (r=-0.55, p=0.01). In summary, MAIT-17 are present in children with asthma and associated with asthma symptoms.

Natural killer (NK) cells are a heterogeneous population of innate lymphocytes whose potent anticancer properties make them ideal candidates for cellular therapeutic application. However, our lack of understanding of the role of NK cell diversity in antitumor responses has hindered advances in this area. In this study, we describe a new CD56(dim) NK cell subset characterized by the lack of expression of DNAX accessory molecule-1 (DNAM-1). Compared with CD56(bright) and CD56(dim)DNAM-1(pos) NK cell subsets, CD56(dim)DNAM-1(neg) NK cells displayed reduced motility, poor proliferation, lower production of interferon-γ, and limited killing capacities. Soluble factors secreted by CD56(dim)DNAM-1(neg) NK cells impaired CD56(dim)DNAM-1(pos) NK cell–mediated killing, indicating a potential inhibitory role for the CD56(dim)DNAM-1(neg) NK cell subset. Transcriptome analysis revealed that CD56(dim)DNAM-1(neg) NK cells constitute a new mature NK cell subset with a specific gene signature. Upon in vitro cytokine stimulation, CD56(dim)DNAM-1(neg) NK cells were found to differentiate from CD56(dim)DNAM-1(pos) NK cells. Finally, we report a dysregulation of NK cell subsets in the blood of patients diagnosed with Hodgkin lymphoma and diffuse large B-cell lymphoma, characterized by decreased CD56(dim)DNAM-1(pos)/CD56(dim)DNAM-1(neg) NK cell ratios and reduced cytotoxic activity of CD56(dim)DNAM-1(pos) NK cells. Altogether, our data offer a better understanding of human peripheral blood NK cell populations and have important clinical implications for the design of NK cell–targeting therapies.

Background Several retrospective case series have suggested rituximab (RTX) might improve patients with refractory Myasthenia Gravis (MG). Objective In this study, we aimed to evaluate prospectively the efficacy of RTX on muscle function in refractory generalized anti-acetylcholine receptor (AChR) MG patients. Methods Enrolled patients received 1 g of RTX at day 0, day 14, and 6-month follow-up (M6). The primary endpoint was improvement of muscle function at 12-month (M12) based on myasthenic muscle score (MMS). Secondary endpoints were an improvement of the MG Foundation of America Postintervention Status (MGFA-PIS), respiratory forced vital capacity, occurrences of acute MG exacerbation and requirement of associated immunosuppressants and immunomodulatory agents. Results Twelve patients were enrolled, and 11 completed the study. Only a single patient presented an improvement of at least 20 points on MMS at M12, although 2 patients displayed an increase of at least 18 points at M12. MGFA-PIS had improved in 55% of patients by M12. The clinical improvement was not associated with a reduction of immunosuppressant burden. Conclusions These results provide data on the effect of RTX in patients with severe, refractory anti-AChR Abs generalized MG. Even though primary outcome was only reached in a single patient at M12, a beneficial effect of RTX on muscle function was seen in half of the patients at M12 and persisted in a third of patients at M18.

A defect in indoleamine 2,3-dioxygenase 1 (IDO1), which is responsible for immunoregulatory tryptophan catabolism, impairs development of immune tolerance to autoantigens in NOD mice, a model for human autoimmune type 1 diabetes (T1D). Whether IDO1 function is also defective in T1D is still unknown. We investigated IDO1 function in sera and peripheral blood mononuclear cells (PBMCs) from children with T1D and matched controls. These children were further included in a discovery study to identify SNPs in IDO1 that might modify the risk of T1D. T1D in children was characterized by a remarkable defect in IDO1 function. A common haplotype, associated with dysfunctional IDO1, increased the risk of developing T1D in the discovery and also confirmation studies. In T1D patients sharing such a common IDO1 haplotype, incubation of PBMCs in vitro with tocilizumab (TCZ) - an IL-6 receptor blocker - would, however, rescue IDO1 activity. In an experimental setting with diabetic NOD mice, TCZ was found to restore normoglycemia via IDO1-dependent mechanisms. Thus, functional SNPs of IDO1 are associated with defective tryptophan catabolism in human T1D, and maneuvers aimed at restoring IDO1 function would be therapeutically effective in at least a subgroup of T1D pediatric patients., The authors wish to thank patients and subjects who participated in this study, as well as nurses and staff of the Pediatric Clinic of S. Maria della Misericordia Hospital (Perugia), Juvenile Diabetes Center-Anna Meyer Children's Hospital (Florence), Unit of Endocrinology and Diabetes-'Bambino Gesu' Children's Hospital (Rome), Hopital Necker-Enfants Malades (Paris), and Diabetes and Metabolism Service-University Hospital Centre of Coimbra (Coimbra). The authors wish also to thank Roberto Gerli for the gift of TCZ, Giovanni Ricci for histologies, and Francisco Carrilho and Eduarda Coutinho for providing and processing, respectively, DNA samples from the Portuguese cohorts. This work was supported by the European Research Council (338954-DIDO to UG) and, in part, by Associazione per l'Aiuto ai Giovani con Diabete Italia e dell'Umbria (to UG) and the Northern Portugal Regional Operational Programme (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (FEDER) (NORTE-01-0145-FEDER-000013 to AC) and the Fundacao para a Ciencia e Tecnologia (contracts IF/00735/2014 to AC, and SFRH/BPD/96176/2013 to CC)., info:eu-repo/semantics/publishedVersion

β-Cells may be a source of IL-1β that is produced as inactive pro-IL-1β and processed into biologically-active IL-1β by enzymatic cleavage mediated by the NLRP1-, NLRP3- and NLRC4-inflammasomes. Little is known about the β-cell inflammasomes. NLRP1-expression was upregulated in islet-cells from T2D-patients and by IL-1β+IFNγ in INS-1 cells in a histone-deacetylase dependent manner. NLRP3 was downregulated by cytokines in INS-1 cells. NLRC4 was barely expressed and not regulated by cytokines. High extracellular K+ reduced cytokine-induced apoptosis and NO production and restored cytokine-inhibited accumulated insulin-secretion. Basal inflammasome expression was JNK1-3 dependent. Knock-down of the ASC interaction domain common for NLRP1 and 3 improved insulin secretion and ameliorated IL-1β and/or glucolipotoxicity-induced cell death and reduced cytokine-induced NO-production. Broad inflammasome-inhibition, but not NLRP3-selective inhibition, protected against IL-1β-induced INS-1 cell-toxicity. We suggest that IL-1β causes β-cell toxicity in part by NLRP1 mediated caspase-1-activation and maturation of IL-1β leading to an autocrine potentiation loop.

Purpose of review Herein our focus will be to revisit peripheral tolerance mechanisms and in particular 'active' or 'dominant' tolerance as originally defined and mediated by regulatory CD4FoxP3 T lymphocytes (Treg) and also T-cell anergy that appears as a major mainstay to support long-term allograft survival. Recent findings It is at the same time interesting and rewarding that the tool that recently guided our efforts along this path is the in-vivo use of CD3 antibody, the first monoclonal introduced in the clinic (Orthoclone OKT3) about 35 years ago to treat and prevent rejection of renal allografts. Beyond their immunosuppressive activity, whenever administered judiciously, CD3 antibodies promote robust allograft tolerance through selective purging of alloreactive effectors, resetting Treg-mediated active tolerance and promoting a unique subset of anergic CD8 T cells. Summary The new findings discussed open up new perspectives from both a fundamental and a clinical point of view. In basic research, concrete molecular signaling paths are now spotted to finely dissect the conditions that lead to the establishment and maintenance of robust T-lymphocyte anergy mediating allograft tolerance. In the clinic, this may rapidly translate into novel biomarkers to be used in parallel to the ones already available, to better adapt posttransplant immunotherapy and monitor for long-term allograft acceptance.

The field of immunotherapy has undergone a major rejuvenation with the development of monoclonal antibodies and fusion proteins targeting specialized receptors of T and B lymphocytes or cytokines relevant for the differentiation of these cells. This review will focus on autoimmunity, probably the field that benefited most from these new biological therapies that very significantly impacted the modalities of patient's care. The aim is to present the agents which constitute major disease modifying drugs for the treatment of chronic invalidating autoimmune diseases. In doing so, we shall distinguish between agents that globally depress immune responses and those that may selectively target the harmful autoimmune response over long-term while preserving the capacity of the host to react normally to exogenous antigens, in other words, agents promoting 'operational tolerance'.

Eosinophilic esophagitis (EoE) is a clinicopathologic disease that presents with a massive infiltration of the esophagus by eosinophils triggered by food antigen(s).To determine the impact of dietary therapy on nutritional parameters in patients who present with EoE.A convenience retrospective study analyzed patients with EoE after a 2-month dietary therapy (6-food elimination diet, avoidance of the 6 most common allergenic foods, plus avoidance of those eliciting positive skin testing, plus amino-acid formula as replacement for dairy products). Pre- and postdiet allergic and nutritional status were evaluated.Of 111 eligible patients, 59 patients, with a median age of 77.7 months (range, 9-189 months) were enrolled. Dietary therapy significantly increased the return to normal endoscopic appearance (47.4%, P.0009) and led to complete remission (5 eosinophils/esophageal HPF and disappearance of symptoms) in 59.3%. All symptoms improved, digestive (98.3%), cutaneous (80%), and respiratory (92.8%). The prediet median weight-for-height (WFH) z score was -0.75 (-3.00 to 5.69), and the postdiet WFH did not significantly differ, -0.51 (-3.09 to 5.00). The prediet WFH z score was less than -2 (moderate malnutrition) in 10.1%. Postdiet blood eosinophils counts decreased in absolute numbers and in counts ≥ 500 × 10(6)/L (P.0001). Evaluation after 1 year of progressive reintroduction of eliminated foods was available in 33 children: the median WFH z score did not significantly improve, from -0.89 (range, -3.00 to 0.67) at enrollment to -0.59 (range, -3.66 to 2.24).The nutritional status of children with EoE was mildly affected and not worsened by the 2-month dietary therapy.

Background Cow's milk allergy (CMA) is a frequent food allergy in young children. The oral food challenge is the gold standard for diagnosis, and there is currently no reliable biological test. Our aim was to evaluate the diagnostic potential of a functional assay quantifying allergen-specific Th2 cells in CMA children. Methods A total of 29 children aged 2.8–10.5 years underwent a double-blind, placebo-controlled food challenge (DBPCFC) to cow's milk. Blood was collected before performing the DBPCFC, and peripheral mononuclear cells were cultured in an 18-h ELISpot assay with casein, α-lactalbumin, or β-lactoglobulin. Numbers of antigen-specific IL-4- and IL-13-secreting lymphocytes and serum-specific IgE, IgG4, and total IgE levels were assessed. Receiver operating characteristic (ROC) curves were generated. Results A total of 17 (59%) children reacted to cow's milk and were therefore considered as allergic to cow's milk (CMA). The mean number of casein-specific IL-4- and IL-13-secreting T cells was higher in CMA than in non-CMA children (P = 0.009, 0.004, respectively). Moreover, it was inversely correlated with the cumulative dose of cow's milk tolerated (P = 0.003, 0.0009, respectively). ROC curve of combined IL-4 and IL-13 analysis showed an area under the curve of 0.98 (95% CI 0.90–1.06). For a cutoff of 10 IL-4- and 12 IL-13-secreting T cells, sensitivity and negative predictive value were 100%. Conclusions Enumeration of casein-specific IL-4- and IL-13-secreting T cells appears a promising tool to improve diagnosis and, if confirmed in larger studies, could permit less frequent use of the oral food challenge.

SummaryBackground Defining suitable markers to diagnose and monitor allergy and its severity is essential to correctly assign patients for specific immunotherapy. Circulating levels of specific IgE are good markers of sensitization, but not of clinically symptomatic allergy. Objective To quantify circulating interleukin (IL)-4- and IL-13-secreting T cells specific for house dust mite (HDM) in children presenting HDM-allergic asthma associated or not with rhinitis and correlate results with clinical symptoms. Methods We analysed 26 children with HDM respiratory disease (allergic rhinitis and asthma) together with six children with non-allergic asthma. Peripheral blood mononuclear cells were stimulated with HDM extract in a 24-h ELISpot assay to quantify the number of HDM-specific IL-4- and IL-13-secreting T cells. Asthma severity and control, and rhinitis severity were scored according to the Global Initiative for Asthma (GINA) and the Allergic Rhinitis and its Impact on Asthma (ARIA) Guidelines. Results The number of HDM-specific IL-4- and IL-13-secreting T cells was higher in patients with allergic asthma as compared to patients with non-allergic asthma. It varied with the season of blood sampling with two peaks in the fall and early spring. Independently of the season, the number of HDM-specific IL-4-secreting T cells correlated with rhinitis severity (OR = 2; 95% IC:1.1–3.8; P = 0.04). Conclusions and Clinical relevance Allergen-specific IL-4- and IL-13-producing T cells were only detected in HDM-allergic asthmatic children (not in patients with non-allergic asthma). Their numbers correlated with clinical severity of allergic rhinitis.

Assess the frequency of anti-HAnti-HAnti-HAutoimmune gastritis presents an age-dependent frequency in children and adolescents with T1D but is rarely symptomatic. Screening for anti-H

Ideally, therapy for autoimmune diseases should eliminate pathogenic autoimmune cells while sparing protective immunity, but feasible strategies for such an approach have been elusive. Here, we show that in the antibody-mediated autoimmune disease pemphigus vulgaris (PV), autoantigen-based chimeric immunoreceptors can direct T cells to kill autoreactive B lymphocytes through the specificity of the B cell receptor (BCR). We engineered human T cells to express a chimeric autoantibody receptor (CAAR), consisting of the PV autoantigen, desmoglein (Dsg) 3, fused to CD137-CD3ζ signaling domains. Dsg3 CAAR-T cells exhibit specific cytotoxicity against cells expressing anti-Dsg3 BCRs in vitro and expand, persist, and specifically eliminate Dsg3-specific B cells in vivo. CAAR-T cells may provide an effective and universal strategy for specific targeting of autoreactive B cells in antibody-mediated autoimmune disease.

Cancer onset and progression involves the accumulation of multiple oncogenic hits, which are thought to dominate or bypass the physiologic regulatory mechanisms in tissue development and homeostasis. We demonstrate in T-cell acute lymphoblastic leukemia (T-ALL) that, irrespective of the complex oncogenic abnormalities underlying tumor progression, experimentally induced, persistent T-cell receptor (TCR) signaling has antileukemic properties and enforces a molecular program resembling thymic negative selection, a major developmental event in normal T-cell development. Using mouse models of T-ALL, we show that induction of TCR signaling by high-affinity self-peptide/MHC or treatment with monoclonal antibodies to the CD3ϵ chain (anti-CD3) causes massive leukemic cell death. Importantly, anti-CD3 treatment hampered leukemogenesis in mice transplanted with either mouse- or patient-derived T-ALLs. These data provide a strong rationale for targeted therapy based on anti-CD3 treatment of patients with TCR-expressing T-ALL and demonstrate that endogenous developmental checkpoint pathways are amenable to therapeutic intervention in cancer cells. Significance: T-ALLs are aggressive malignant lymphoid proliferations of T-cell precursors characterized by high relapse rates and poor prognosis, calling for the search for novel therapeutic options. Here, we report that the lineage-specific TCR/CD3 developmental checkpoint controlling cell death in normal T-cell progenitors remains switchable to induce massive tumor cell apoptosis in T-ALL and is amenable to preclinical therapeutic intervention. Cancer Discov; 6(9); 972–85. ©2016 AACR. See related commentary by Lemonnier and Mak, p. 946. This article is highlighted in the In This Issue feature, p. 932

Regulatory macrophages (M regs) were administered to two living-donor renal transplant recipients. Both patients were minimized to low-dose tacrolimus monotherapy within 24 wk of transplantation and subsequently maintained excellent graft function. After central venous administration, most M regs remained viable and were seen to traffic from the pulmonary vasculature via the blood to liver, spleen, and bone marrow. By 1 y posttransplantation, both patients displayed patterns of peripheral blood gene expression converging upon the IOT-RISET signature. Furthermore, both patients maintained levels of peripheral blood FOXP3 and TOAG-1 mRNA expression within the range consistent with nonrejection. It is concluded that M regs warrant further study as a potential immune-conditioning therapy for use in solid-organ transplantation. The results of this work are being used to inform the design of The ONE Study, a multinational clinical trial of immunomodulatory cell therapy in renal transplantation. Copyright © 2011 by The American Association of Immunologists, Inc.

The ability to induce specific tolerance during transplantation or autoimmunity has considerable clinical applications. Fundamental and clinical immunologists gathered at a recent meeting 2 2The meeting ‘Tolerance Induction in the Adult: Means and Mechanisms' was held in Le Bischenberg, Strasbourg, 3–6 September 1994, and was sponsored by the European Science Foundation. to discuss mechanisms underlying the induction and maintenance of tolerance in mature T cells.

AIMS/HYPOTHESIS: The aim of the study was to examine the 48 month outcome of treating recent-onset type 1 diabetic patients for 6 days with humanised CD3-antibody, ChAglyCD3. METHODS: Eighty patients, aged 12-39 years, were recruited for a phase 2 multicentre trial and randomised to placebo (n=40) or ChAglyCD3 (n=40) treatment by a third party member; participants and care-givers were blinded. The change in insulin dose (U kg(-1)day(-1)) over 48 months was chosen as primary endpoint and compared in 31 placebo-and 33 ChAglyCD3-treated patients. Adverse events were followed in 35 and 38 patients, respectively. RESULTS: Treatment with ChAglyCD3 delayed the rise in insulin requirements of patients with recent-onset diabetes and reduced its amplitude over 48 months (+0.09 vs +0.32 U kg(-1)day(-1) in the placebo group). Using multivariate analysis this effect was correlated with higher baseline residual beta cell function and a younger age. It was associated with better outcome variables in subgroups selected according to these variables. In the ChAglyCD3 subgroup with higher initial beta cell function, 0/11 patients became C-peptide-negative over 48 months vs 4/9 in the corresponding placebo subgroup. In the subgroup aged 80% decline within 24 months in the placebo subgroup

In this brief review we propose to discuss salient data showing the importance of immune regulatory mechanisms, and in particular of Treg, for the control of pathogenic anti-β-cell response in autoimmune diabetes. Disease progression that culminates with the massive destruction of insulin-secreting β-cells and advent of hyperglycemia and glycosuria tightly correlates with a functional deficit in immune regulation. Better dissection of the cellular and molecular mechanisms through which the immune system normally sustains tolerance to "self", and which become defective when autoimmune aggression is overt, is the only direct and robust way to learn how to harness these effectively, so as to restore immune tolerance in patients with insulin-dependent type 1 diabetes. No doubt that regulatory T cells are a privileged mechanism underlying this self-tolerance in the periphery. The discovery of the key role of the transcription factor FoxP3, represented the cornerstone leading to the great advances in the field we are witnessing today. Type 1 diabetes is certainly one of the prototypic T cell-mediated autoimmune diseases where immune regulatory mechanisms relying on specialized subsets of T cells have been the most thoroughly analyzed from the fundamental point of view and also largely exploited in a translational therapeutic perspective.

BACKGROUND Persistent CD4 T-cell lymphopenia after kidney transplantation has been associated with an increased occurrence of opportunistic infections, malignancies and even mortality, but studies have focussed only on the first few years after kidney transplantation. In this study, we investigated the risk factors and clinical significance of long-term profound CD4 lymphopenia detected ≥10 years after renal transplantation. METHODS Between 2007 and 2010, 6206 CD4 T-cell counts, including 1507 counts

Type 1 diabetes results from the destruction of β-cells by an autoimmune T-cell response assisted by antigen-presenting B cells producing autoantibodies. CD8+ T-cell responses against islet cell antigens, thought to play a central role in diabetes pathogenesis, can be monitored using enzyme-linked immunosorbent spot (ELISpot) assays. However, such assays have been applied to monitoring of adult patients only, leaving aside the large and increasing pediatric patient population. The objective of this study was twofold: 1) to develop a CD8+ T-cell interferon-γ ELISpot assay for pediatric patients and 2) to determine whether zinc transporter 8 (ZnT8), a recently described target of autoantibodies in a majority of patients, is also recognized by autoreactive CD8+ T cells. Using DNA immunization of humanized mice, we identified nine HLA-A2–restricted ZnT8 epitopes. Among 36 HLA-A2+ children with diabetes, 29 responded to ZnT8 epitopes, whereas only 3 of 16 HLA-A2+ control patients and 0 of 17 HLA-A2− control patients responded. Some single ZnT8 epitopes performed as well as the group of epitopes in discriminating between patients and control individuals. Thus, ZnT8 is a major CD8+ T-cell autoantigen, and ELISpot assays display similar performance in adult and pediatric type 1 diabetes.

OBJECTIVES The diagnosis of celiac disease (CD) is based on the histological identification of gluten-sensitive enteropathy and detection of anti-tissue transglutaminase antibodies (tTGA) and/or endomysial antibodies. Serial measurements of tTGA are now recommended as a follow-up strategy to monitor compliance with a gluten-free diet (GFD). We evaluated the performances of a quantitative radiobinding assay (RBA) of tTGA immunoglobulin A at diagnosis and during monitoring of GFD in pediatric CD. METHODS Eighty children with confirmed CD were selected. Levels of serum tTGA measured by RBA and a commercial enzyme-linked immunosorbent assay (ELISA) were compared at diagnosis. The relation between RBA-tTGA levels and histological damage was analyzed, as well as the time course of tTGA clearance during GFD. RESULTS Both RBA and ELISA showed high sensitivity and specificity for tTGA detection at diagnosis. There was no relation between RBA-tTGA levels at diagnosis and severity of mucosal damage. Upon initiation of GFD, the rate of RBA-tTGA positivity declined slower than that of endomysial antibodies positivity, with >50% of the children still tTGA positive at year 5; however, tTGA levels decreased rapidly during the first year of GFD and more slowly thereafter. Children who seroreverted had lower tTGA levels at diagnosis (2080±1554 cpm) than those who remained tTGA positive throughout follow-up (3688±1435 cpm). CONCLUSIONS The high sensitivity of RBA is likely responsible for higher tTGA positivity rates during GFD than previously reported with ELISA. A decreasing trend for tTGA levels may represent a better surrogate marker of compliance with GFD than absolute normal tTGA levels.

Type 1 diabetes (T1D) is a prototypic organ-specific autoimmune disease resulting from the selective destruction of insulin-secreting β-cells within the pancreatic islets of Langerhans. It is caused by an immune-mediated inflammation, involving autoreactive CD4⁺ and CD8⁺ T lymphocytes that infiltrate the islets and initiate insulitis. The use of exogenous insulin is the current standard treatment. However, in spite of significant advances, this therapy is still associated with major constraints, including risk of hypoglycemia and severe degenerative complications. As T1D mainly affects children and young adults, any candidate immune therapy must be safe, and it must avoid a sustained depression of immune responses with all its attendant problems of recurrent infection and drug toxicity. In this context, inducing or restoring immune tolerance to target autoantigens would be the ideal approach. We refer to immune tolerance here as the selective damping of the damaging autoimmune response following a short treatment, while keeping intact the capacity of the host to respond normally to exogenous antigens. The therapeutic approach we discuss in this article originates from attempts to induce tolerance both to soluble antigens and tissue antigens (i.e. alloantigens and autoantigens) by using biological agents that selectively interfere with lymphocyte activation, namely polyclonal and monoclonal anti-T cell antibodies. The challenged dogma was that, in an adult-primed immune system, it was not possible to restore self-tolerance therapeutically without the use of exogenous autoantigen administration. The reality has been that, in diabetes, endogenous host autoantigen can fulfill this role because a significant amount of functioning β-cells remains, even at the time of established hyperglycemia. Experimental results obtained in the 1990s showed that a short-term CD3 antibody treatment in recently diagnosed diabetic non-obese diabetic (NOD) mice induced permanent remission of the disease by restoring self-tolerance. Based on these findings, phase I, II, and III trials were conducted using two distinct humanized Fc-mutated antibodies to human CD3, namely ChAglyCD3 (otelixizumab) and OKT3γ1 Ala-Ala (teplizumab). Overall, when dosing was adequate, the results demonstrated that CD3 antibodies preserved β-cell function very efficiently, maintaining significantly high levels of endogenous insulin secretion in treated patients for up to 24 months after treatment. These data provided the first proof of concept for a long-term therapeutic effect in T1D following a short course administration of a therapeutic agent. Our aim is to review these data and to discuss them in the context of the pitfalls linked to pharmaceutical development, especially in the context of pediatric patients, as in autoimmune diabetes.

Type 1 diabetes is an autoimmune disease. It was thus logical to attempt preventing or stopping the progression of the disease by immunotherapy. Following the strategies used in organ transplantation, the first trials in the 80s used cyclosporin in patients presenting recently diagnosed Type 1 diabetes. The effect was spectacular but waned when the treatment was stopped as the effect was non antigen-specific. Going back from bed to bench-side major efforts were then devoted to device strategies allowing induction or restoration of self-tolerance. Two major approaches provided encouraging results when used in spontaneous models of autoimmune diabetes that are the use of β-cell autoantigens and of monoclonal antibodies to CD3. Based on these results academic phase II trials and subsequently pharmaceutically driven phase III trials were launched. Results are now available and when critically analyzed in the frame of these last three decades they provide support to the possibility of making step by step immunotherapy available to all new onset diabetic patients with a hope of inducing long-term remission of the disease if the treatment is started sufficiently early, immediately after diagnosis.

Purpose of review The continuing problem of late graft loss and immunosuppressive drug toxicity forces us to explore new treatments for the induction of transplant tolerance. Monoclonal antibodies targeting molecules implicated in lymphocyte activation, in particular CD3/TCR, constitute a promising strategy. Recent findings Promising results were obtained from the use of antibodies targeting CD3/TCR, coreceptors or costimulatory pathways as tolerance-promoting tools in experimental transplantation. These antibodies do not uniformly depress the immune system but act in an antigen-specific manner by preferentially targeting effector T cells while preserving regulatory T cells. However, translation to the clinic proved to be more difficult than expected. New generation CD3 antibodies, currently used in phase II/III trials in autoimmunity, constitute a promising approach as, beside their immunosuppressive effect, they also express potent tolerogenic capacities. Importantly, CD3 therapy is effective especially when applied in primed hosts, highlighting the importance of the therapeutic window for tolerance induction. Summary Further investigations are required for adapting to the clinic monoclonal antibodies as substitutes for current immunosuppression. Our aim is to show that development of new therapeutic strategies/molecules may come from transversal-type research, in particular from experience in autoimmunity, as immune responses leading to autoimmunity and graft rejection involve similar pathways.

BACKGROUND FOXP3-expressing regulatory T cells (Tregs) play a crucial role in maintaining allogeneic transplant tolerance in experimental models. In clinical transplantation, there are few data about their role in chronic inflammation. We hypothesized that Tregs might accumulate within the graft since enrichment of Tregs has been frequently described in chronically inflamed tissues. METHODS Sixty-seven biopsies, indicated by a rise in creatinine level, were studied. Thirty-four biopsies showing acute T-cell-mediated rejection and 33 displaying inflamed fibrosis were selected. Tregs frequency was calculated for each infiltrate by counting FOXP3+ and CD3+ cells on two contiguous serial sections. RESULTS A total of 121 infiltrates were scored with a mean of 309 CD3+ cells per infiltrate (range: 50-700). Tregs were enriched within allografts exhibiting inflamed fibrosis versus acute cellular rejection (10.6 +/- 6.8% versus 5.5 +/- 2.6%, respectively, P = 0.005). In those with inflammation within scarred areas, the subset of patients with a low FOXP3/CD3 ratio (below the median value) displayed a lower frequency of B-cell-enriched nodular cell clusters (20% versus 86%, P = 0.001) and had a significantly lower graft survival (log-rank, P = 0.02). In multivariate analysis, the low FOXP3/CD3 ratio remained an independent indicator of outcome (P = 0.03). Consistently, the FOXP3+/IL-17+ cell ratio was higher in nodular than in diffuse infiltrates. CONCLUSIONS Our results suggest that Tregs may dampen the graft injury in chronic (versus acute) inflammation and stress the importance of devising strategies to enhance Tregs efficiency.

Aim Infliximab (IFX) therapy is highly efficacious for the induction and maintenance of remission in pediatric Crohn disease (CD). However, to date it is unclear how long patients should be given IFX. Given the increasing safety concerns about the concomitant and prolonged use of IFX and azathioprine in CD, we wanted to address the clinical outcome in pediatric CD patients who responded to IFX medication, once IFX was stopped. Patients and methods Upon induction therapy with 3 IFX infusions, 36 of 38 patients with CD were in clinical remission at 3 months. These 36 responders were separated into 2 groups: 16 patients received no further IFX infusions, whereas 20 patients received scheduled maintenance therapy with IFX for 12 months. Results Among the 16 patients who received no further IFX infusions, 12 experienced relapse during the 12-month follow-up interval after IFX was stopped. In the group receiving maintenance therapy, 11 of 20 patients remained in clinical remission at 12 months of therapy, whereas 8 patients required adjustment of IFX doses or intervals. Among the 11 children who were in clinical remission and receiving maintenance therapy without dose adjustment, 8 experienced relapse within 12 months after IFX maintenance therapy was stopped. Overall, the relapse rates after IFX induction or maintenance therapy was stopped were 75% and 72%, respectively. Conclusions These data indicate that IFX is efficacious in controlling severe pediatric CD; however, to induce and maintain clinical remission, repeated IFX infusions are required, with a need for dose adjustment in a substantial number of patients.

Type 1 diabetes (T1D) results from a chronic and selective destruction of insulin-secreting β-cells within the islets of Langerhans of the pancreas by autoreactive CD4(+) and CD8(+) T lymphocytes. The use of animal models of T1D was instrumental for deciphering the steps of the autoimmune process leading to T1D. The non-obese diabetic (NOD) mouse and the bio-breeding (BB) rat spontaneously develop the disease similar to the human pathology in terms of the immune responses triggering autoimmune diabetes and of the genetic and environmental factors influencing disease susceptibility. The generation of genetically modified models allowed refining our understanding of the etiology and the pathogenesis of the disease. In the present review, we provide an overview of the experimental models generated and used to gain knowledge on the molecular and cellular mechanisms underlying the breakdown of self-tolerance in T1D and the progression of the autoimmune response. Immunotherapeutic interventions designed in these animal models and translated into the clinical arena in T1D patients will also be discussed.

International audience; The contribution of memory B cells in alloreactive humoralresponses remains poorly understood. Here we testedthe presence of circulating alloreactive memory B cells in69 patients with end-stage renal disease under renalreplacement therapy, using an in vitro memory Bcell–stimulation assay combined with identification of IgGhuman leukocyte antigen (HLA) antibodies in culturesupernatant. HLA antibody–producing memory B cells wereevidenced only in patients carrying serum HLA antibodiesfollowing multiple classical HLA-immunizing events. Inpatients with a previous renal allograft, alloreactive memoryB cells could be detected ranging from 6 to 32 years (mean13.2 years) after transplantation. HLA antibodies produced bymemory B cells were also detected in the corresponding seraand showed a restricted reactivity, targeting only a fewepitopes shared by several HLA antigens. In contrast, serumHLA antibodies, not associated with the detection of specificmemory B cells, showed a broader pattern of specificities.Thus, expansion and survival of alloreactive memory B cells isalloantigen driven, and their frequency is related to the‘strength’ of HLA immunization.

Heart failure is still a major cause of hospitalization and mortality in developed countries. Many clinical trials have tested the use of multipotent stem cells as a cardiac regenerative medicine. The benefit for the patients of this therapeutic intervention has remained limited. Herein, we review the pluripotent stem cells as a cell source for cardiac regeneration. We more specifically address the various challenges of this cell therapy approach. We question the cell delivery systems, the immune tolerance of allogenic cells, the potential proarrhythmic effects, various drug mediated interventions to facilitate cell grafting and, finally, we describe the pathological conditions that may benefit from such an innovative approach. As members of a transatlantic consortium of excellence of basic science researchers and clinicians, we propose some guidelines to be applied to cell types and modes of delivery in order to translate pluripotent stem cell cardiac derivatives into safe and effective clinical trials.

Tumor necrosis factor (TNF)-alpha plays a critical role in the initiation and progression of Crohn's disease, a chronic inflammatory disorder of the gastrointestinal tract. Infliximab, a chimeric monoclonal antibody blocking TNF-alpha, has proven effective as an induction and maintenance therapy for refractory Crohn's disease in adult and pediatric patients. However, infliximab therapy induces the appearance of neutralizing anti-infliximab antibodies. In the pediatric cohort, we analyzed (n=28) sensitization occurred in 35.7% patients and was associated with a loss of response to maintenance infusions. In two patients presenting high titers of anti-infliximab antibodies, severe infusion reactions were observed, possibly IgE-mediated, precluding further use of the medication. Serum concentrations of TNF-alpha and infliximab were influenced by the presence of anti-infliximab antibodies. We propose that surveillance of circulating infliximab and/or TNF-alpha concentration during maintenance therapy represents an indirect but reliable method to monitor anti-infliximab immunization.

Background The responsibility of sirolimus (SRL) for postrenal transplant anemia has never been proven, because SRL is usually combined with myelotoxic drugs, and because of the high incidence of anemia in the posttransplant period. Methods We retrospectively analyzed anemia in 46 renal transplant recipients, who had been switched from calcineurin inhibitors to SRL for biopsy-proven chronic allograft nephropathy. Results The mean decrease in hemoglobin (Hb) after SRL introduction was 2.8 g/dl. The 24 patients, whose Hb fell by >or=2 g/dl, displayed microcytic aregenerative anemia with low serum iron despite high ferritinemia, consistent with anemia of chronic inflammatory states. Fibrinogen and CRP levels increased in these patients after sirolimus introduction. We subsequently focused our study on eight patients without confounding factors of anemia. Anemia improved in all eight after SRL withdrawal. IL6 and TNFalpha at the nadir of anemia were significantly higher than before SRL introduction and after its withdrawal. Decreases in Hb correlated with increases in proinflammatory cytokine levels in a linear regression model. Unchanged serum IL10 levels measured at the nadir of anemia were discordant with the inflammatory state. Conclusions Late introduction of SRL may induce anemia and correlates with biochemical evidence of a chronic inflammatory state possibly due to defective IL10-dependent inflammatory autoregulation.

Current therapeutic approaches in transplantation and autoimmunity are essentially focused on immunosuppression, which is non-specific (i.e. unrelated to the antigens involved). The major drawback is their relative ineffectiveness in the long term, with the likely risk of recurrence of the pathogenic immune process once the drug is withdrawn necessitating indefinite drug administration; this has attendant problems of recurrent infections and drug toxicity. Instead, CD3-specific monoclonal antibodies possess the unique capacity to induce immunological tolerance: an antigen-specific unresponsiveness in the absence of long-term generalised immunosuppression, as is well-established in experimental models. Clinical application using humanised non-mitogenic CD3-specific antibodies is presently underway. The future challenge will be to define the modalities allowing the widespread application of this strategy through a better understanding of the underlying immune mechanisms.

Although they were used initially as non-specific immunosuppressants in transplantation, CD3-specific monoclonal antibodies have elicited renewed interest owing to their capacity to induce immune tolerance. In mouse models of autoimmune diabetes, CD3-specific antibodies induce stable disease remission by restoring tolerance to pancreatic beta-cells. This phenomenon was extended recently to the clinic--preservation of beta-cell function in recently diagnosed patients with diabetes was achieved by short-term administration of a CD3-specific antibody. CD3-specific antibodies arrest ongoing disease by rapidly clearing pathogenic T cells from the target. Subsequently, they promote long-term T-cell-mediated active tolerance. Recent data indicate that transforming growth factor-beta-dependent CD4+CD25+ regulatory T cells might have a central role in this effect.

Major progress has been made over the last 10 years in understanding the immune mechanisms underlying autoimmune Type I (insulin-dependent) diabetes mellitus. It seems that immunotherapy is the best approach for preventing and/or arresting beta cell destruction. For immunotherapeutic approaches to be clinically applicable to Type I diabetes, they cannot rely on the use of non-specific immunosuppressants. Their relative ineffectiveness over the long term and the risk of recurrence, once the drug is withdrawn, requires permanent drug treatment which augments problems of infection and drug toxicity. One solution is to selectively tackle the pathogenic immune response. Operationally, this requires establishing immunological tolerance, i. e. a state of durable antigen-specific unresponsiveness in the absence of generalised immunosuppression. This review aims to present the rationale and clinical applicability of possible immunointervention strategies in Type I diabetes aimed at restoring self-tolerance to beta cells. [Diabetologia (2001) 44: 521–536]

Tolerance to β cell autoantigens represents a fragile equilibrium. Autoreactive T cells specific to these autoantigens are present in most normal individuals but are kept under control by a number of peripheral tolerance mechanisms, among which CD4+CD25+CD62L+T cell–mediated regulation probably plays a central role. The equilibrium may be disrupted by inappropriate activation of autoantigen-specific T cells, notably following to local inflammation that enhances the expression of the various molecules contributing to antigen recognition by T cells. Even when T cell activation finally overrides regulation, stimulation of regulatory cells by CD3 antibodies may reset the control of autoimmunity. Other procedures may also lead to disease prevention. These procedures are essentially focused on Th2 cytokines, whether used systemically or produced by Th2 cells after specific stimulation by autoantigens. Protection can also be obtained by NK T cell stimulation. Administration of β cell antigens or CD3 antibodies is now being tested in clinical trials in prediabetics and/or recently diagnosed diabetes.

The immune mechanisms underlying autoimmune diseases and allograft rejection are similar, and T cells play a major role in both processes. Current therapeutic strategies rely on the application of nonspecific immunosuppression using chemicals such as corticosteroids, azathioprine, methotrexate, cyclophosphamide and cyclosporin. One major drawback of these drugs is their relative ineffectiveness over the long term; withdrawal leads to recurrence of the disease, but chronic administration puts patients at risk of being overimmunosuppressed, which can result in an increased incidence of infections and tumors. This review describes the advantages of developing biological agents that target immune receptors on T cells. In some cases, these agents can induce a state of durable, antigen-specific unresponsiveness in the absence of generalized immunosuppression, which could be useful in transplantation and autoimmunity.

A long-held dogma was that immune tolerance could only be achieved through central deletion of potentially reactive cells at their site of origin, a concept supported by the direct demonstration of natural negative selection in bone marrow and thymus; however, it is now extensively documented that although deletion mechanisms ensure the elimination of most autoreactive cells, this purging is far from complete. There are major immune mechanisms that ensure tolerance in the periphery which are clearly non-deletional. A paper in this issue of the European Journal of Immunology demonstrates that T cell-mediated immunoregulation involving in particular the CTLA-4 signaling pathway plays a key role in controlling the pathogenic potential of fully differentiated autoreactive T cells.

It is now well established that a distinct subset of T lymphocytes is essential for downregulating immune responses to both endogenous (self) and exogenous antigens. These Tregs are CD4+ and express high levels of CD25 (the alpha chain of the IL-2 receptor) and the transcription factor Foxp3. The mechanisms determining the lifespan, homeostasis, and in vivo generation of these Tregs are still ill defined. A study by Vukmanovic-Stejic et al. in this issue of the JCI shows that in humans, Tregs are present throughout life but that despite their high throughput, they are short lived (see the related article beginning on page 2423). It is thus unlikely that all CD4+ CD25hi Foxp3+ Tregs are generated as a separate lineage in the thymus. The authors propose that during adulthood, Tregs essentially emerge at the periphery from the memory T cell pool.

The detection of preformed donor-specific alloantibodies (DSA) with multiplex-bead arrays has led to the common observation that individuals without a history of pregnancy, transfusion or transplantation can have circulating anti-HLA antibodies of unknown etiology. We retrospectively analyzed the risk of antibody-mediated rejection (AMR) and graft outcome in 41 kidney transplant recipients with DSA of unknown etiology (DSA cause-unk) at the time of transplantation. Twenty-one patients received a posttransplantation desensitization protocol, and 20 received standard immunosuppressive therapy. The mean number of DSA was 1.4 ± 0.8, ranging from 1 to 5. Complement-dependent cytotoxicity crossmatches were negative for all the patients. Flow cytometry crossmatches were positive in 47.6% of cases. The incidence of acute AMR was 14.6% at 1 year, regardless of the immunosuppressive regimen. No patients experienced graft loss following AMR. At month 12, across the entire population of patients with DSA cause-unk, the outcomes were favorable: the measured glomerular filtration rate was 63.8 ± 16.4 mL/min/1.73 m(2), the screening biopsies showed low frequencies of microvascular inflammation and no transplant glomerulopathy, and graft and patient survival were 100%. In conclusion, patients with DSA cause-unk are able to mount AMR but have favorable 1-year outcomes.

Two humanized, anti-CD3 mAbs with reduced FcR binding, teplizumab and otelixizumab, have been evaluated in over 1500 subjects, ages 7-45, with new and recently diagnosed T1D with a range of intravenous doses (3-48mg) and regimens (6-14 days, single or repeat courses). In general, studies that used adequate dosing demonstrated improvement in stimulated C-peptide responses and reduced need for exogenous insulin for two years and even longer after diagnosis. Drug treatment causes a transient reduction in circulating T cells, but the available data suggest that the mechanism of action may involve induction of regulatory mechanisms. The adverse effects of anti-CD3 treatment are infusion-related and transient. The studies have identified significant differences in efficacy among patient groups suggesting that a key aspect for development of this immune therapy is identification of the demographic, metabolic, and immunologic features that distinguish subjects who are most likely to show beneficial clinical responses.