Implication de la protéine du syndrome de Wiskott-Aldrich dans la migration et la rencontre des lymphocytes T CD4+ avec les cellules présentatrices de l'antigène

Les lymphocytes T CD4+ sont des acteurs clés dans l'orchestration d'une réponse immunitaire dirigée spécifiquement contre un pathogène. L'efficacité de la réponse immunitaire dépend de la génération de lymphocytes T CD4+, de l'acquisition de leurs fonctions effectrices et de l'exercice de ces dernières. Ces différentes étapes se déroulent successivement dans différents sites spécialisés de l'organisme. Ceci impose aux lymphocytes T CD4+ une grande mobilité ainsi que l'établissement de contacts privilégiés avec les cellules présentatrices de l'antigène, responsables de leur différenciation et de leur activation. La mobilité ainsi que les interactions cellulaires des lymphocytes T CD4+ requièrent une réorganisation continue de leur cytosquelette d'actine. Un des régulateurs majeurs du cytosquelette d'actine des lymphocytes T est la protéine du syndrome de Wiskott-Aldrich (WASP) dont la déficience provoque le syndrome de Wiskott-Aldrich (WAS), une immunodéficience primaire sévère. Le dysfonctionnement des lymphocytes T est généralement considéré comme la cause majeure du dérèglement du système immunitaire observé chez les patients WAS. Mes travaux de thèse ont consisté à étudier l'implication de WASP dans la mobilité des lymphocytes T CD4+ humains, au cours des phases de migration et de rencontre avec les cellules présentatrices de l'antigène dans les contextes d'interaction non spécifique (antigènes du soi) et spécifique de l'antigène (antigènes exogènes). Nous avons tout d'abord montré que WASP n'est pas requit pour la migration dirigée par les chimiokines des lymphocytes T CD4+ in vitro. Ensuite, nous nous sommes intéressés au rôle de WASP dans l'interaction des lymphocytes T CD4+ avec des cellules présentatrices de l'antigène présentant des antigènes du soi. Dans cette situation, il a été précédemment décrit que les cellules présentatrices de l'antigène induisaient la mobilité des lymphocytes T et participaient au maintien de l'homéostasie lymphocytaire. De façon surprenante, nous avons observé que l'absence de WASP conduisait à une mobilité anormalement élevée des lymphocytes T CD4+ au contact des cellules présentatrices de l'antigène. Nous avons ensuite montré que cet excès de mobilité était lié à une adhésion LFA-1/ICAM-1 anormalement régulée. En conséquence de cet excès de mobilité, nous avons également observé un maintien favorisé de la survie homéostatique lymphocytaire. Enfin, nous avons poursuivi cette étude dans un contexte d'interaction avec des cellules présentatrices de l'antigène présentant des antigènes exogènes. Dans cette situation, il est connu que l'engagement du TCR avec les antigènes exogènes induit un arrêt de mobilité des lymphocytes T. Nous avons observé qu'en absence de WASP, les lymphocytes T CD4+ présentaient des défauts d'arrêt de migration sur les cellules présentatrices de l'antigène. Nous avons aussi montré un défaut d'organisation moléculaire de la synapse immunologique, formée entre un lymphocyte T et une cellule présentatrice de l'antigène. En conséquence de cette mauvaise organisation, des défauts de prolifération et de production de cytokines des lymphocytes T ont été révélés. Finalement, nos résultats indiquent que WASP agirait comme un frein sur la mobilité des lymphocytes T CD4+ au contact des cellules présentatrices de l'antigène pour permettre une interaction cellulaire optimale dans le contexte de la rencontre avec le soi et le non-soi. Nous proposons que ces défauts soient en partie responsables des symptômes auto-immuns et de l'immunodéficience observés chez les patients WAS.CD4+ T cells orchestrate immune response directed specifically against pathogens. Immune response efficiency depends on the maturation of CD4+ T lymphocytes and on their ability to exert effectors functions. These steps occur successively in different sites of the organism in which T cells will interact with multiple antigen presenting cells. Tissue homing and antigen presenting cell scanning require a very high and regulated migratory capacity. This is in part controlled by actin cytoskeleton reorganization. A key regulator of actin cytoskeleton in T lymphocyte is the Wiskott-Aldrich syndrome protein (WASP) which deficiency in humans causes the Wiskott-Aldrich syndrome (WAS), a severe primary immunodeficiency. T cell dysfunction is contributing to the immune system dysregulation observed in WAS patients. My thesis research consisted in studying the role of WASP in human CD4+ T lymphocyte motility, in real time, during their migration phases and during antigen presenting cell encounter in the context of both non specific (self antigens) and specific interactions (exogenous antigens). First, we demonstrated that WASP is not required for CD4+ T lymphocyte chemotaxis in vitro. Then, we investigated the role of WASP in CD4+ T lymphocyte interaction with antigen presenting cells presenting self-antigens. In this context, it was previously described that antigen-presenting cells drive T cell motility and maintain T cell homeostasis. Surprisingly, we observed that the absence of WASP lead to an abnormally high CD4+ T cell motility at the contact with antigen presenting cells. Then, we showed that this increased motility is linked to an abnormal regulation of LFA-1/ICAM-1 adhesion. Consequently of this exacerbated motility, we also observed that T cell homeostatic survival is favored. Finally, we pursued our study in the context of exogenous antigen encounter. In this condition, it is known that TCR engagement with exogenous antigens drive T cell migration arrest. We observed that in the absence of WASP, CD4+ T lymphocytes are unable to stop efficiently at the contact with antigen presenting cells. We also demonstrated that signaling molecules at the immunological synapse established between T lymphocytes and antigen presenting cells are dispersed in the absence of WASP. Consequently to this defective organization, we showed that CD4+ T cell proliferation and cytokine production are impaired. All together, our results indicate that WASP acts as a brake on CD4+ T cell motility at the contact with antigen presenting cells permitting optimal cellular interaction in the context of self and non-self encounter. We hypothezed that these defects could be in part responsible for the autoimmune development and immunodeficiency observed in WAS patients

A Task Force Against Local Inflammation and Cancer: Lymphocyte Trafficking to and Within the Skin

The skin represents a specialized site for immune surveillance consisting of resident, inflammatory and memory populations of lymphocytes. The entry and retention of T cells, B cells, and ILCs is tightly regulated to facilitate detection of pathogens, inflammation and tumors cells. Loss of individual or multiple populations in the skin may break tolerance or increase susceptibility to tumor growth and spread. Studies have significantly advanced our understanding of the role of skin T cells and ILCs at steady state and in inflammatory settings such as viral challenge, atopy, and autoimmune inflammation. The knowledge raised by these studies can benefit to our understanding of immune cell trafficking in primary melanoma, shedding light on the mechanisms of tumor immune surveillance and to improve immunotherapy. This review will focus on the T cells, B cells, and ILCs of the skin at steady state, in inflammatory context and in melanoma. In particular, we will detail the core chemokine and adhesion molecules that regulate cell trafficking to and within the skin, which may provide therapeutic avenues to promote tumor homing for a team of lymphocytes

Kartezio: Evolutionary Design of Explainable Pipelines for Biomedical Image Analysis

An unresolved issue in contemporary biomedicine is the overwhelming number and diversity of complex images that require annotation, analysis and interpretation. Recent advances in Deep Learning have revolutionized the field of computer vision, creating algorithms that compete with human experts in image segmentation tasks. Crucially however, these frameworks require large human-annotated datasets for training and the resulting models are difficult to interpret. In this study, we introduce Kartezio, a modular Cartesian Genetic Programming based computational strategy that generates transparent and easily interpretable image processing pipelines by iteratively assembling and parameterizing computer vision functions. The pipelines thus generated exhibit comparable precision to state-of-the-art Deep Learning approaches on instance segmentation tasks, while requiring drastically smaller training datasets, a feature which confers tremendous flexibility, speed, and functionality to this approach. We also deployed Kartezio to solve semantic and instance segmentation problems in four real-world Use Cases, and showcase its utility in imaging contexts ranging from high-resolution microscopy to clinical pathology. By successfully implementing Kartezio on a portfolio of images ranging from subcellular structures to tumoral tissue, we demonstrated the flexibility, robustness and practical utility of this fully explicable evolutionary designer for semantic and instance segmentation.Comment: 36 pages, 6 main Figures. The Extended Data Movie is available at the following link: https://www.youtube.com/watch?v=r74gdzb6hdA. The source code is available on Github: https://github.com/KevinCortacero/Kartezi

Type I interferon induces CXCL13 to support ectopic germinal center formation.

Ectopic lymphoid structures form in a wide range of inflammatory conditions, including infection, autoimmune disease, and cancer. In the context of infection, this response can be beneficial for the host: influenza A virus infection-induced pulmonary ectopic germinal centers give rise to more broadly cross-reactive antibody responses, thereby generating cross-strain protection. However, despite the ubiquity of ectopic lymphoid structures and their role in both health and disease, little is known about the mechanisms by which inflammation is able to convert a peripheral tissue into one that resembles a secondary lymphoid organ. Here, we show that type I IFN produced after viral infection can induce CXCL13 expression in a phenotypically distinct population of lung fibroblasts, driving CXCR5-dependent recruitment of B cells and initiating ectopic germinal center formation. This identifies type I IFN as a novel inducer of CXCL13, which, in combination with other stimuli, can promote lung remodeling, converting a nonlymphoid tissue into one permissive to functional tertiary lymphoid structure formation

Implication de la protéine du syndrome de Wiskott-Aldrich dans la migration et la rencontre des lymphocytes T CD4+ avec les cellules présentatrices de l'antigène

TOULOUSE3-BU Sciences (315552104) / SudocSudocFranceF

The Wiskott-Aldrich syndrome protein permits assembly of a focused immunological synapse enabling sustained T-cell receptor signaling

International audienceBACKGROUND: T-cell activation relies on the assembly of the immunological synapse, a structure tightly regulated by the actin cytoskeleton. The precise role of the Wiskott-Aldrich syndrome protein, an actin cytoskeleton regulator, in linking immunological synapse structure to downstream signaling remains to be clarified. DESIGN AND METHODS: To address this point, CD4$^+$ T cells from patients with Wiskott-Aldrich syndrome were stimulated with antigen-presenting cells. The structure and dynamics of the immunological synapse were studied by confocal and video-microscopy. RESULTS: Upon stimulation by antigen-presenting cells, Wiskott-Aldrich syndrome protein-deficient T cells displayed reduced cytokine production and proliferation. Although Wiskott-Aldrich syndrome T cells formed conjugates with antigen-presenting cells at normal frequency and exhibited normal T-cell receptor down-regulation, they emitted actin-rich protrusions away from the immunological synapse area and their microtubule organizing center failed to polarize fully towards the center of the immunological synapse. In parallel, abnormally dispersed phosphotyrosine staining revealed unfocused synaptic signaling in Wiskott-Aldrich syndrome T cells. Time-lapse microscopy confirmed the anomalous morphology of Wiskott-Aldrich syndrome T-cell immunological synapses and showed erratic calcium mobilization at the single-cell level. CONCLUSIONS: Taken together, our data show that the Wiskott-Aldrich syndrome protein is required for the assembly of focused immunological synapse structures allowing optimal signal integration and sustained calcium signaling

Phenotypic and Histological Distribution Analysis Identify Mast Cell Heterogeneity in Non-Small Cell Lung Cancer

Mast cells (MCs) are multifaceted innate immune cells often present in the tumor microenvironment (TME). However, MCs have been only barely characterized in studies focusing on global immune infiltrate phenotyping. Consequently, their role in cancer is still poorly understood. Furthermore, their prognosis value is confusing since MCs have been associated with good and bad (or both) prognosis depending on the cancer type. In this pilot study performed on a surgical cohort of 48 patients with Non-Small Cell Lung Cancer (NSCLC), we characterized MC population within the TME and in matching non-lesional lung areas, by multicolor flow cytometry and confocal microscopy. Our results showed that tumor-associated MCs (TAMCs) harbor a distinct phenotype as compared with MCs present in non-lesional counterpart of the lung. Moreover, we found two TAMCs subsets based on the expression of CD103 (also named alphaE integrin). CD103+ TAMCs appeared more mature, more prone to interact with CD4+ T cells, and located closer to cancer cells than their CD103− counterpart. In spite of these characteristics, we did not observe a prognosis advantage of a high frequency of CD103+ TAMCs, while a high frequency of total TAMC correlated with better overall survival and progression free survival. Together, this study reveals that TAMCs constitute a heterogeneous population and indicates that MC subsets should be considered for patients’ stratification and management in future research

Stochastic asymmetric repartition of lytic machinery in dividing CD8+ T cells generates heterogeneous killing behavior

National audienceI calibrate a Multiple‐Risk Susceptible–Infected–Recovered model on the covid pandemic to analyze the impact of the age‐specific confinement and polymerase chain reaction (PCR) testing policies on incomes and mortality. Two polar strategies emergeas potentially optimal. The suppression policy would crush the curve by confining 90% of the population for 4 months to eradicate the virus. The flatten‐thecurvepolicy would reduce the confinement to 30% of the population for 5 months, followed by almost 1 year of free circulation of the virus to attain herd immunity without overwhelming hospitals.Both strategies yield a total cost of around 15% of annualgross domestic product (GDP) when combining the economic cost of confinement with the value of lives lost. I show that hesitating between the two strategies can have a huge societal cost, in particular if the suppression policy is stopped too early. Becauseseniors are much more vulnerable, a simple recommendation emerges to shelter them as one deconfines young and middle‐aged people to build our collective herd immunity. By doing so, one reduces the death toll of the pandemic together with the economic cost of the confinement, and the total cost is divided by a factor 2. I also show that expandingthe mass testing capacity to screen people sent back to work has a large benefit under various scenarios.This analysis is highly dependent upon deeply uncertain epidemiologic, sociological, economic, and ethical parameters

Blood leukocytes and macrophages of various phenotypes have distinct abilities to form podosomes and to migrate in 3D environments

International audienc

Plasmacytoid dendritic cell heterogeneity is defined by CXCL10 expression following TLR7 stimulation

Plasmacytoid dendritic cells (pDCs) play a critical role in bridging the innate and adaptive immune systems. pDCs are specialized type I interferon (IFN) producers, which has implicated them as initiators of autoimmune pathogenesis. However, little is known about the downstream effectors of type I IFN signaling that amplify autoimmune responses. Here, we have used a chemokine reporter mouse to determine the CXCR3 ligand responses in DCs subsets. Following TLR7 stimulation, conventional type 1 and type 2 DCs (cDC1 and cDC2, respectively) uniformly upregulate CXCL10. By contrast, the proportion of chemokine positive pDCs was significantly less, and stable CXCL10(+) and CXCL10(-) populations could be distinguished. CXCL9 expression was induced in all cDC1s, in half of the cDC2 but not by pDCs. The requirement for IFNAR signaling for chemokine reporter expression was interrogated by receptor blocking and deficiency and shown to be critical for CXCR3 ligand expression in Flt3-ligand-derived DCs. Chemokine-producing potential was not concordant with the previously identified markers of pDC heterogeneity. Finally, we show that CXCL10(+) and CXCL10(-) populations are transcriptionally distinct, expressing unique transcriptional regulators, IFN signaling molecules, chemokines, cytokines, and cell surface markers. This work highlights CXCL10 as a downstream effector of type I IFN signaling and suggests a division of labor in pDCs subtypes that likely impacts their function as effectors of viral responses and as drivers of inflammation