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During thymic development, murine invariant natural killer T cells go through different maturation stages differentiating into distinct sublineages, namely, invariant natural killer T1, 2, and 17 cells. Recent reports indicate that invariant natural killer T2 cells display immature properties and give rise to other subsets, whereas invariant natural killer T1 cells seem to be terminally differentiated. Whether human invariant natural killer T cells follow a similar differentiation model is still unknown. To define the maturation stages and assess the sublineage commitment of human invariant natural killer T cells during thymic development, in this study, we performed single-cell RNA sequencing analysis on human V\u03b124+V\u03b211+ invariant natural killer T cells isolated from thymocytes. We show that these invariant natural killer T cells displayed heterogeneity, and our unsupervised analysis identified 5 clusters representing different maturation stages, from an immature profile with high expression of genes important for invariant natural killer T cell development and proliferation to a mature, fully differentiated profile with high levels of cytotoxic effector molecules. Evaluation of expression of sublineage-defining gene sets revealed mainly cells with an invariant natural killer T2 signature in the most immature cluster, whereas the more differentiated ones displayed an invariant natural killer T1 signature. Combined analysis with a publicly available single-cell RNA sequencing data set of human invariant natural killer T cells from peripheral blood suggested that the 2 main subsets exist both in thymus and in the periphery, while a third more immature one was restricted to the thymus. Our data point to the existence of different maturation stages of human thymic invariant natural killer T cells and provide evidence for sublineage commitment of invariant natural killer T cells in the human thymus.<\/jats:p>","DOI":"10.1093\/jleuko\/qiad113","type":"journal-article","created":{"date-parts":[[2023,9,21]],"date-time":"2023-09-21T16:18:11Z","timestamp":1695313091000},"page":"401-409","source":"Crossref","is-referenced-by-count":13,"title":["Single-cell transcriptomics reveal different maturation stages and sublineage commitment of human thymic invariant natural killer T cells"],"prefix":"10.1093","volume":"115","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-1788-2807","authenticated-orcid":false,"given":"Kristina","family":"Maas-Bauer","sequence":"first","affiliation":[{"name":"Division of Blood and Marrow Transplantation and Cellular Therapy, Stanford University , Center for Clinical Sciences Research Building, 269 W. Campus Drive, Stanford, CA 94305, United States"},{"name":"Department of Hematology, Oncology, and Stem Cell Transplantation, Medical Center\u2014University of Freiburg, Faculty of Medicine , Hugstetter Str. 55, Freiburg 79106 , Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Natalie","family":"K\u00f6hler","sequence":"additional","affiliation":[{"name":"Department of Hematology, Oncology, and Stem Cell Transplantation, Medical Center\u2014University of Freiburg, Faculty of Medicine , Hugstetter Str. 55, Freiburg 79106 , Germany"},{"name":"CIBSS\u2014Centre for Integrative Biological Signalling Studies, University of Freiburg , Sch\u00e4nzlestr. 18, Freiburg 79104 , Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anna-Verena","family":"Stell","sequence":"additional","affiliation":[{"name":"Department of Hematology, Oncology, and Stem Cell Transplantation, Medical Center\u2014University of Freiburg, Faculty of Medicine , Hugstetter Str. 55, Freiburg 79106 , Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Melissa","family":"Zwick","sequence":"additional","affiliation":[{"name":"Department of Hematology, Oncology, and Stem Cell Transplantation, Medical Center\u2014University of Freiburg, Faculty of Medicine , Hugstetter Str. 55, Freiburg 79106 , Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Swati","family":"Acharya","sequence":"additional","affiliation":[{"name":"Sean N. Parker Center for Asthma and Allergy Research, Department of Medicine, Stanford University , 240 Pasteur Dr, Stanford, CA 94304 , United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anne","family":"Rensing-Ehl","sequence":"additional","affiliation":[{"name":"Institute for Immunodeficiency, Center for Chronic Immunodeficiency, Medical Center-University of Freiburg, Faculty of Medicine , Breisacher Str. 115, Freiburg 79106 , Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Christoph","family":"K\u00f6nig","sequence":"additional","affiliation":[{"name":"Institute for Immunodeficiency, Center for Chronic Immunodeficiency, Medical Center-University of Freiburg, Faculty of Medicine , Breisacher Str. 115, Freiburg 79106 , Germany"},{"name":"Faculty of Biology, University of Freiburg , Sch\u00e4nzlestr. 1, Freiburg 79104 , Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Johannes","family":"Kroll","sequence":"additional","affiliation":[{"name":"Department of Cardiovascular Surgery, Heart Center Freiburg University , Hugstetter Stra\u00dfe 55, Freiburg 79106 , Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jeanette","family":"Baker","sequence":"additional","affiliation":[{"name":"Division of Blood and Marrow Transplantation and Cellular Therapy, Stanford University , Center for Clinical Sciences Research Building, 269 W. Campus Drive, Stanford, CA 94305, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Stefanie","family":"Ko\u00dfmann","sequence":"additional","affiliation":[{"name":"Department of Hematology, Oncology, and Stem Cell Transplantation, Medical Center\u2014University of Freiburg, Faculty of Medicine , Hugstetter Str. 55, Freiburg 79106 , Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Amandine","family":"Pradier","sequence":"additional","affiliation":[{"name":"Division of Hematology, Department of Oncology, Geneva University Hospitals , Rue Gabrielle-Perret-Gentil 4, Geneva 1205 , Switzerland"},{"name":"Translational Research Center for Oncohematology, Department of Medicine, Faculty of Medicine, University of Geneva , Rue Michel-Servet 1, Geneva 1211 , Switzerland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sisi","family":"Wang","sequence":"additional","affiliation":[{"name":"Translational Research Center for Oncohematology, Department of Medicine, Faculty of Medicine, University of Geneva , Rue Michel-Servet 1, Geneva 1211 , Switzerland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Myl\u00e8ne","family":"Docquier","sequence":"additional","affiliation":[{"name":"iGE3 Genomics Platform, University of Geneva , Rue Michel-Servet 1, Geneva 1211 , Switzerland"},{"name":"Department of Genetics & Evolution, University of Geneva , Rue Michel-Servet 1, Geneva 1211 , Switzerland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David B","family":"Lewis","sequence":"additional","affiliation":[{"name":"Division of Allergy, Immunology and Rheumatology, Department of Pediatrics, Stanford University School of Medicine , 240 Pasteur Dr, Stanford, CA 94304 , United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Robert S","family":"Negrin","sequence":"additional","affiliation":[{"name":"Division of Blood and Marrow Transplantation and Cellular Therapy, Stanford University , Center for Clinical Sciences Research Building, 269 W. Campus Drive, Stanford, CA 94305, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Federico","family":"Simonetta","sequence":"additional","affiliation":[{"name":"Division of Blood and Marrow Transplantation and Cellular Therapy, Stanford University , Center for Clinical Sciences Research Building, 269 W. 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