The heterogeneity and sexual-dimorphism of human TCRαβ+CD4–CD8– double negative T cells

double negative T (DNT) cells are increasingly being recognized for their critical roles in the progression of various diseases. However, the molecular and functional signatures remain poorly understood and controversial. In this study, we characterize the single-cell transcriptome, TCR repertoire, and chromatin accessibility of peripheral DNT cells from 10 healthy donors. Naïve, regulatory, cytotoxic, antibody-dependent cell-mediated cytotoxicity, and helper-like DNT cell subsets exhibit specific gene expression, transcriptome regulation, development signature, and sexual heterogeneity, respectively. Using publicly available data, we observe changes in the proportion and gene expression of DNT cell subsets, especially the helper subset, in patients with ulcerative colitis and type 1 diabetes. Overall, human DNT cells exhibit phenotypic and functional diversity, providing a valuable framework for understanding the development and progression of immune-related diseases.