Fig. 3: In obesity, As-iNKT1 cells give rise to Ac-iNKT1 cells.

a Unsupervised clustering of TCRβ^int/CD1d.PBS57 tetramer⁺ iNKT cells from WAT of NCD-, 1-week, or 8-week HFD-fed mice. 1,287 cells from NCD-, 1,800 cells from 1-week, and 2,400 cells from 8-week HFD-fed mice on a UMAP plot. b Heatmap showing the expression levels of subpopulation marker genes. c Proportion of each adipose iNKT cell subpopulation in scRNA-seq data. d Representative FACS plots and proportion of each subpopulation among total adipose iNKT cells in NCD- (n = 7) or 8-week HFD-fed mice (n = 8). e Gene expression levels of Nr4a1, Hmgb2, Trbv13-1, Cd160, and Klrk1 in adipose iNKT cell subpopulations under each condition. p-values were adjusted for multiple comparisons using Bonferroni correction. f In silico pseudotime analysis of adipose iNKT1 cells. g Clonotype overlap analysis of adipose iNKT cell subpopulations. h Representative clones showing clonotype overlapping between As-iNKT1, Ac-iNKT1, and A-Cycling iNKT1 cells. i Experimental scheme for adoptive transfer of CD45.1⁺ Au-iNKT1 and As-iNKT1 cells. iNKT cells were sorted from CD45.1 mice 1-week after α-GC injection and injected into each WAT fat pad of 16-week HFD-fed CD45.2 Jα18 KO mice. j, k Representative FACS plots and composition of injected CD45.1⁺ donor iNKT cells in recipient mice after 3 weeks (Before injection (n = 2), Au-iNKT1 post injection (n = 2), and As-iNKT1 post injection (n = 3)). l Schematic diagram of iNKT1 differentiation process in adipose tissue. m CDR3β amino acid sequences of most prevalent CDR3β length of each adipose iNKT1 cell subpopulation. Data were collected from 8-week-HFD condition. Data are represented as mean ± SD. n.s., non-significant. Two-tailed unpaired Student’s t test (d, e, j, and k). Au-iNKT1; Adipose universal iNKT1, As-iNKT1; Adipose specific iNKT1. Ac-iNKT1; Adipose cytotoxic iNKT1.