Extended Data Fig. 1: Adoptively transferred young-origin CD4 T cell differentiation in an aged environment.

a. Representative flow cytometry plots showing the gating strategy used to define CD4 T cell subsets. b. Representative flow cytometry plots showing phenotypic characteristics of the young CD45.1 cells used for adoptive transfer experiments. c. Quantitative analysis of CD4⁺ cells among the transferred cells in young (n = 6) or old (n = 4) WT mice (CD45.2). d. Histogram plots showing fluorescence of CFSE-labeled CD45.1⁺CD4⁺ cells relative to unlabeled cells. e. Graph showing the analysis of transferred cells (CD45.1⁺); the percentage of CD4⁺CD44⁺ T cells out of CFSE⁻ or CFSE⁺ cells in old mice (CD45.2; n = 6). f. Representative confocal images showing γH2A.X immunostaining in the Vehicle (Left) and Senolytic (Right) groups. Scale bars are indicated in the figures. g. Representative immunofluorescence images showing liver sections of Vehicle mice immunolabeled with a secondary antibody as a negative control (Lower) or stained with DAPI (Upper). Scale bars are indicated in the figures. h-i. The percentages of Treg (FOXP3⁺, Left), naïve (CD3⁺CD4⁺CD62L⁺CD44⁻, Middle), effector (CD3⁺CD4⁺CD62L⁻CD44⁺, Right) (h) and PD1⁺ (CD3⁺CD4⁺CD62L⁻CD44⁺PD1⁺) (i) CD4⁺ T cells originating from CD45.1 young mice and transferred into CD45.2 old control (n = 11) and senolytic drug-treated (n = 10) mice. Each dot represents the percentages in a single mouse. Bars represent mean ± SEM from two (c and e) or three (h-i) independent experiments. Data were analyzed using two-tailed unpaired (c, h-i) and paired (e) Student’s t-tests.