Figure 1. Unlike CD8⁺ T_E cells, CD8⁺ T_M cells do not acquire substantial amounts of extracellular fatty acids.

OT-I cells were injected i.v. into congenic recipients. Mice were infected with LmOVA to generate T_E (7 days) and T_M (≥ 21 days) cells, and peripheral blood was collected 1h after i.v. Bodipy FL C₁₆ injection and uptake quantified by FACS. (A) Representative plots (histogram) and average MFI (bar graph) of Bodipy FL C₁₆ in T_E and T_M K^bOVA-specific CD8⁺ OT-I⁺ cells from 2 experiments (n=7-8 mice/group). Bar graphs show mean ± SEM, **p <0.01. (B) Representative plots (histogram) of Bodipy FL C₁₆ MFI in polyclonal T_E (CD44^hi CD62L^lo), T_M (CD44^hi CD62L^hi), and T_N (CD44^lo CD62L^hi) CD8⁺ T cells 7 days post-infection and average MFI (bar graph) of polyclonal CD8⁺ T cells from 2 experiments (n=9 mice/group). Bar graphs show mean ± SEM ***p <0.001. (C) OT-I cells were activated with OVA peptide and IL-2 for 3 days and subsequently cultured in IL-15 or IL-2 for 4 more days to generate IL-15 T_M and IL-2 T_E cells respectively. (D) OTI cells were incubated with Bodipy FL C₁₆ on d4-7 of culture, and uptake was measured. MFIs were normalized to the daily MFI of IL-2 T_E cells. Data from 3 experiments shown as mean ± SEM, *p <0.05 by one sample t-test. (E) CD36 expression on d7 IL-2 T_E and IL-15 T_M cells, or polyclonal T cells. Data represent 3 experiments. (F-H) Day 6 IL-2 T_E or IL-15 T_M cells were cultured overnight ± oleate then stained with Bodipy. (F) Representative plots showing Bodipy MFI and (G) average Bodipy MFI from 3 experiments show mean ± SEM, *p <0.05, **p <0.01. (H) Images showing Bodipy (green) and CD8 (red) represent 4 experiments. Scale bar = 2 µM (I) OT-I cells expressing a lipid droplet-targeting construct (LD-GFP) were cultured overnight ± oleate. Images show LD-GFP (green) and CD8 (red) and represent 2 experiments. (J) Cells were incubated with Bodipy-LDL or left unstained (Control), data represent 2 experiments.