CD8⁺ T-cell mitochondrial mass and long-chain fatty acid uptake are altered during progression of MM and in the BM microenvironment. (A-B) Mitochondrial mass of (A) CD8⁺ and (B) CD4⁺ T cells within BM mononuclear cells from controls (n = 10), individuals with MGUS (n = 13), asymptomatic MM (asymptomatic; n = 11), or MM at diagnosis (diagnosis; n = 12) was assessed by flow cytometry using MVG and is expressed relative to the mean of each experiment. (C-D) Mitochondrial mass of BM and PB CD8⁺ and CD4⁺ T cells is directly compared for each group and for all patient samples combined. (E) CoxIV MFI of CD8⁺ T cells positive or negative for IFN-γ or TNF-α as indicated from control (n = 4) or MM at diagnosis (n = 4) samples. (F) Correlation of mitochondrial mass (MVG fluorescence) with frequency of BM CD8⁺ T cells positive (+) for the indicated cytokines for all patient samples combined. (G) C₁₆-BODIPY uptake of BM CD8⁺ T cells within indicated groups expressed relative to the mean of each experiment. (H) C₁₆-BODIPY uptake of BM and PB CD8⁺ T cells is directly compared for each group and for all patient samples combined. (I) C₁₆-BODIPY uptake of BM and PB CD8⁺ and CD4⁺ T cells directly compared for all patients combined. (J) C₁₆-BODIPY uptake of panel J NV, CM, EM, and EMRA BM CD8⁺ T-cell subsets and subsets expressing TIGIT and/or PD-1 as indicated from samples with MM at diagnosis (n = 12). (K) Lipid peroxidation capacity of CD8⁺ T cells within BM and PB mononuclear cells from patients with MM at diagnosis (n = 4), assessed using the lipid peroxidation probe Bodipy 581/591. For panels A-B,G,I-J, significance was calculated using 1-way ANOVA or for panels C-D (bars) and E,H (bars) 2-way ANOVA and Sidak multiple comparison tests; for panels C-D,H (circles) and K, a paired t test was used; and for panel F, Pearson correlation; ∗P < .05, ∗∗P < .01, ∗∗∗P < .0001, and ∗∗∗∗P < .00001.