Figure 3.
CD8+ T-cell function is impaired by BM long-chain fatty acid uptake via FATP1. (A-D) BM mononuclear cells from controls (n = 6) or patients with MM who had relapsed (n = 6) were cultured in autologous PB or BM plasma as indicated for 72 hours with stimulation via CD3/CD28. (A) CD8+ T-cell mitochondrial mass was assessed by flow cytometry using MVG and is shown for each group (left) and both groups combined (right). (B) At 72 hours, T cells were activated with PMA/ionomycin and expression of IFN-γ was assessed within CD8+ T cells by flow cytometry. (C-D) IFN-γ and TNF-α in cell culture supernatants was measured by ELISA. (E-H) BM mononuclear cells were cultured and assessed as in panels A-D but in either complete or lipid-depleted BM autologous plasma as indicated. (I-K) BM mononuclear cells from patients with MM at diagnosis (n = 6) were cultured in autologous PB (I,K) or BM (J-K) plasma as in panels A-D, in absence or presence of ferrostatin and assessed for IFN-γ expression by ELISA. (L) Proportion of CD8+ T cells expressing CD36 within indicated BM mononuclear cell samples from controls (n = 10), individuals with MGUS (n = 8), asymptomatic MM (asymptomatic; n = 9), or MM at diagnosis (diagnosis; n = 7) were assessed by flow cytometry. (M) Expression of IFN-γ, TNF-α, and IL-2 by CD36+ or CD36− CD8+ T cells within indicated BM mononuclear cell samples as in panel L. (N-O) BM mononuclear cells from patients with MM at diagnosis (n = 7) were cultured in autologous BM plasma as in panel A-D, in absence or presence of the CD36 inhibitor, SSO, and assessed for IFN-γ expression within CD8+ T cells by flow cytometry (N) or by ELISA (O). (P) single-cell RNA-sequencing UMAP projections of BM mononuclear cells from 2 patients with newly diagnosed MM, with cell type annotation (top left) and transcript abundance of indicated genes overlaid. (Q-S) BM mononuclear cells from patients with MM at diagnosis (n = 7) were cultured in autologous PB (Q,S) or BM (R-S) plasma as in panels A-D, in absence or presence of the FATP1 inhibitor, FATP1in2, and assessed for IFN-γ expression by ELISA. Significance was calculated using Wilcoxon matched-pairs signed rank test for panels A-H (circles), I-J,N-O,Q-R; and paired t test for panels K,S; ∗P < .05, ∗∗P < .01, and ∗∗∗P < .0001. UMAP, Uniform Manifold Approximation and Projection.

CD8+ T-cell function is impaired by BM long-chain fatty acid uptake via FATP1. (A-D) BM mononuclear cells from controls (n = 6) or patients with MM who had relapsed (n = 6) were cultured in autologous PB or BM plasma as indicated for 72 hours with stimulation via CD3/CD28. (A) CD8+ T-cell mitochondrial mass was assessed by flow cytometry using MVG and is shown for each group (left) and both groups combined (right). (B) At 72 hours, T cells were activated with PMA/ionomycin and expression of IFN-γ was assessed within CD8+ T cells by flow cytometry. (C-D) IFN-γ and TNF-α in cell culture supernatants was measured by ELISA. (E-H) BM mononuclear cells were cultured and assessed as in panels A-D but in either complete or lipid-depleted BM autologous plasma as indicated. (I-K) BM mononuclear cells from patients with MM at diagnosis (n = 6) were cultured in autologous PB (I,K) or BM (J-K) plasma as in panels A-D, in absence or presence of ferrostatin and assessed for IFN-γ expression by ELISA. (L) Proportion of CD8+ T cells expressing CD36 within indicated BM mononuclear cell samples from controls (n = 10), individuals with MGUS (n = 8), asymptomatic MM (asymptomatic; n = 9), or MM at diagnosis (diagnosis; n = 7) were assessed by flow cytometry. (M) Expression of IFN-γ, TNF-α, and IL-2 by CD36+ or CD36− CD8+ T cells within indicated BM mononuclear cell samples as in panel L. (N-O) BM mononuclear cells from patients with MM at diagnosis (n = 7) were cultured in autologous BM plasma as in panel A-D, in absence or presence of the CD36 inhibitor, SSO, and assessed for IFN-γ expression within CD8+ T cells by flow cytometry (N) or by ELISA (O). (P) single-cell RNA-sequencing UMAP projections of BM mononuclear cells from 2 patients with newly diagnosed MM, with cell type annotation (top left) and transcript abundance of indicated genes overlaid. (Q-S) BM mononuclear cells from patients with MM at diagnosis (n = 7) were cultured in autologous PB (Q,S) or BM (R-S) plasma as in panels A-D, in absence or presence of the FATP1 inhibitor, FATP1in2, and assessed for IFN-γ expression by ELISA. Significance was calculated using Wilcoxon matched-pairs signed rank test for panels A-H (circles), I-J,N-O,Q-R; and paired t test for panels K,S; ∗P < .05, ∗∗P < .01, and ∗∗∗P < .0001. UMAP, Uniform Manifold Approximation and Projection.

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