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. 2020 Jan 21;30(3):687–698.e6. doi: 10.1016/j.celrep.2019.12.050

Figure 4.

Figure 4

Enhanced Autophagy Levels Are Linked to Effector Function and Mitochondrial Fitness in Human T Cells

(A) Example plots of IFN-γ, LC3, granzyme B (GzB), and perforin (perf; gated on CD8+ T cells) and histograms of LC3 staining for PBMC ex vivo or after anti-CD3/CD28 stimulation (3 days; see also Figure S3).

(B) LC3 staining of CD8+ T cells from unstimulated PBMCs (IFN-γ−), IFN-γ− and IFN-γ+ CD8+ T cells after anti-CD3/CD28 stimulation (3 days; eight biological replicates).

(C and D) LC3 staining on GzB and perf-expressing CD8+ T cells ex vivo (C) and after anti-CD3/CD28 stimulation (D) (3 days; eight biological replicates; box whisker, Tukey).

(E) Example mitochondrial staining of CD8+ T cells in blood (PBMCs; black) and liver (IHLs; red) and summary data for the ex vivo percentage of total CD8+ T cells with depolarized mitochondria (mitoTracker deep red [MtDR]lo; see also Figure S4; PBMCs, 10; and IHLs, 15 biological replicates).

(F) Ex vivo percentage of CD8+ TRM cell subsets in the liver with depolarized mitochondria (14 biological replicates; box whisker, Tukey; outliers shown as dots).

(G) The percentage of total CD8+ T cells or CD8+ TRM cell subsets with depolarized mitochondria after overnight culture of IHLs with DMSO (untreated), MRT68921 dihydrochloride (10 μM), bafA1 (0.1 μM), or reagent A (chloroquine diphosphate, 1:1000, FlowCellect LC3 kit; 13–15 biological replicates).

Cells were treated with bafA1 (A–D). Bars at mean (B, E, and G). Friedman test (ANOVA) with Dunn’s post hoc test for pairwise multiple comparisons (B, C, D, and F). Kruskal-Wallis test with Dunn’s post hoc test for unpaired multiple comparisons (G). Mann-Whitney unpaired t test for total CD8+ PBMCs versus IHLs (E). Wilcoxon paired t test for untreated versus that treated with MRT68921 (G). Bars at mean (E and G). ∗p < 0.05, ∗∗p < 0.005, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001.