Fig. 2. Murine T cells are metabolically impaired by AML cells in vitro and in vivo.
(A) Recipient mice (BALB/c background) were transplanted with 5 × 106 C57BL/6 bone marrow (BM) cells (allo BM) with or without additional 1 × 105 syngeneic leukemia cells (WEHI-3B luc) and 2 × 105 C57BL/6 T cells (allo Tc). Tumor growth was monitored by bioluminescence imaging (BLI). d0, day 0. (B) Analysis of ECAR of reisolated CD8+ T cells (n = 6, means ± SEM). (C) Analysis of OCR (n = 6, means ± SEM). The experiment was performed six times (with 18 mice in total); n values represent biologically independent experiments, pooling together three mice for each experiment. P values were determined using the one-sample Student’s unpaired t test. (D) ECAR measured at baseline and in response to glucose, oligomycin, and 2-deoxyglucose (2-DG) for CD8+ T cells isolated from 48-hour cell culture under CD3/CD28 stimulation either alone or in coculture with AML cells or incubated in medium originating from 48-hour AML cell culture. Graph is representative of four biologically independent experiments. (E) ECAR of CD8+ T cells (n = 4 to 6). (F) OCR of CD8+ T cells (n = 4 to 6). (G) ECAR of CD8+ T cells isolated from 48-hour cell culture and stimulated with CD3/CD28, from T cells incubated alone, in coculture with BM-derived dendritic cells (BMDCs), or T cells incubated in medium originating from 48-hour BMDC cell culture (n = 6). (H) OCR of CD8+ T cells (n = 6). The n values represent biologically independent experiments (means ± SEM). P values were determined using one-way ANOVA. (I) Microarray-based analysis of gene expression of CD8+ T cells isolated after 48-hour CD3/CD28 stimulation and incubated in fresh medium or medium originating from 48-hour AML or BMDC cell culture. Bar plot depicts the top 10 down-regulated gene sets in T cells cultured in AML medium compared to T cells cultured in DC medium. MTORC1, mammalian target of rapamycin complex 1. (J) Tile display shows differentially regulated OXPHOS genes as determined from linear model analysis. Only regulated genes with an adjusted P < 0.05 were selected. The color scale represents the row-wise Z score of gene expression with red being the highest and blue being the lowest. (K) Tile display shows differentially regulated glycolysis genes.