Journal: Frontiers in Immunology

Article Title: Unspecific CTL Killing Is Enhanced by High Glucose via TNF-Related Apoptosis-Inducing Ligand

doi: 10.3389/fimmu.2022.831680

Figure Lengend Snippet: Metabolic processes in CTLs are reprogrammed by high glucose. Primary human CD8 + T cells were stimulated with CD3/CD28 beads for 3 days in NG (5.6 mM) or HG (25 mM) medium. (A–D) Oxidative phosphorylation (n = 5 donors) and glycolysis (n = 3 donors) of CTLs from two independent experiments were determined with seahorse assay. One representative donor for oxidative phosphorylation and glycolysis is shown in (A, C) , respectively. (E) Heatmap of log2-transformed gene expression data. The microarray data was normalized using quantile normalization. Duplicate probes for genes were aggregated by taking the median intensity. Genes were filtered for those with an absolute fold change > 1.5, and a Benjamini-Hochberg adjusted p -value < 0.05. (F) Expression of glucose transporters at mRNA level from the transcriptomics data ( Table S1 ) of 6 donors collected from five independent stimulations and two independent microarray analyses. A.U. stands for arbitrary units. (G, H) ROS production in CD8 + T cells was determined at 6 hours after CD3/CD28 bead stimulation by DCFDA (n = 5 donors from three independent experiments). One representative donor is shown in (G) Connected lines in (H) are the data from the same donor. (I, J) H 2 O 2 enhances TRAIL expression in CTLs in NG. CD8 + T cells were stimulated with CD3/CD28 beads in presence or absence of H 2 O 2 for 3 days (n = 5 from three independent experiments). One representative donor is shown in (I, K–N) Inhibition of ROS production abolishes HG-enhanced TRAIL expression in CTLs. NAC ( K , L , 10 mM, n = 6 donors) or MitoQ ( M, N , 0.4 μM, n = 5 donors) from three independent experiments was added during the activation for 3 days. One representative donor for NAC and MitoQ is shown in (K, M) , respectively. Results are represented as Mean ± SD. Data were analyzed by two-tailed unpaired Student’s t test (B, D) , two-tailed paired Student’s t test (H) or one-way ANOVA with Bonferroni’s multiple comparison test (J, L, N) .

Article Snippet: Differential expression analysis of the Agilent microarray data was performed with the Linear Models for Microarray Data (limma) R package ( ).

Techniques: Transformation Assay, Expressing, Microarray, Inhibition, Activation Assay, Two Tailed Test