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. Author manuscript; available in PMC: 2018 Aug 27.
Published in final edited form as: Cell Rep. 2018 Jul 31;24(5):1136–1150. doi: 10.1016/j.celrep.2018.06.065

Figure 2. Autophagy Inhibition during Stimulation Induces T Cell Anergy.

Figure 2.

(A) Expression of anergy-associated genes (qPCR) in Th1 cells activated in the presence or absence of L/N or 3MA.

(B) Expression of anergy-associated genes (qPCR) in cells transfected with non-targeting (Ctrl) or Atg7-specific siRNAs and activated for 24 hr.

(C) Foxp3 gene expression (qPCR) in Th1 cells activated in the presence or absence of 3MA. mRNA obtained from purified Tregs is included as control.

(D)Th1 cells were anergized with anti-CD3 (Anerg), activated in the absence (Preact) or presence of L/N (Preact-L/N) or 3MA (Preact-3MA) for 24 hr. Cells were washed to remove inhibitors, cultured for 5 days with 50 ng/mL IL-2, and re-stimulated. IL-2 production and cell proliferation were measured.

(E) IL-2 and cell proliferation were measured as in (D) in cells transfected with Ctrl or Atg7 siRNAs 24 hr prior to pre-activation.

(F) Wild-type or NFAT1-deficient Th1 cells were stimulated for 24 hr in the presence or absence of 3MA. Cells were washed and re-stimulated after 5 days to measure IL-2 production and cell proliferation.

Data represent mean + SEM from 3 (A–C), 4 (D and E), or 5 (F) different experiments. *p < 0.05; **p < 0.01; ***p < 0.001, ANOVA with Tukey post-test for (A), (C), (D), and (F); two-tailed t test for (B) and (E). See also Figures S2–S4.