Figure 3: Accumulation of succinate and itaconate is independent of alterations to oxidative phosphorylation.

A) A schematic depicting itaconate and succinate in the context of the tricarboxylic acid cycle (TCA cycle). IRG1, cis-aconitase decarboxylase 1; SDH, succinate dehydrogenase. B) Intracellular abundances of itaconate and succinate from control (Ctl) and BMDMs treated with Pam3, Poly I:C, or Pam3 + Poly I:C for 24 hr. (n = 7). C) Intracellular abundances of itaconate and succinate from control (Ctl) and BMDMs treated with 10 ng/mL LPS, 20 ng/mL IFN-γ, or LPS + IFN-γ for 24 hr. (n = 6). D) Intracellular abundances of itaconate and succinate in wildtype (WT) or IRG1-null (Irg1^−/−) control (Ctl) and BMDMs treated with LPS + IFN-γ for 24 hr. (n = 6). E&F) Representative oxygen consumption rates, where not visible, error bars are obscured by the symbol, O, oligomycin; F, FCCP; R/A, rotenone/antimycin A (n = 1 biological with 5 technical replicates) and (E) ATP-linked and maximal respiration for WT or Irg1^−/− control (Ctl) and BMDMs treated with LPS + IFN-γ for 24 hr. (n = 4) (F). G) ATP-linked and maximal respiration rates for control (Ctl) and HMDMs treated with 50 ng/mL LPS for 24 hr. (n = 5). H) Intracellular abundances of itaconate and succinate from control (Ctl) and HMDMs treated with 50 ng/mL LPS for 24 hr. (n = 2). All data are mean ± SEM with statistical analysis conducted on data from biological replicates, each of which included multiple technical replicates, unless otherwise indicated. Statistical analysis for (B) and (C) was performed as an ordinary one-way, ANOVA followed by Tukey’s post hoc multiple comparisons test. Statistical analysis for (D) and (F) was performed as an ordinary two-way, ANOVA followed by Sídák’s post hoc multiple comparisons test. Statistical analysis for (G) was performed as an unpaired, two-tailed t-test.