Figure 4: Neither Pam3 nor Poly I:C induce characteristic breaks in the TCA cycle.

A) Intracellular abundance of citrate from control (Ctl) and BMDMs treated with Pam3, Poly I:C, or Pam3 + Poly I:C for 24 hr. (n = 7). B) State 3 respiration from permeabilized control (Ctl) and BMDMs treated with Pam3, Poly I:C, or Pam3 + Poly I:C for 24 hr. Permeabilized BMDMs were offered pyruvate/malate, citrate, glutamate/malate, or succinate/rotenone as substrates (n = 4). C) A graphical schematic depicting the labeling patterns of TCA cycle metabolites from ¹³C₆ Glucose (black circles) and ¹³C₅ Glutamine (grey circles). CS, citrate synthase; Aco., aconitase; IDH, isocitrate dehydrogenase; α-KGDH, alpha-ketoglutarate dehydrogenase; SCS, succinyl CoA synthetase; SDH, succinate dehydrogenase; FH, fumarate hydratase; MDH, malate dehydrogenase; AST, aspartate transaminase. D&E) Percent enrichment of denoted isotopologues from either ¹³C₆ Glucose or ¹³C₅ Glutamine from control (Ctl) and BMDMs treated with Pam3, Poly I:C, or Pam3 + Poly I:C for 24 hr. BMDMs were treated in unlabeled media for 16 hr. then changed to tracing media with stimuli for 6 hr. (n = 4). 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 (A) and (B) was performed as an ordinary one-way, ANOVA followed by Tukey’s post hoc multiple comparisons test. Statistical analysis for (D) and (E) was performed as an ordinary two-way, ANOVA followed by Tukey’s post hoc multiple comparisons test.