Fig. 1: NO fluxes regulate PDH and alter PDH-E2 cofactor lipoate in macrophages.

a Enzymatic activity of PDH in total cell lysates from WT BMDMs from male and female mice (mixed sex in all groups) after 16 h activation with LPS + IFNγ or DETA/NO (100 μM) (n > 3 mice). b Representative Seahorse analysis of permeabilized WT BMDMs from male and female mice (mixed sex in all groups) where state 3-OCR was elicited by a mixture of pyruvate and malate to measure PDH flux. Bar graphs show quantified respiration in 16 h activated WT (n > 8 wells). Data in (a) and (b) were analyzed by one-way ANOVA with Dunnet’s multiple comparisons test. c Quantified pyruvate/malate respiration from Seahorse analysis of permeabilized WT BMDMs from male and female mice (mixed sex in all groups) after 8 h activation, performed in the presence of vehicle or lipoic acid (n > 3 mice) shown as percentage of OCR relative to untreated. Data were analyzed by two-way ANOVA (Sidak’s post-tests). d Schematic illustration of PDH enzyme with emphasis on E2 subunit. e Interrogated peptide of PDH-E2 predicted to bind lypoyl group on functional lysine (K). f–h Major products, possible structures and abundances of E2 K¹³¹-lipoylated peptide in mitochondrial lysates from WT BMDMs from male mice after 16 h stimulation with LPS + IFNγ or DETA/NO (500 μM) (n = 3 technical repeats) Data were analyzed by two-way ANOVA with Dunnet’s multiple comparisons test. All error bars display mean ± SEM. Data shown are representative of two or more independent experiments. Source data are provided as a Source Data file.