Extended Data Fig. 6: Production of itaconate upon TLR stimulation leads to increased peroxide accumulation.
From: Itaconate modulates immune responses via inhibition of peroxiredoxin 5
a Representative immunoblot of Irg1 expression in WT or Irg1-/- BMDMs non-stimulated or stimulated with R848 (1 μg/ml, 24 h). GAPDH serves as loading control (n = 3). b Intracellular levels of itaconate in BMDMs stimulated as in (a). To rescue itaconate levels in Irg1-/- cells were treated with itaconic acid (IA) (1 mM, at 4 h post stimulation) (n = 3 biological replicates). c-d mtPY1 levels and representative histograms in BMDMs non-stimulated or stimulated as with LPS (0.1 ug/ml) or R848 (1 μg/ml) or poly(I:C) (20 μg/ml) for 24 h (n = 6 (c), n = 5 (d) independent cultures). e Intracellular itaconate levels in WT and Irg1-/- BMDMs treated as in (c) (n = 3 biological replicates). f Nitrite+Nitrate levels in supernatant of WT or Irg1-/- BMDMs stimulated as in (c) (n = 4 biological replicates). g Nitrite levels in supernatants of BMDMs stimulated as in (c) (n = 4 independent cultures) h Randomized design assignment of TMT10 reagents used for multiplexing of enriched samples. Data represent mean ±SEM. Statistical analysis: c-d One sample t-test.
