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. 2021 Oct 18;118(42):e2102698118. doi: 10.1073/pnas.2102698118

Fig. 2.

Fig. 2.

Heme training relies on Syk/JNK activation in human monocytes. (A) Western blot of p-S6 24 h after heme ± rapamycin stimulation. (B) TNF production after restimulation with LPS at day 7 of vehicle- or heme-trained Mφ ± rapamycin. The representative Western blots of at least three independent donors are pictured. (C) Western blot of Syk phosphorylation after heme treatment of human monocytes at indicated time points. (D) Syk phosphorylation 20 min after heme stimulation ± the Syk inhibitor R406. (E) TNF production after restimulation with LPS at day 7 of vehicle- or heme-trained Mφ ± R406. (F) Western blot of total H3K27ac 24 h after heme treatment ± R406. (G) ChIP-qPCR analysis of H3K27ac at the IL8 promoter (n = 3 independent donors). (H) Western blot of JNK phosphorylation after heme treatment of human monocytes at indicated time points. (I) Western blot of JNK phosphorylation 20 min after heme stimulation ± R406. (J) Western blot and densitometry of JNK activation 20 min after heme stimulation ± the JNK inhibitor SP600125. (K) TNF production after restimulation with LPS at day 7 of vehicle- or heme-trained Mφ ± SP600125. All enzyme-linked immunosorbent assay results derived ≥3 independent experiment à 2 to 3 donors; Two-way ANOVA with Fisher’s LSD; mean ± SEM, *P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001; ****P ≤ 0.0001. GAPDH, glyceraldehyde 3-phosphate dehydrogenase; H3, histone 3; H3K27ac, histone 3 lysine 27 acetylation; JNK-I, JNK inhibitor SP600125.