Inflammatory signals are integrated through TNF via NF-κB to directly regulate PU.1 in HSCs in vivo. (A) NF-κB-signaling is essential for PU.1 induction by TNF. Quantification of HSC PU.1-eYFP levels 12 hours after stimulation with NF-κB inhibitor (NEMO-binding domain peptide, 30 μM) or p38 MAPK inhibitor (SB239063, 50 μM). Cells: Ctrl: 162, TNF: 223, NF-κB-i: 233, p38 MAPK-i: 48. ***P < .002, ANOVA. (B) TNF is required for PU.1 induction by LPS in vivo. Immunofluorescence quantification of PU.1 protein in C57BL/6 or TNF^−/− HSCs 24 hours after in vivo LPS or TNF stimulation. Data presented from wild-type: PBS, n = 4; LPS, n = 4; TNF, n = 2 mice. Knockout: PBS, n = 4; LPS, n = 4; TNF, n = 3 mice. *P < .05; **P < .01, ANOVA.