Figure 1. LPS stimulates DLL4 expression in HPMECs and mouse lung.

A, HPMEC RNA obtained at 3, 12 and 24 h after LPS was used to quantify DLL4 expression by qRT‐PCR, P < 0.01 (Control vs. LPS at 3 h^*, 12 h^** and 24 h^***), n = 3. B, DLL4 protein in HPMECs was quantified by immunoblotting 24 h after LPS treatment, with or without TLR4‐blocking antibody (TLR4‐I), n ≥ 3. C, DLL4, IKK‐β phosphorylation [(p)IKKβ], and phosphorylated NF‐κB [(p)p65] in HPMECs were quantified 30 min after treatment with LPS, and with or without TLR4‐I, n ≥ 3. D, HPMEC cell death (%) was quantified by Trypan Blue staining 24 h after LPS treatment, P < 0.01 (^*Control vs. LPS), n = 3. E, cleaved caspase 3 protein level was quantified by immunoblotting HPMEC lysates 24 h after LPS treatment. F, total RNA obtained from whole lung of 7‐day old TLR4^+/+ and TLR4^−/− mice 24 h after i.p. saline or i.p. LPS treatment was used to probe DLL4 mRNA expression by qRT‐PCR. Fold change in expression is relative to TLR4^+/+ pups with saline injection, P < 0.05 (^*TLR4^+/+ vs. TLR4^+/+ LPS; ^**TLR4^+/+ LPS vs. TLR4^−/− LPS), n ≥ 4 mice per group. G, DLL4 protein expression in whole lung lysates of 7‐day old TLR4^+/+ and TLR4^−/− mice, n ≥ 3. H, DLL4 mRNA was quantified in mouse lung EC isolated at 6 and 24 h after i.p. LPS treatment by qRT‐PCR, P < 0.01 (Control vs. LPS at 6 h^* and 24 h^**), n ≥ 3 mice per group. I, DLL4 protein expression was quantified in mouse lung EC isolated 24 h after i.p. LPS, with densitometric quantification shown graphically (J), ^* P < 0.05 (Control vs. LPS), n ≥ 3. [Color figure can be viewed at http://wileyonlinelibrary.com]