Fig. 4

OMVs induce proinflammatory signaling cascades in human lung ECs. A) Human lung microvascular ECs (HULEC-5a) were stimulated with SEC-purified KpOMVs (MOV₁₀₀₀) or left untreated as control (Ctrl). Expression and phosphorylation of IRAK-1, p38 and STAT1 after OMV exposure for up to 180 min were determined by Western Blot. β-actin served as a loading control. B) HULEC-5a were stimulated for 180 min with Kp or cCOMVs (MOV₁₀₀₀) alone or in combination with PB (20 µg/ml) or left untreated for Ctrl. Expression and phosphorylation of IRAK-1 and p38 was determined by Western Blot. β-actin served as a loading control. C) HULEC-5a were incubated with KpOMV for up to 45 min or D) incubated with KpOMV for 30 min with or without 1 h pretreatment with PB (20 µg/ml). Samples were fractionated into cytosolic and nuclear proteins and analyzed by Western Blot for p65 and p38. Tubulin served as a cytoplasmic loading control, while Lamin A/C served as a nuclear loading control. One representative result of three biological independent replicates is shown