CD14 is required for LNFPIII-NGC induced ERK and NFκB activation.

<p>a) HEKTLR4_AP cells were pretreated with anti-CD14 antibody or isotype control for 90 mins at 37˚C followed by stimulation with LPS (100ng/ml), LNFPIII (50μg/ml), Dex (50μg/ml) or media control for 20–24 hrs at 37˚C. The supernatants were collected and levels of alkaline phosphatase were determined. Statistical significance was calculated using GraphPad student’s unpaired <i>t</i> test. b-c) RAW 264.7 cells were treated with anti-CD14 antibody or isotype control for 90 mins at 37°C followed by stimulation with LPS (100ng/ml), LNFPIII (50μg/ml), Dex (50μg/ml) or media alone for 60 min. The nuclear extracts or the whole cell lysate was then fractionated using SDS-PAGE and probed with antibody against NFκB subunit p50 (b) or phospho-ERK1/2 (c) and the blots were re-probed with anti-TFIID (b) or anti-total-ERK antibodies (c). Bar graph shows the ratio of intensities of the pERK and the control (tERK) bands. d) BMDCs from WT (C57BL/6J) or CD14-/- knock out (C57BL/6J) were stimulated with LPS (100ng/ml), LNFPIII (50μg/ml), Dex (50μg/ml) for 60 min. The whole cell lysate was then fractionated using SDS-PAGE and probed with anti-pERK antibody and the blot was re-probed with anti-total ERK antibody.