Figure 7
MBL decreases LPS-stimulated NF-κB binding activity and translocation in imMDCs. (A) LPS-induced DNA-binding activity of NF-κB is inhibited by MBL. ImMDCs (5 × 105 cells/sample) were stimulated with LPS (100 ng/ml) in the presence of 15 μg/ml of HSA, anti-MBL pAb and MBL, or MBL for 1 h, then harvested to prepare nuclear extracts. The nuclear extracts were mixed with radiolabeled NF-κB oligonucleotide probe and analyzed with EMSA. (B) MBL inhibits NF-κB translocation in imMDCs. ImMDCs (5 × 105 cells/sample) were stimulated with LPS (100 ng/ml) in the presence of 15 μg/ml HSA, anti-MBL pAb and MBL, or MBL for 1 h, then the cells were harvested to prepare nuclear extracts. The proteins in the nuclei-free supernatants were separated by 10% SDS-PAGE, followed by transfer to a nitrocellulose membrane. After blocking, the membrane was incubated with the NF-κB-specific mouse anti-human mAb p65, followed by HRP-conjugated secondary antibody. ECL was used to visualize the protein bands. As an internal control, actin was used.