FIGURE 6.
KLF6 regulates macrophage polarization by modulating functions of NF-κB. A, wild-type BMDMs were stimulated with 10 ng/ml LPS, and KLF6 ChIP analysis was performed on IL-1α (−976 to −980) and IL-1β (−601 to −605) promoters. Fold changes in KLF6 enrichment over control are indicated. ChIP analysis performed using IgG was used as negative control. B, RAW264.7 cells were co-transfected with NF-κB concatemer luciferase reporter plasmid in the presence of KLF6 or NF-κB-p65 plasmid. These cells were stimulated with LPS, and induction in luciferase was documented and is indicated as relative luciferase activity over the control group. C and D, RAW264.7 cells overexpressed with Klf6 and Lyz2cre, Lyz2cre:Klf6fl/fl mouse BMDMs were induced with 10 ng/ml LPS. ChIP analysis was performed on IL-1α promoter (−1787 to −1796) utilizing anti-p65 and anti-p300 antibody. ChIP analysis performed using IgG was used as negative control. E, RAW264.7 cells were co-transfected with Klf6 or NF-κB-p65 plasmid. These cells were stimulated with LPS, and total RNA from these cells was isolated. IL-1α mRNA expression was analyzed by quantitative PCR and normalized to 36B4. F, RAW264.7 cells were transfected with Klf6 plasmid. These cells were stimulated with LPS in the presence of NF-κB peptide inhibitor SN-50, and total RNA from these cells was isolated. IL-1α mRNA expression was analyzed by quantitative PCR and normalized to 36B4.