FIG. 3.

SirT1 knockdown in WAT results in NF-κB nuclear localization and gene expression through reduction of H3K9 deacetylation. A and B: Representative (n = 5/group) images showing that SirT1 knockdown stimulates nuclear translocation of NF-κB in rat WAT. Adipocytes are stained for caveolin (green) and NF-κB p65 nuclear localization sequence (red). Nuclei are stained with DAPI (blue). Inset box is expanded to individual channels in B.1-B.3 to demonstrate NF-κB and DAPI colocalization. Some NF-κB–positive nuclei (arrows) are in adipocytes. Because the nuclear localization sequence is masked in the cytosol, nonnuclear staining is nonspecific (i.e., erythrocyte or other autofluorescence). C: SirT1 knockdown increases abundance of phosphorylated (Ser536) and total p65 in rat WAT nuclear lysates. Line separates noncontiguous lanes from the same blot. D: Chow-fed fat-specific SirT1 knockout mice have increased phospho-p65 in WAT ∼12 h after stimulation with LPS (50 μg i.p.). E: SirT1 knockdown increases adipose tissue cytokine, complement, and TLR mRNA expression (n = 5–7/group). *P < 0.05, **P < 0.01. F: SirT1 knockdown increases H3K9 acetylation. (A high-quality digital representation of this figure is available in the online issue.)