Fig. 8

TonEBP modulates LCN2 gene expression under inflammatory and hyperglycemic LTM/HG-treated conditions in vitro and in vivo. a Western-blot analysis of TonEBP and LCN2 in HT22 cells transfected with Scramble (Scr) or TonEBP or LCN2 siRNAs. b Expression of mRNA encoding TonEBP (F = 59.76, p < 0.001) and LCN2 (F = 290.1, p < 0.001) in LTM/HG-treated HT22 cells was measured by quantitative RT-PCR. c Representative immunofluorescence staining for TonEBP and LCN2 in LTM/HG-treated HT22 cells transfected with Scr or TonEBP or LCN2 siRNA. DAPI was used for nuclear staining. d, e Expression of mRNA encoding IL-6 (F = 84.22, p < 0.001) and TNF-α (F = 6.274, p = 0.017) (d), or Arg1 (F = 2.783, p = 0.11) and IL-10 (F = 56.45, p < 0.001) e in LTM/HG-treated HT22 cells was measured by quantitative RT-PCR. f A schematic drawing of the TonEBP binding motif in the mouse LCN2 promoter. g, h ChIP assay for TonEBP in the LCN2 promoter region as indicated in LPS/HG-treated RAW 264.7 macrophages g (F = 7.123, p = 0.028) or LTM/HG-treated HT22 h (F = 85.04, p < 0.001) cells. i, j ChIP assay for TonEBP at the − 2827 location in the LCN2 promoter in LPS/HG-treated RAW 264.7 macrophage i (F = 202.2, p < 0.001) or LTM/HG-treated HT22 j (F = 161.4, p < 0.001) cells transfected with TonEBP siRNA. k–l ChIP assay for TonEBP at the − 2827 location in the LCN2 promoter in WT mice k (F = 55.79, p < 0.001) or TonEBP ( ±) l (F = 12.94, p = 0.007). Data (n = 3) are shown as mean ± SEM. The indicated p values represent a one-way ANOVA in c, e, f, and l, or a two-way ANOVA in h, i, j, k, and m followed by Tukey’s post-hoc test. *p < 0.05 versus Scr or CTL or WT ND, ^†p < 0.05 versus LTM/HG-treated HT22 cells or WT HFD/STZ