Tunicamycin-induced ER stress reduces 11β-HSD1 expression in 3T3-L1 adipocytes.

<p>(<b>A–E</b>) Real-time PCR measurement of levels of mRNA encoding (<b>A</b>) 11β-HSD1, (<b>B</b>) C/EBPβ, (<b>C</b>) CHOP, (<b>D</b>) calreticulin and (<b>E</b>) GRP78 in untreated differentiated 3T3-L1 adipocytes (white bars) or treated with tunicamycin (4 µg/ml, black bars) for 6 or 16 h. Data are normalized to TBP and expressed relative to levels of control cells (arbitrarily set to 100%). (<b>F</b>) Representative western blot (40 µg protein/lane) and quantification showing levels of C/EBPβ-LAP (38 kDa LAP* +35 kDa LAP) and -LIP (20 kDa) isoforms. Inset shows C/EBPβ-LIP:LAP ratio in untreated 3T3-L1 adipocytes (C, white bar) or treated with tunicamycin for 0.5 h (0.5 h, grey bar) or 2 h (2 h, black bar). Blots were stripped and reprobed with β-tubulin antibody, as loading control. All samples were analysed in the same gel but not in adjacent lanes. C/EBPβ levels were quantified relative to β-tubulin and are expressed in arbitrary units (AU). Values are mean±SEM of 2–3 independent adipocyte differentiations with each treatment tested in duplicate or triplicate. *, Significantly different from control. *, p≤0.05; **, p<0.001; ***, p<0.0001.