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. 2010 Sep 13;116(25):5679–5687. doi: 10.1182/blood-2010-04-277954

Figure 5.

Figure 5

Hfe transgenes do not induce endoplasmic reticulum (ER) stress or the unfolded protein response (UPR). Dithiothreitol (DTT) induction (A-B) of the endoplasmic reticulum (ER) stress response in mouse embryonic fibroblast (MEF) cells. (A) Semiquantitative RT-PCR for unspliced (205 bp, uXbp-1) or spliced (179 bp, sXbp-1) Xbp-1 mRNA forms after treatment with increasing concentrations of DTT that induces ER stress. Total cell lysates (B) were analyzed for the KDEL motif-containing proteins Grp94 (Hsp90b1) and BiP (Hspa5) in MEF cells by Western blot. Equivalent loading of liver lysates was confirmed by immunoblot analysis for β-actin. ER stress in transgenic mouse livers was evaluated by measuring Xbp-1 mRNA splicing (C) using semiquantitative RT-PCR (n = 3, results presented as percentage spliced) and protein expression of BiP (D, n = 3), an ER-resident chaperone, relative to β-actin. Mean protein expression for Hfe−/− mice was set as 1.0 and all other data were expressed in relation to this. (D-E) All comparisons are statistically not significant.