Figure 3.

MDX up-regulates the IRE1-dependent UPR pathway in intestinal epithelial cells. (A) Volcano plot and heat map showing microarray-based differential expression, log₂ (fold change) of genes related to lipid and carbohydrate metabolism, protein glycosylation, and UPR pathway of colon samples isolated from mice exposed to drinking water in the presence or absence of 5% MDX for 45 days without DSS. (B) Ern-2, Ern-1, and Xbp1s RNA expression was assessed by real-time PCR in total colonic samples. Data were generated using 11–12 mice per group from 3 independent experiments (*P ≤ .05, **P ≤ .01). Each point in the graph indicates the RNA expression of the specific transcript in the colon of a single mouse; horizontal bars indicate median value. Differences among groups were compared using the Mann–Whitney U test. (C) Scatter plots showing Grp78, Atf6, Atf4, and Chop RNA transcripts in colonic samples isolated from mice treated as indicated in panel A. Data were generated using 7–8 mice per group of 3 independent experiments. Each point in the graph indicates the RNA expression of the specific transcript in the colon of a single mouse; horizontal bars indicate median value or means ± SD. Differences among groups were compared using the Mann–Whitney U test or the 2-tailed Student t test. (D) Representative histograms showing Ern-2 and Ern-1 RNA expression in IECs and LPMCs isolated from a pool of 3–4 mice treated as indicated in panel A. The example is representative of 2 independent experiments in which similar results were obtained. (E) Scatter plots showing Ern-2, Ern-1, and Xbp1s RNA expression in intestinal crypts isolated from the colons of untreated mice and cultured in the presence or absence of MDX for 30 minutes. Data were generated using crypts isolated from 6–7 mice from 3 independent experiments (*P ≤ .05, **P ≤ .01). Left: Horizontal bars indicate median value and differences between groups were compared using 2-tailed Mann–Whitney U test. Middle and right: Horizontal bars indicate means ± SD and differences between groups were compared using the Student t test. mRNA, messenger RNA; UT, untreated.