1. Volume 141, Issue 3, Pages 1024-1035 (September 2011)
Altered Endoplasmic Reticulum Stress Affects Translation in Inactive Colon Tissue From Patients With Ulcerative Colitis 
Xavier Tréton, Eric Pédruzzi, Dominique Cazals–Hatem, Alain Grodet, Yves Panis, André Groyer, Richard Moreau, Yoram Bouhnik, Fanny Daniel, Eric Ogier–Denis 
Gastroenterology 
Volume 141, Issue 3, Pages (September 2011)
DOI: /j.gastro
Copyright © 2011 AGA Institute Terms and Conditions

2. Figure 1
Extended IRE1β and ATF6α branch signaling of ER stress in unaffected UC mucosa. (A) The mRNA levels of spliced (XBP-1s) and unspliced (XBP-1u) XBP-1 forms, GRP78, GRP94, and EDEM1 were determined by quantitative polymerase chain reaction from unaffected UC (n = 12 individual data points shown) and control (Ctrl) (n = 10 individual data points shown) mucosa. Mann–Whitney test (*P < .05, **P < .01 vs control). (B) Representative immunoblot analysis of GRP94, GRP78, and ATF6α expression in normal and unaffected colon biopsy samples from 2 controls (Ctrl) and 5 UC patients, respectively. β-actin served as a loading control, and densitometric analyses are shown. Individual data points from normal mucosa of 10 controls and unaffected mucosa of 12 UC patients are shown. Statistics as in panel A. (C) Representative immunohistologic analysis of GRP78 in unaffected colon sections from 11 controls (Ctrl) and 12 UC patients. Scale bars, 80 μm. Magnification of the micrographs shows increased immunostaining of GRP78 in unaffected UC mucosa; scale bars, 50 μm. (D) Ultrastructural evidence of ER stress in the unaffected colon epithelium of patients with UC. Representative transmission electron micrographs from the unaffected colon of 8 controls (Ctrl) and 8 UC patients. Scale bars, 5 μm (upper panels) and 1 μm (lower panels). Inset: The bars represent the mean values of ER cell surface area, expressed as square micrometers ± standard deviation counted in 20 different fields of electron micrographs. **P < .01 vs control values. T, thecae.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2011 AGA Institute Terms and Conditions

3. Figure 2
Aberrant attenuation of the eIF2α-dependent integrated stress response in unaffected colonic mucosa from patients with UC. (A) Representative immunoblot analysis using anti-P-eIF2α (Ser51) and anti-eIF2α antibodies were performed on normal and unaffected colon biopsy specimens from 2 controls (Ctrl) and 5 UC patients, respectively. β-actin served as a loading control. The ratio P-eIF2α/eIF2α was depicted, and individual data points from normal mucosa of 10 controls and unaffected mucosa of 12 UC patients are shown. Mann–Whitney test (***P < .001 vs control). (B) The mRNA levels of ATF4, CHOP, and GADD34 were determined by quantitative polymerase chain reaction and expressed relative to β-actin from unaffected UC (n = 12) and control (Ctrl) (n = 10) mucosa. Mann–Whitney test (*P < .05, **P < .01 vs control). (C) Representative immunoblot analysis using anti-ATF4, anti-CHOP, and anti-GADD34 antibodies were performed on normal and unaffected colon biopsy specimens from 2 controls (Ctrl) and 5 UC patients, respectively. β-actin served as a loading control, and densitometric analyses are shown. Individual data points from normal mucosa of 10 controls and unaffected mucosa of 12 UC patients are shown. Mann–Whitney test (**P < .01, ***P < .001 vs control).
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2011 AGA Institute Terms and Conditions

4. Figure 3
Stress granule formation is altered in unaffected UC mucosa. (A) Double indirect immunofluorescence of colon sections from normal and unaffected UC mucosa stained with antibodies against TIA-1 (green) and eIF3 (red). Nuclei (blue) were stained with 4′,6-diamidino-2-phenylindole or TO-PRO-3 iodide. Original magnification, ×40. Photomicrographs are representative sections from 15 UC patients and 10 controls. Inset boxes are enlarged views showing separate and merged color channels. Original magnification, ×60. Inset: Quantification of SGs in unaffected UC and normal (Ctrl) colonic mucosa was determined by counting the number of epithelial cells per millimeter of colon epithelium that contained 5 or more SGs. Mann–Whitney test (***P < .001 vs control). (B) Representative immunoblot analysis using anti-TIA-1 and anti-eIF3 antibodies were performed on normal and unaffected colon biopsy specimens from 2 controls (Ctrl) and 4 UC patients, respectively. β-actin served as a loading control, and densitometric analyses are shown. Individual data points from the normal mucosa of 10 controls and unaffected mucosa of 12 UC patients are shown. Mann–Whitney test (*P < .05).
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2011 AGA Institute Terms and Conditions

5. Figure 4
Increased expression of the translation initiation factors in unaffected UC mucosa and microarray analysis of polysomal mRNAs (A) Representative immunoblot analysis using anti-P-4E-BP1 (Ser65), anti-4E-BP1, anti-P-eIF4E (Ser209), and anti-eIF4E antibodies was performed on normal and unaffected colon biopsy specimens from 2 controls (Ctrl) and 3 UC patients, respectively. The ratios of P-4E-BP1/4E-BP1 and P-eIF4E/eIF4E were reported, and individual data points from normal mucosa of 10 controls and unaffected mucosa of 12 UC patients are shown. Mann–Whitney test (*P < .05, **P < .01 vs control). (B) Representative immunoblot analysis using anti-eIF4A and anti-eIF4G antibodies was performed on normal and unaffected colon biopsy extracts from 2 controls (Ctrl) and 3 UC patients, respectively. β-actin was used as a loading control, and densitometric analyses are shown. Individual data points from the normal mucosa of 10 controls and 12 unaffected UC mucosa are shown. Statistics as in panel A. (C) Representative polysome profiles in lysates from colon mucosa of controls and inactive UC patients. RNA bound to polysomes were subjected to microarray analysis. Three experiments were performed from pooled unaffected mucosa from 6 patients with UC and pooled normal mucosa from 6 healthy subjects for each experiment. (D) Gene ontology analysis that indicated >2- or <2-fold change at P < .05 (Student t test with Benjamini and Hochberg false discovery rate as multiple testing correction) in the polysome fractions of unaffected UC mucosa compared with controls.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2011 AGA Institute Terms and Conditions

6. Figure 5
Validation of 6 candidate genes identified by microarray analysis in unaffected UC and control mucosa. (A) Representative immunoblot analysis of p21WAF/Cip, eIF5A, RanBP2, and SOD1 expression in normal and unaffected colon biopsy specimens from 2 controls (Ctrl) and 3 UC patients, respectively. β-actin served as a loading control, and densitometric analyses are shown. Individual data points from the normal mucosa of 6 controls and the unaffected mucosa of 6 patients with UC and P values from Mann–Whitney test are shown. (B) Representative immunoblot analysis of PIgR and TST/CES2 in normal and unaffected colon biopsy specimens from 2 controls (Ctrl) and 3 UC patients, respectively. β-actin served as a loading control, and densitometric analyses are shown. Individual data points from the normal mucosa of 6 controls and the unaffected mucosa of 6 UC patients and P values from Mann–Whitney tests are shown. Immunohistologic analysis of PIgR and TST/CES2 in colon sections from the normal mucosa of 8 controls (Ctrl) and the unaffected mucosa of 8 patients with UC. Scale bars, 80 μm. Magnification of the micrographs shows decreased immunostaining of PIgR and TST/CES2 in unaffected UC mucosa; scale bars, 50 μm.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2011 AGA Institute Terms and Conditions