1. Mesalamine blocks tumor necrosis factor growth inhibition and nuclear factor κB activation in mouse colonocytes 
Greg C. Kaiser, Fang Yan, D.Brent Polk 
Gastroenterology 
Volume 116, Issue 3, Pages (March 1999)
DOI: /S (99)
Copyright © 1999 American Gastroenterological Association Terms and Conditions

2. Fig. 1
The antiproliferative effect of high-dose TNF-α is inhibited by mesalamine. YAMC monolayers were serum-starved at nonpermissive conditions for 24 hours before use. Cells were incubated for 24 hours at 37°C with EGF and/or TNF-α in the presence or absence of mesalamine (5-ASA) pretreatment. Cell numbers were determined as described in Materials and Methods. *P < compared with control; ΦP < compared with 100 ng/mL TNF-α; ωP < compared with 10 ng/mL EGF; ΣP < compared with 1 ng/mL TNF-α and 10 ng/mL EGF; and ξP < compared with 100 ng/mL TNF-α and 10 ng/mL EGF.
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

3. Fig. 2
TNF-α–stimulated ERK1/ERK2 activation is blocked by mesalamine. YAMC cells were incubated with EGF for 2 minutes or TNF-α for 15 minutes in the presence or absence of mesalamine (5-ASA) at 37°C. Triton-soluble lysates were obtained, proteins separated by SDS-PAGE, and transferred to a polyvinylidene difluoride membrane, and Western blot analysis was conducted with an antiactive ERK1/ERK2 antibody (top panel) and an anti–pan ERK1/ERK2 antibody (lower panel) as described in Materials and Methods. Proteins were visualized by enhanced chemiluminescence.
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

4. Fig. 3
Inhibition of TNF-α–stimulated ERK1/ERK2 and JNK/SAPK activation by mesalamine is both concentration and time dependent. In the top panels, YAMC cells were incubated with various concentrations of mesalamine (5-ASA), then cultured with TNF-α for 15 minutes at 37°C. (A) Antiactive ERK1/ERK2 was detected as described in Figure 2. (B) Antiactive JNK/SAPK was identified by Western blot analysis of Triton-soluble lysates as described in Materials and Methods.
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

5. Fig. 4
ERK1/ERK2 activation by ceramide is blocked by mesalamine. YAMC cells were treated with TNF-α or ceramide in dimethyl sulfoxide for 15 minutes at 37°C in the presence or absence of mesalamine (5-ASA). Antiactive ERK1/ERK2 was detected as described in Figure 2.
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

6. Fig. 5
TNF-α–stimulated nuclear translocation of NF-κB is inhibited by mesalamine. YAMC cells were grown on collagen-coated Thermanox culture slides at 37°C and received (A) no treatment, (B) 100 ng/mL TNF-α for 30 minutes, (C) 20 mmol/L mesalamine for 30 minutes, or (D) the combination of TNF-α and mesalamine. Cells were then fixed, probed with an anti–NF-κB p65 antibody, and incubated with a secondary antibody linked to Cy3 as described in Materials and Methods. Images were obtained using confocal laser immunofluorescent microscopy.
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

7. Fig. 6
Iκ-Bα degradation is inhibited by mesalamine. YAMC cells were treated with 100 ng/mL TNF-α for various times at 37°C as indicated in the presence or absence of (A) mesalamine (5-ASA) or (B) treated with TNF-α for 15 minutes in the presence of mesalamine at various concentrations. Cytosolic fractions were prepared; proteins were separated by SDS-PAGE and transferred to a polyvinylidene difluoride membrane; and Western blot analysis was conducted with an anti–Iκ-Bα antibody as described in Materials and Methods. Proteins were visualized by enhanced chemiluminescence.
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

8. Fig. 7
TNF-α–stimulated degradation of IκB and activation of MAP kinase are not affected by pH nor osmolarity. YAMC cells were treated with TNF-α for 15 minutes alone or in combination with a 30-minute pretreatment with mesalamine (5-ASA), equivalent osmotic concentrations of mannitol, equivalent pH-corrected media with butyric acid, or the combined osmotic and pH effects or various concentrations of 4-aminosalicylic acid at 37°C. Cell lysates were prepared for Western blot analysis with anti-IκB or antiactive ERK1/ERK2 or anti-ERK1/ERK2, as indicated.
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions