Fig 1. Characterization of oxazolone-induced colitis model.

(A) Wild type BALB/c mice were sensitized by epicutaneous application of 3% oxazolone at a dilution of 4:1 in a mixture of acetone and olive oil (100μl) on day 0, followed by intracolonic administration of 1% oxazolone dissolved in 50% ethanol (100μl) (or 50% ethanol for control group EtOH-treated mice) on day 7 using an intravenous catheter inserted 3 cm in the colon. Mice were sacrificed 6, 24 hours and 5 days post oxazolone administration and tissue samples were collected for different analysis. (B) Survival proportion was assessed daily. Mortality is expressed as survival rate and shown by Kaplan-Meier survival curves; statistical significance of Kaplan-Meier survival curves was determined with Gehan-Breslow-Wilcoxon test. *p < 0.05; (n = 12–18 mice per group). (C) Repeated measurements of body temperature of EtOH- and oxazolone-treated mice were taken every 30 minutes and followed until 6 hours after treatment with oxazolone. Body temperature is shown as mean ± SEM, as determined by the repeated-measures two-way ANOVA test. ****p < 0.001; (n = 8 mice per group). (D) Bar graphs represent serum levels of HMGB1 in naïve, EtOH- and oxazolone-treated mice 6h post colitis induction as determined by the Mann-Whitney test. Ns, not significant; (n = 3–4 mice per group). (E) Paraffin-embedded colon sections were stained with hematoxylin and eosin (H&E) assessment of tissue alteration. Representative images of colonic sections stained with H&E from control naïve, EtOH- and oxazolone-treated mice at 6 and 24 hours after colitis induction. Scale bars are 40 μm in the upper panel and 100 μm in the lower panel. (F) Th1, chemokines and Th2 (G) gene expression was quantified in colonic tissue isolated from EtOH- and oxazolone-treated mice. Data are expressed as mean ± SEM of fold increase versus naïve mice as determined by the repeated-measures two-way ANOVA test. Ns, not significant; *p < 0.05; (n = 3–6 mice per group).