Mouse colon size changes during DSS treatment.

<p>C57BL/6 mice were exposed to 3.5% DSS in their drinking water for indicated days. Colon length removed from euthanized mice is depicted as means±SEM length of the colon in each group. * P<0.05, ** P<0.01.</p

Hematoxylin-stained colon sections of mice treated with DSS at 0, 1, 3, 5, 9 (withdraw after 5 days treatment, 4 days to recover) and 14 (withdraw after 5 days treatment, 9 days to recover) days, with magnification of 20 times.

<p>(A): Distal colon; (B) Proximal colon.</p

DSS can increase the production significantly of proinflammatory mediators TNF-α, IL-1β, IL-6, IL-10, IL-12, IFN-γ and chemokines KC and MIP-2.

<p>This stimulation happened very early with the treatment, TNF-α, IL-1β, IL-10, IL-12, and IFN-γ production increased as early as the first day. The production of these cytokines reached to the peak at the day 5 of DSS treatment, then decreased after the DSS withdrawal, but all of them are still high significantly than those in control mice. * P<0.05, ** P<0.01.</p

Determination of MPO enzymatic activity as an index of neutrophils infiltration into the injured tissue.

<p>(A): Distal colon; (B): Proximal colon. Results are expressed as MPO mUnits per µg protein and represent mean±SEM of 3 determinations. * P<0.05, ** P<0.01.</p

<i>In vivo</i> permeability assay in colon epithelium of mice treated with DSS.

<p>Results are presented as FITC-dextran in mg/µg protein and represent mean±SEM of 4 determinations. * P<0.05, ** P<0.01.</p

Mouse body weight changes during DSS treatment.

<p>C57BL/6 mice were administered to 3.5% DSS by drinking water for indicated days. Body weight changes are depicted as means±SEM body weight changes in each group. * P<0.05, ** P<0.01.</p

5 days of DSS treatment stimulates the production of proinflammatory cytokines differently in proximal and distal parts of colon.

<p>After DSS treatment, the production of cytokines TNF-α, IL-1β, IL-6, IL-10, IL-12, IFN-γ and chemokines KC and MIP-2 increases both in proximal and distal parts of colon. But the increasing folds of TNF-α is higher in proximal colon than that in distal colon, the increasing folds of IL-1β, IL-10 and KC are almost same in both proximal and distal colon. There are higher folds increases in distal colon than in proximal colon for IL-6, IL-12, IFN-γ and chemokine MIP-2. But the normalized cycles thresholds (NCT) are always lower in distal colon than in proximal colon which means higher absolute production of these mediators in distal part than in proximal parts of the colon. I: Water drinking mice proximal colon, II: DSS treated mice proximal colon, III: Water drinking mice distal colon, IV: DSS treated mice distal colon. * P<0.05, ** P<0.01.</p

Dextran-loaded nanoparticles but not free dextran induce colitis.

<p>Mouse body weight changes during treatment with DSS supplemented with dextran-loaded NPs (“dextran-loaded NPs”) or empty NPs (“DSS 3%”) compared to those of control mice gavaged with empty NPs suspended in water. C57BL/6 mice drank a 3% (w/v) DSS solution, or water, for the indicated numbers of days. (A) Body weight changes are shown as means±SEMs. * P<0.05. (B) Colon length (mm) of DSS-treated mice given dextran-loaded or empty NPs, compared to that of control animals (drinking water only). * P<0.05. (C) Determination of MPO enzymatic activity as an index of neutrophil infiltration into injured tissue. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0032084#s3" target="_blank">Results</a> are expressed as units of MPO per µg of protein, are compared to the values in control mice, and are means±SEMs of three independent determinations. ** P<0.01, *** P<0.001. (D) Dextran-loaded NPs can significantly increase production of IL-1β and IL-6, compared to use of empty NPs, in animals drinking DSS (3% w/v). mRNA expression was measured in test and control animals. * P<0.05. (E) Hematoxylin-stained colon sections of mice receiving daily gavage with dextran-loaded or empty NPs (“DSS”) in the interval during which the drinking water contained DSS (3% w/v); thus on day 7. (F) Histological scores. (G) Macroscopic inflammation was assessed using a mouse colonoscope. Photographs were obtained on the day of sacrifice and, (H), endoscopic scores were calculated.</p

DSS induces colitis.

<p>(A) Mouse body weight changes during DSS treatment compared to that of control animals (drinking water). C57BL/6 mice were exposed to 3% (w/v) DSS in the drinking water for the indicated numbers of days. Body weight changes are shown as means±SEMs. * P<0.05, ** P<0.01. (B) Hematoxylin-stained colonic sections of mice treated with DSS; the mice were sacrificed on day 8. (C) Associated histological scores. *** P<0.001. (D) Macroscopic inflammation was assessed using a mouse colonoscope. Photographs were obtained on the day of sacrifice and (E) an endoscopic score was calculated. *** P<0.001. (F) KC mRNA expression was measured in test and control animals. *** P<0.001. (G) Determination of MPO enzymatic activity as an index of neutrophil infiltration into injured tissue. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0032084#s3" target="_blank">Results</a> are expressed as MPO –fold increases compared to those of control mice and represent means±SEMs of three independent determinations. *** P<0.01.</p

DSS and DSS-associated molecules affect luminal intestinal osmolarity.

<p>(A) Macroscopic observations of the effects of DSS-associated molecules [DSS = dextran sodium sulfate, D+S = dextran (1.49% w/v) in sulfate solution (2.6% w/v), D = dextran (1.49% w/v), S = sulfate solution (2.6% w/v), and Glc+S = glucose (1.73% w/v) in sulfate solution (2.6% w/v)] on colonic epithelium were assessed using a mouse colonoscope. Photographs were obtained from all treatment groups on the day of sacrifice. (B) Fecal relative osmolarity values (means±SEM, ANOVA statistical test, * P<0.05).</p