The delay in the development of experimental colitis from isomaltosyloligosaccharides in rats is dependent on the degree of polymerization

Abstract

Background: Isomaltosyloligosaccharides (IMO) and dextran (Dex) are hardly digestible in the small intestine and thus influence the luminal environment and affect the maintenance of health. There is wide variation in the degree of polymerization (DP) in Dex and IMO (short-sized IMO, S-IMO; long-sized IMO, L-IMO), and the physiological influence of these compounds may be dependent on their DP.

Methodology/principal findings: Five-week-old male Wistar rats were given a semi-purified diet with or without 30 g/kg diet of the S-IMO (DP = 3.3), L-IMO (DP = 8.4), or Dex (DP = 1230) for two weeks. Dextran sulfate sodium (DSS) was administered to the rats for one week to induce experimental colitis. We evaluated the clinical symptoms during the DSS treatment period by scoring the body weight loss, stool consistency, and rectal bleeding. The development of colitis induced by DSS was delayed in the rats fed S-IMO and Dex diets. The DSS treatment promoted an accumulation of neutrophils in the colonic mucosa in the rats fed the control, S-IMO, and L-IMO diets, as assessed by a measurement of myeloperoxidase (MPO) activity. In contrast, no increase in MPO activity was observed in the Dex-diet-fed rats even with DSS treatment. Immune cell populations in peripheral blood were also modified by the DP of ingested saccharides. Dietary S-IMO increased the concentration of n-butyric acid in the cecal contents and the levels of glucagon-like peptide-2 in the colonic mucosa.

Conclusion/significance: Our study provided evidence that the physiological effects of α-glucosaccharides on colitis depend on their DP, linkage type, and digestibility.

Figure 1. Changes in DAI score and colon weight per length.

(A) DAI was calculated by the sum of three clinical scores (stool consistency, rectal bleeding, and weight loss) during the DSS treatment period. Significant differences comparing scores to values on day 0 (before DSS treatment) were determined by Dunnett’s multiple comparison test (**P<0.01, ***P<0.001). (B) Colon weight per length with or without DSS treatment. Two-way ANOVA P values for the colon weight per length (diet and treatment) were 0.0047 for diet, <0.0001 for treatment, and 0.0053 for the interaction between diet and treatment. No significant difference was observed for strain. Significant differences between the untreated group and the 2% DSS-treated group were determined by an unpaired two-tailed Student’s t-test (**P<0.01, ***P<0.001). Values are expressed as means ± SEM (n = 5–7).

Figure 2. MPO activity and histological appearance.

(A) MPO activity in the distal colon on day 7. Significant differences in activity between rats with or without DSS treatment were determined by an unpaired two-tailed Student’s t-test (*P<0.05, **P<0.01). Two-way ANOVA P values for MPO activity (diet and treatment) were <0.0001 for treatment. No significant difference was observed for treatment and the interaction between diet and treatment. Values are expressed as means ± SEM (n = 5–7). (B) Hematoxylin and eosin staining in a distal colonic section fixed with formalin. The scale bar represents 500 µm.

Figure 3. Inflammatory cytokine expression in distal colonic mucosa.

The mRNA expression of IL-1β (A) and IFN-γ (B) was evaluated in the distal colon of rats with or without DSS treatment using a TaqMan gene expression assay. GAPDH was used as the internal control. Significant differences between rats with or without DSS treatment were determined by a median test (*P<0.05). Two-way ANOVA P values for IL-1β (A) and IFN-γ mRNA expression (diet and treatment) were 0.0042 and 0.0087 for treatment, respectively. No significant difference was observed for treatment and the interaction between diet and treatment. Values are expressed as means ± SEM (n = 6).

Figure 4. Concentrations of organic acids in cecal content.

Organic acids (succinic, lactic, acetic, propionic, iso-butyric, n-butyric, iso-valeric and n-valeric acids) were measured via HPLC in the homogenates of cecal contents of (A) untreated or (B) DSS-treated rats. Significant differences in each organic acid were determined by Dunnett’s multiple comparison test (**P<0.01, ***P<0.001). Values are expressed as means ± SEM (n = 6).

Figure 5. Concentrations of GLP-1 and GLP-2 in ileal and colonic mucosa.

Active GLP-1 concentrations in the mucosa of (A) the terminal ileum and (B) the proximal colon were measured using a GLP-1 ELISA kit. Total GLP-2 concentrations in the mucosa of (C) the terminal ileum and (D) the proximal colon of rats treated with DSS were measured using a GLP-2 ELISA kit. Significant differences were determined by Dunnett’s multiple comparison test (*P<0.05, **P<0.01). Values are expressed as means ± SEM (n = 5–6).

Figure 6. Phenotypic analysis of PBL.

(A) Representative dot plots of phenotypes in PBL expressing CD8α and CD161 molecules in rats fed the control diet. (B) The percentage of CD8⁺ CD161⁻, CD8⁻CD161⁺, and CD8⁺CD161⁺ cells in the CD45⁺ cell populations. Significant differences between rats with or without DSS treatment were determined by an unpaired two-tailed Student’s t-test (*P<0.05, ***P<0.001). Values are expressed as means ± SEM (n = 3–5).