Dietary Supplementation with Nondigestible Oligosaccharides Reduces Allergic Symptoms and Supports Low Dose Oral Immunotherapy in a Peanut Allergy Mouse Model

Abstract

Scope: A major downside of oral immunotherapy (OIT) for food allergy is the risk of severe side effects. Non-digestible short- and long-chain fructo-oligosaccharides (scFOS/lcFOS) reduce allergy development in murine models. Therefore, it is hypothesized that scFOS/lcFOS can also support the efficacy of OIT in a peanut allergy model.

Methods and results: After sensitization to peanut extract (PE) using cholera toxin, C3H/HeOuJ mice are fed a 1% scFOS/lcFOS or control diet and receive OIT (1.5 or 15 mg PE). Hereafter, mice are exposed to PE via different routes to determine the safety and efficacy of treatment in clinical outcomes, PE-specific antibody production, and numbers of various immune cells. scFOS/lcFOS increases short-chain fatty acid levels in the caecum and reduce the acute allergic skin response and drop in body temperature after PE exposure. Interestingly, 15 mg and 1.5 mg OIT with scFOS/lcFOS induce protection against anaphylaxis, whereas 1.5 mg OIT alone does not. OIT, with or without scFOS/lcFOS, induces PE-specific immunoglobulin (Ig) IgG and IgA levels and increases CD103+ dendritic cells in the mesenteric lymph nodes.

Conclusions: scFOS/lcFOS and scFOS/lcFOS combined with low dose OIT are able to protect against a peanut-allergic anaphylactic response.

Keywords: mouse model; non-digestible oligosaccharides; oral immunotherapy; peanut allergy.

Figure 1

Schematic overview of the experimental set‐up. CT, cholera toxin; i.d., intradermal; i.g., intragastric; i.p., intraperitoneal; OIT, oral immunotherapy; PE, peanut extract.

Figure 2

Allergic manifestations evaluated in PE‐sensitized mice after having received OIT or OIT + scFOS/lcFOS (FF). A) Change in body temperature after intraperitoneal challenge on day 77. B) Anaphylactic shock symptom scores determined 40 min after intraperitoneal challenge on day 77. C) Acute allergic skin response measured as Δ ear swelling 1 h after intradermal challenge on day 64. D) Concentrations of MMCP‐1 in serum collected 30 min after intragastric challenge on day 70. E) Peripheral blood basophil activation was measured after whole blood stimulation with αIgE on day 68. Data are represented as mean ± SEM n = 5/6 mice/group. Statistical analysis was performed using repeated measures two‐way ANOVA and Bonferroni's post hoc test (body temperature), one‐way ANOVA, and Bonferroni's post hoc test to compare preselected combinations (ear swelling, MMCP‐1, and basophil activation) or a Kruskal–Wallis test with Dunn's post hoc test (clinical score). ^# p < 0.05; compared to sham‐sensitized control; *p < 0.05 compared to PE‐sensitized control. FF, scFOS/lcFOS dietary supplementation; i.p., intraperitoneal; IT, immunotherapy; MMCP‐1, mast cell protease‐1; OIT, oral immunotherapy; PE, peanut extract.

Figure 3

PE‐specific IgE, IgA, IgG1, and IgG2a levels in serum determined by ELISA and IgA and IgG1 production by splenocytes determined by ELISPOT. Blood was taken on day 35, 50, 63, 70, and 78. A,B) Allergen‐specific IgE and IgA measured in serum by ELISA. C,D) Allergen‐specific IgG2a and IgG1 measured in serum by ELISA. E,F) Number of allergen‐specific IgA and IgG1 antigen secreting splenocytes (ASCS) per spleen. Data are represented as mean ± SEM n = 5/6 mice/group. Statistical analysis of the antibody levels was performed on each individual time point, after log transformation, using one‐way ANOVA and Bonferroni's post hoc test for multiple comparisons. All treatment groups were compared to the sensitized control group and significant differences were indicated with letters: Letters used: s for sham‐sensitized control; a for no OIT plus scFOS/lcFOS; b for 1.5 mg OIT; c for 1.5 mg OIT plus scFOS/lcFOS; d for 15 mg OIT; and e for 15 mg OIT plus scFOS/lcFOS when p < 0.05. Statistical analysis for the ELISPOT results was performed on each time point using one‐way ANOVA and Bonferroni's post hoc test for multiple comparisons. ^# p < 0.05 compared to sham‐sensitized control; *p < 0.05 compared to PE‐sensitized control; ^p < 0.05 compared to scFOS/lcFOS control. FF, scFOS/lcFOS dietary supplementation; id, intradermal challenge; ig, intragastric challenge; ip, intraperitoneal challenge; OIT, oral immunotherapy; PE, peanut extract.

Figure 4

Flow cytometric analysis of DC and T‐cell populations in the MLN. Cells were gated based on FSC–SSC properties and the fluorescence‐minus‐one (FMO) technique. For DC, the live cells were gated on: CD64 negative, MHC Class II positive and CD11c positive. The CD11c/MHCII positive population was further characterized on the basis of CD103 and CD11b expression. A,B) Percentage of CD103+CD11b+ DC and percentage of CD103+CD11b‐DC. C) Percentage of regulatory T cells (CD25+FoxP3+ of CD4+) and D) activated TH₁ cells (CXCR3+CD69+ of CD4+). All data are represented as mean ± SEM n = 5/6 mice/group. Statistical analysis was performed for each time point using a one‐way ANOVA and Bonferroni's post hoc test. *p < 0.05 compared to PE‐sensitized control; ^p < 0.05; compared to scFOS/lcFOS control; ^$ p < 0.05 compared to the OIT control diet group. FF, scFOS/lcFOS dietary supplementation; IT, immunotherapy; OIT, oral immunotherapy; PE, peanut extract.

Figure 5

Levels of total and individual short chain fatty acids (SCFA). A) Mean total SCFA content (mmol L⁻¹). B–E) Butyric, acetic, propionic, and valeric acid content (mmol L⁻¹). All data are represented as mean ± SEM n = 5/6 mice/group. Statistical analysis was performed for each time point using one‐way ANOVA and Bonferroni's post hoc test for multiple comparisons to compare preselected combinations. *p < 0.05 compared to PE‐sensitized control; ^p < 0. 05 compared to scFOS/lcFOS control; ^$ p < 0.05 compared to the OIT alone group. FF, scFOS/lcFOS dietary supplementation; IT, immunotherapy; OIT, oral immunotherapy; PE, peanut extract; SCFA, short‐chain fatty acids.