Genetic diversity between mouse strains allows identification of the CC027/GeniUnc strain as an orally reactive model of peanut allergy

Abstract

Background: Improved animal models are needed to understand the genetic and environmental factors that contribute to food allergy.

Objective: We sought to assess food allergy phenotypes in a genetically diverse collection of mice.

Methods: We selected 16 Collaborative Cross (CC) mouse strains, as well as the classic inbred C57BL/6J, C3H/HeJ, and BALB/cJ strains, for screening. Female mice were sensitized to peanut intragastrically with or without cholera toxin and then challenged with peanut by means of oral gavage or intraperitoneal injection and assessed for anaphylaxis. Peanut-specific immunoglobulins, T-cell cytokines, regulatory T cells, mast cells, and basophils were quantified.

Results: Eleven of the 16 CC strains had allergic reactions to intraperitoneal peanut challenge, whereas only CC027/GeniUnc mice reproducibly experienced severe symptoms after oral food challenge (OFC). CC027/GeniUnc, C3H/HeJ, and C57BL/6J mice all mounted a T_H2 response against peanut, leading to production of IL-4 and IgE, but only the CC027/GeniUnc mice reacted to OFC. Orally induced anaphylaxis in CC027/GeniUnc mice was correlated with serum levels of Ara h 2 in circulation but not with allergen-specific IgE or mucosal mast cell protease 1 levels, indicating systemic allergen absorption is important for anaphylaxis through the gastrointestinal tract. Furthermore, CC027/GeniUnc, but not C3H/HeJ or BALB/cJ, mice can be sensitized in the absence of cholera toxin and react on OFC to peanut.

Conclusions: We have identified and characterized CC027/GeniUnc mice as a strain that is genetically susceptible to peanut allergy and prone to severe reactions after OFC. More broadly, these findings demonstrate the untapped potential of the CC population in developing novel models for allergy research.

Keywords: Ara h 2; Collaborative Cross; anaphylaxis; food allergy; mouse model; peanut allergy.

Figure 1.

Collaborative Cross screening approach. Schematic shows 4 representative strains of the 16 strains screened. Six female mice between the ages of 4 and 6 weeks from each strain were mixed so that each cage contained 3–5 mice from different strains. Mice were then transferred from the UNC Systems Genetics Core to the UNC Food Allergy Initiative where researchers were blinded to the identification of each strain. Mice were sensitized intragastrically with peanut extract and Cholera toxin for 4 weeks before undergoing either an OFC (n=3/strain) or IP challenge (n=3/strain) with peanut extract.

Figure 2.

Anaphylaxis in peanut-sensitized Collaborative Cross strains following peanut challenge. Oral and IP challenges revealed Collaborative Cross strains that are non-reactors (A, D), mild reactors (B, E) and severe reactors (C, F) as measured by decreased body temperature. Challenges were repeated with an IP reactor control (G), a non-reactor control (H), and oral reactors (I, J).

Figure 3.

Immune response of CC027/GeniUnc to peanut extract relative to that of C3H/HeJ, C57BL/6J, and BALB/cJ mice. CC027/GeniUnc is represented as CC027 in figures. Body temperatures following oral challenge with peanut extract (n≥12/strain; representative of 3 independent experiments) (A). Serum levels of immunoglobulins following 4 weeks of sensitization (B-H). Mann-Whitney U Test *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001. Statistical significance represents comparisons of C3H/HeJ, C57BL/6J, and BALB/cJ relative to CC027/GeniUnc (A).

Figure 4.

Cellular responses in CC027/GeniUnc, C3H/HeJ, and C57BL/6J. Splenic cytokines 96 hours following peanut-stimulation (n=10/strain) (A-H), mRNA expression (I-J), and CD4+CD25+FoxP3+ regulatory T cells (K) 1 week following oral challenge. Mann-Whitney U Test *p<0.05, **p<0.01, ***p<0.001 (A-H; K); unpaired t-test *p<0.05, **p<0.01 (I-J).

Figure 5.

Enumeration of effector cells in CC027/GeniUnc, C3H/HeJ, and C57BL/6J. Percent IgE+CD49b+ basophils (A) and basophil inhibitory receptor, CD200R1 in whole blood (B). Jejunal tryptase+ mast cells quantified 1–3 weeks following challenge (C) and representative staining images shown with arrows indicating tryptase+ cells (D). Mann-Whitney U Test *p<0.05, **p<0.01, ***p<0.001.

Figure 6.

Post-OFC serum levels of mast cell degranulation marker and the major peanut allergen Ara h 2. Serum levels of MMCP-1 (A) and Ara h 2 (C) 60 min after oral challenge; Mann-Whitney U Test *p<0.05, **p<0.01, ****p<0.0001. Correlations between MMCP-1 (B) or Ara h 2 (D) and maximum body temperature decrease following oral challenge in CC027/GeniUnc.

Figure 7:

Anaphylaxis in peanut-sensitized mice following oral peanut challenge. Body temperatures after oral challenge with peanut extract in C3H/HeJ (A), BALB/cJ (B), and CC027/GeniUnc (C) mice sensitized with PBS, peanut extract once per week (PN 1X), peanut extract plus Cholera toxin once per week (PN+CT), or peanut extract three times per week (PN 3X). Peanut-specific IgE quantities in mice pre- and post-sensitization (D). Mann-Whitney U Test *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001.