Basophil responses in susceptible AKR mice upon infection with the intestinal helminth parasite Trichuris muris

Abstract

Intestinal helminth infection promotes a Type 2 inflammatory response in resistant C57BL/6 mice that is essential for worm clearance. The study of inbred mouse strains has revealed factors that are critical for parasite resistance and delineated the role of Type 1 versus Type 2 immune responses in worm clearance. In C57BL/6 mice, basophils are key innate immune cells that promote Type 2 inflammation and are programmed via the Notch signalling pathway during infection with the helminth Trichuris muris. However, how the host genetic background influences basophil responses and basophil expression of Notch receptors remains unclear. Here we use genetically susceptible inbred AKR/J mice that have a Type 1-skewed immune response during T. muris infection to investigate basophil responses in a susceptible host. Basophil population expansion occurred in AKR/J mice even in the absence of fulminant Type 2 inflammation during T. muris infection. However, basophils in AKR/J mice did not robustly upregulate expression of the Notch2 receptor in response to infection as occurred in C57BL/6 mice. Blockade of the Type 1 cytokine interferon-γ in infected AKR/J mice was not sufficient to elicit infection-induced basophil expression of the Notch2 receptor. These data suggest that the host genetic background, outside of the Type 1 skew, is important in regulating basophil responses during T. muris infection in susceptible AKR/J mice.

Keywords: AKR-inbred strain; Trichuris muris; Type 2 inflammation; basophils; notch signalling.

Figure 1.. Susceptible AKR/J mice retain T. muris and have a Type 1 immune skew.

(A) Experimental schematic. (B) Worm burden in AKR/J mice. Real time PCR analysis of proximal colon homogenate from naïve and T. muris (Tm)-infected mice, quantifying (C) Ifng (D) Il4, (E) Il13, (F) Fcer1a, and (G) Mcpt8 (normalized to Actb). Data from all combined experiments are presented as the mean ± SEM. Statistical analysis was performed using a linear fixed-effect model with pairwise comparison; *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001. n=2–3 for each time point per experiment; 3 independent experiments.

Figure 2.. The basophil population expands upon T. muris infection in AKR/J mice.

Flow cytometric analysis for basophils in naïve and T. muris (Tm)-infected mice. (A) Cecum basophil gating (CD45⁺Lineage (Lin)⁻FcεRIα⁺IgE⁺CD49b⁺ckit^-). Basophil percentage of (B) Lin^- and (C) CD45⁺ cecum cells. (D) Spleen basophil gating (CD45⁺Lineage (Lin)⁻FcεRIα⁺IgE⁺CD49b⁺ckit^-). Basophil percentage of (E) Lin^- and (F) CD45⁺ spleen cells. Data from all combined experiments are presented as the mean ± SEM. Statistical analysis was performed using a linear fixed-effect model with pairwise comparison; *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001. n=2–3 for each time point per experiment; 3 independent experiments.

Figure 3.. Basophil Notch2 expression does not substantially increase following T. muris infection in AKR/J mice.

Flow cytometric analysis of basophils in the cecum in naïve and T. muris (Tm)-infected mice, displaying (A) representative plots for Notch2 staining, (B) percent Notch2⁺ basophils, and (C) Notch2 geometric mean fluorescence intensity (gMFI) on cecum basophils. Data from all combined experiments are presented as the mean ± SEM. n=2–3 for each time point per experiment; 3 independent experiments.

Figure 4.. Basophil Notch2 expression does not changes at day 14 post-T. muris infection upon IFN-γ neutralization in AKR/J mice.

(A) Experimental schematic. IFN-γ protein levels as measured by ELISA in (B) serum and (C) proximal colon tissue lysate, and (D) real time PCR analysis of proximal colon homogenate quantifying Ifng (normalized to Actb) from naïve and T. muris (Tm)-infected mice. Flow cytometric analysis of basophils in the cecum in naïve and Tm-infected mice, displaying (E) representative plots for Notch2 staining, (F) percent Notch2⁺ basophils, and (G) Notch2 geometric mean fluorescence intensity (gMFI) on basophils. Data from all combined experiments are presented as the mean ± SEM. Statistical analysis was performed using a linear fixed-effect model with pairwise comparison; *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001. n=2–3 for each time point per experiment; 3 independent experiments.