Distinct hepatic myeloid and lymphoid cell repertoires are associated with susceptibility and resistance to Ascaris infection

Abstract

The soil-transmitted helminth Ascaris lumbricoides infects ~800 million people worldwide. Some people are heavily infected, harbouring many worms, whereas others are only lightly infected. The mechanisms behind this difference are unknown. We used a mouse model of hepatic resistance to Ascaris, with C57BL/6J mice as a model for heavy infection and CBA/Ca mice as a model for light infection. The mice were infected with the porcine ascarid, Ascaris suum or the human ascarid, A. lumbricoides and immune cells in their livers and spleens were enumerated using flow cytometry. Compared to uninfected C57BL/6J mice, uninfected CBA/Ca mice had higher splenic CD4+ and γδ T cell counts and lower hepatic eosinophil, Kupffer cell and B cell counts. Infection with A. suum led to expansions of eosinophils, Kupffer cells, monocytes and dendritic cells in the livers of both mouse strains and depletions of hepatic natural killer (NK) cells in CBA/Ca mice only. Infection with A. lumbricoides led to expansions of hepatic eosinophils, monocytes and dendritic cells and depletions of CD8+, αβ, NK and NK T cells in CBA/Ca mice, but not in C57BL/6J mice where only monocytes expanded. Thus, susceptibility and resistance to Ascaris infection are governed, in part, by the hepatic immune system.

Keywords: Ascaris lumbricoides; Ascaris suum; flow cytometry; liver; mouse model; resistance; susceptibility.

Fig. 1.

Lymphoid cell subtype numbers in the spleens of uninfected and Ascaris suum-infected C57BL/6J and CBA/Ca mice. (A) Gating strategy for the definition of lymphoid cell populations in spleens and livers. Following flow cytometric acquisition of MNCs, an electronic gate was placed on the lymphocytes based on forward and side scatter areas (FSC-A vs SSC-A) followed by gating of singlets (FSC-A vs FSC-H). Next, the live cells were gated upon in a dot plot of FSC-A vs dead cell stain (DCS). From these live cells, the activated cells were identified as CD69⁺ cells, T cells were identified as CD3⁺ cells and NKT cells were identified as CD3⁺ NK1.1⁺ cells. B cells were identified as CD19⁺ cells. αβ T cells (CD3⁺ and TCR γ/δ⁻) and γδ T cells (CD3⁺ and TCR γ/δ⁺) were identified after gating on CD3⁺ NK1.1⁻ cells. Finally, the αβ T cells were used to identify CD4⁺ and CD8⁺ T cells. (B) Scatter plots showing the lymphoid cell subtype numbers in the spleens of uninfected and A. suum-infected C57BL/6J and CBA/Ca mice. The numbers of cells per whole spleen for the different cell types for each sample are shown. The means are indicated with the red horizontal bars. **P < 0.01.

Fig. 2.

Lymphoid cell subtype numbers in the livers of uninfected and A. suum-infected C57BL/6J and CBA/Ca mice. The numbers of cells per mL of liver extract for the different cell types for each sample are shown. The means are indicated with the red horizontal bar. *P < 0.05; **P < 0.01.

Fig. 3.

Myeloid cell subtypes in the livers of uninfected and A. suum-infected C57BL/6J and CBA/Ca mice. (A) Gating strategy for the definition of myeloid cell populations in livers. For analysis of myeloid cells, debris was eliminated by gating based on FSC-A vs SSC-A followed by isolation of singlets (FSC-A vs FSC-H) and live cells (FSC-A vs DCS). Next, the monocytes were identified by plotting CD11b against F4/80. Eosinophils and KCs were identified by gating on the F4/80⁺ CD11b⁺ cells and plotting FSC-A against CD170. Myeloid and plasmacytoid DC were identified from gated F4/80⁻ CD11b⁻ cells and plotting CD317 against CD11c. Finally, basophils and mast cells were identified by gating on CD11c⁻ and CD137⁻ cells and plotting FSC-A against CD200R3. Gates for spleen and liver lymphoid and myeloid cells were manually adjusted for every sample. (B) Scatter plots showing numbers of myeloid cell subtypes in the livers of uninfected and A. suum-infected C57BL/6J and CBA/Ca mice. The number of cells per mL of liver extract for the different cell types for each sample is shown. The means are indicated with the red horizontal bar. **P < 0.01.

Fig. 4.

Lymphoid cell subtypes in the spleens of uninfected and Ascaris lumbricoides-infected C57BL/6J and CBA/Ca mice. The number of cells per whole spleen for the different cell types for each sample is shown. The means are indicated with the red horizontal bar. *P < 0.05; **P < 0.01.

Fig. 5.

Lymphoid cell subtype numbers in the livers of uninfected and A. lumbricoides-infected C57BL/6J and CBA/Ca mice. The number of cells per mL of liver extract for the different cell types for each sample is shown. The means are indicated with the red horizontal bar. *P < 0.05; **P < 0.01.

Fig. 6.

Myeloid cell subtype numbers in the livers of uninfected and A. lumbricoides-infected C57BL/6J and CBA/Ca mice. The number of cells per mL of liver extract for the different cell types for each sample is shown. The means are indicated with the red horizontal bar. *P < 0.05; **P < 0.01.