West Nile virus triggers intestinal dysmotility via T cell–mediated enteric nervous system injury
Hana Janova, Fang R. Zhao, Pritesh Desai, Matthias Mack, Larissa B. Thackray, Thaddeus S. Stappenbeck, Michael S. Diamond
Hana Janova, Fang R. Zhao, Pritesh Desai, Matthias Mack, Larissa B. Thackray, Thaddeus S. Stappenbeck, Michael S. Diamond
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Research Article Gastroenterology Infectious disease

West Nile virus triggers intestinal dysmotility via T cell–mediated enteric nervous system injury

Abstract

Intestinal dysmotility syndromes have been epidemiologically associated with several antecedent bacterial and viral infections. To model this phenotype, we previously infected mice with the neurotropic flavivirus West Nile virus (WNV) and demonstrated intestinal transit defects. Here, we found that within 1 week of WNV infection, enteric neurons and glia became damaged, resulting in sustained reductions of neuronal cells and their networks of connecting fibers. Using cell-depleting antibodies, adoptive transfer experiments, and mice lacking specific immune cells or immune functions, we show that infiltrating WNV-specific CD4+ and CD8+ T cells damaged the enteric nervous system (ENS) and glia, which led to intestinal dysmotility; these T cells used multiple and redundant effector molecules including perforin and Fas ligand. In comparison, WNV-triggered ENS injury and intestinal dysmotility appeared to not require infiltrating monocytes, and damage may have been limited by resident muscularis macrophages. Overall, our experiments support a model in which antigen-specific T cell subsets and their effector molecules responding to WNV infection direct immune pathology against enteric neurons and supporting glia that results in intestinal dysmotility.

Authors

Hana Janova, Fang R. Zhao, Pritesh Desai, Matthias Mack, Larissa B. Thackray, Thaddeus S. Stappenbeck, Michael S. Diamond

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Figure 4

Damage to the neuronal and glial networks is caused by CD4+ and CD8+ T cells.

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Damage to the neuronal and glial networks is caused by CD4+ and CD8+ T c...
(A and F) GI transit was measured after oral gavage of carmine red dye. (A) Transit time for sham- or WNV-infected WT or TCRbd–/– mice at 7 dpi. (B–D, G, and H) The muscularis externa was isolated from (B–D) middle and distal regions of the small intestine of sham- or WNV-infected WT or TCRbd–/– mice at 7 dpi, (G and H) middle regions of the small intestine of sham- or WNV-infected WT mice at 7 dpi that were treated with anti-CD4 and/or anti-CD8β or isotype control mAbs and stained for (B and G) calretinin+ and nNOS+ neurons, (C) WNV antigen, (D) S100β+ glia, or (H) and S100β+ glia and WNV antigen. The fraction of the area that stained positive for calretinin, nNOS, or S100β was determined, and the values were normalized to values for (B and D) WT sham-infected mice or (G and H) animals treated with an isotype control mAb. Representative images from the myenteric plexus of the middle region of the small intestine. Scale bars: 100 μm. Original magnification, ×2.5 (enlarged insets). (C) Data are presented as the percentage of WNV antigen+ area in the field of view. (E) Counts of live CD45+TCRβ+CD4+ or CD8+ T cells in the muscularis of sham- or WNV-infected C57BL6/J mice at 7 dpi. (F) Transit time for sham- or WNV-infected mice at 7 dpi; mice were treated with anti-CD4 (α-CD4) and/or anti-CD8β or isotype control mAbs. Data were pooled from (A) 3; (C–E, and G) 2; and (F) 4 experiments. The numbers of mice per group were as follows: (A) n = 7–13; (C and D) n = 5–8; (E) n = 6–7; (F) n = 8–18; (G) n = 7–8; (H) n = 6–8. Lines in A, E, and F and column heights in B–D, G, and H indicate mean values. *P < 0.05, **P < 0.01, and ***P < 0.001, by (A, B, D, G, and H) Kruskal-Wallis ANOVA with Dunn’s post test (all groups compared with each other); (F) Kruskal-Wallis ANOVA with Dunn’s post test (compared with the isotype control group); and (C) 2-tailed Mann-Whitney U test.