<i>Giardia duodenalis</i> Directly Depletes Mucins in Intestinal Goblet Cells

Giardia duodenalis (syn. G. lamblia , G. intestinalis ) is a small intestinal, diarrheagenic parasite that infects humans and animals across the world. While some mechanisms of acute giardiasis have been explained, the pathogenesis remains largely unclear. Giardiasis has also been shown to induce post‐infectious chronic illnesses, including post‐infectious irritable bowel syndrome (PI‐IBS) and extra‐intestinal complications. The pathology of these requires elucidation. While the mucus layers of the small and large intestines are protective against enteric infections, the effects that G. duodenalis has on host mucus remain unknown. Recent evidence indicates that G. duodenalis , despite its selective colonization of the small intestine, is able to affect the large intestine, where it causes hypersensitivity. We hypothesized that G. duodenalis may disrupt the protective mucus layers along the intestinal tract, facilitating the onset and propagation of disease. Objectives and Methods The objectives of the study were threefold: i) To determine the effects of G. duodenalis on human mucus‐producing goblet cells. This was observed by exposing human colonic biopsies or purified human mucus to live G. duodenalis trophozoites ex vivo . ii) To determine the effects of G. duodenalis on host intestinal cells, the mucus lining, and the commensal microbiota layers within a living system. This was explored by inoculating trophozoites into C57Bl/6 mice (WT) and Muc2 −/− (KO) mice. iii) To determine the direct effects of G. duodenalis on goblet cells and particularly, the main component of the mucus layer, mucin‐2 (MUC2). The use of an in vitro model, exposing G. duodenalis trophozoites to a human colonic goblet cell line, LS174T, allowed the study of the direct effects of G. duodenalis in the absence of confounding host microbial, immune, endocrine, or other local factors. Results Periodic acid‐Schiff/Alcian blue (PAS/AB) staining revealed that colonic biopsies exposed to G. duodenalis exhibited smaller mucin granules containing less mucin content. Western blotting showed that G. duodenalis degraded purified human mucins. PAS/AB staining confirmed depletion of goblet cell mucins in WT mice exposed to G. duodenalis , in the small, as well as the large, intestines. Fluorescence imaging using in situ hybridization (FISH) and wheat germ agglutinin (WGA) staining demonstrated that in Giardia ‐infected tissues of WT mice, the mucus layer was entirely lost and commensal bacteria translocated into the epithelial cell layer and lamina propria. Lastly, G. duodenalis reduced intracellular MUC2 in human colonic goblet cells LS174T. These effects were inhibited by a broad‐spectrum cysteine protease inhibitor. Conclusions While G. duodenalis colonizes the upper small intestine, it depletes mucus throughout the small and large intestines, in human biopsies and mice. Mucin depletion is also observed in monocultures of human intestinal goblet cells. These data suggest that G. duodenalis causes mucin depletion in the host intestine, through a combination of direct mucus breakdown and hypersecretion of goblet cells. The findings also indicate a potential role for parasitic cysteine proteases in these effects. Support or Funding Information Supported by HPI NSERC CREATE, NSERC Discovery and Queen Elizabeth II Master's Scholarship

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