Authors

E Tydén¹; M Sellin²; F Martin¹; C Fossum¹; S Hellman¹;  ¹ Swedish University of Agricultural Sciences, Sweden;  ² Uppsala University, Sweden

Discussion

Organoid cultures derived from stem cells have become increasingly popular as experimental models for studying infections caused by various gastrointestinal pathogens in different host species. However, the size of infectious nematode larvae and the closed structure of 3-dimensional organoids often pose challenges when studying the natural route of infection. In order to address this issue, the present study utilized enteroids, organoids derived from the equine small intestine, to establish monolayer cultures on the apical surface of the epithelium, allowing for easier administration of infectious agents. To evaluate the functionality of these monolayers, they were stimulated with IL-4 and IL-13, and/or exposed to infectious stage larvae of equine nematodes such as Parascaris univalens, cyathostominae, and Strongylus vulgaris. The effects of these stimuli were assessed through qPCR analysis, histochemistry, immunofluorescence, live-cell imaging, and scanning electron microscopy. These analyses revealed that the monolayers were heterogeneous, consisting of both immature and differentiated cells including tuft cells and mucus-producing goblet cells. Stimulation with IL-4/IL-13 led to an increase in the differentiation of tuft and goblet cells, as evidenced by the expression of DCLK1 and MUC2. Co-culture with P. univalens further enhanced the expression of MUC2 in these cytokine-primed monolayers. Additionally, live-cell imaging showed morphological changes in the epithelial cells upon exposure to larvae, even in the absence of cytokine stimulation. Overall, this study presents the design, characterization, and usability of an experimental model representing the equine nematode-infected small intestinal epithelium. The presence of tuft cells and goblet cells, whose mucus production is influenced by Th2 cytokines and/or the presence of larvae, provides an opportunity for mechanistic studies on the physical interactions between nematodes and the equine intestinal mucosa.