Epithelial thrombin-PAR1 signaling and Nippostrongylus brasiliensis infection drive small intestinal villus vascularization

Introduction: Gut microbiota is known to affect the capillary network in small intestinal villus structures in a protease-activated receptor-1 (PAR1; F2r)-dependent fashion. The ectopic expression of thrombin (F2), a PAR1-activating protease, was recently reported in intestinal epithelial cells (IEC) affecting microbial spatial organization and intestinal injury. Bacterial and parasitic metabolites can be sensed through specialized small intestinal epithelial cells, called tuft cells, whose activation triggers type 2 immunity involved in the expulsion of nematodes like Nippostrongylus brasiliensis.

Our studies aim to shed light on the interaction between epithelial PAR1 signaling, tuft cells and vascularization in the small intestine.

Method: Mice deficient for intestinal epithelial thrombin(F2^ΔIEC) or PAR1 (F2r^ΔIEC) were generated using a Villin1-specific Cre-loxP system.

Wildtype C57BL/6J mice were injected subcutaneously with infective N. brasiliensis larvae.

Experimental groups were sex and age matched.

Paraffin sections of the mid small intestine were stained by immunofluorescence for the endothelial cell marker CD31 and the tuft cell marker DCLK1 to quantify the vascularized area and tuft cell counts in the intestinal villus structures.

RNA extraction was performed on isolated IECs and whole small intestinal tissue, and converted to cDNA by reverse transcription. Gene expression was analyzed by qPCR.

Results: Both, mice lacking epithelial thrombin synthesis (F2^ΔIEC) and mice lacking the thrombin receptor PAR1 (F2r^ΔIEC) showed a decrease in vascular density of small intestinal villus structures that was paralleled by a reduced expression of vascular markers. Additionally, these mice showed lower numbers of tuft cells in their small intestinal epithelial lining and reduced tuft cell counts positively correlated with reduced intestinal villus vascularization. In line, deletion of intestinal epithelial PAR1 changed the expression of epithelial differentiation markers. Interestingly, nematode infection expanded tuft cell counts in the small intestine and increased villus vascularization in wildtype mice.

Conclusion: Our results indicate that a gut epithelial thrombin-PAR1-signaling axis affects vascularization and tuft cell counts in small intestinal villi. Parasitic infection with the nematode Nippostrongylus brasiliensis expanded tuft cell counts as well as small intestinal villus vascularization. Taken together, our findings strengthen the hypothesis, that PAR1 signaling in intestinal epithelial cells is involved in vascular remodeling of villus capillaries, potentially via tuft cells.

 

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Conflict of Interest:

The authors claim no conflict of interest.

Publication History

Article published online:
13 February 2025