Intraluminal neutrophils limit epithelium damage by reducing pathogen assault on intestinal epithelial cells during Salmonella gut infection

Abstract

Recruitment of neutrophils into the gut epithelium is a cardinal feature of intestinal inflammation in response to enteric infections. Previous work using the model pathogen Salmonella Typhimurium (S. Tm) established that invasion of intestinal epithelial cells by S.Tm leads to recruitment of neutrophils into the gut lumen, where they can reduce pathogen loads transiently. Notably, a fraction of the pathogen population can survive this defense, re-grow to high density, and continue triggering enteropathy. However, the functions of intraluminal neutrophils in the defense against enteric pathogens and their effects on preventing or aggravating epithelial damage are still not fully understood. Here, we address this question via neutrophil depletion in different mouse models of Salmonella colitis, which differ in their degree of enteropathy. In an antibiotic pre-treated mouse model, neutrophil depletion by an anti-Ly6G antibody exacerbated epithelial damage. This could be linked to compromised neutrophil-mediated elimination and reduced physical blocking of the gut-luminal S.Tm population such that the pathogen density remained high near the epithelial surface throughout the infection. The removal of luminal S. Tm by gentamicin, an antibiotic restricted to the gut lumen, reversed the effect of neutrophil depletion on epithelial cell loss. Strikingly, when using germ-free mice and an S. Tm ssaV mutant capable of epithelium invasion, but attenuated for survival and growth within host tissues, neutrophil depletion caused exacerbated immune activation of the gut mucosa and a complete destruction of the epithelial barrier. Together, our data indicate that intraluminal neutrophils are central for maintaining epithelial barrier integrity during acute Salmonella-induced gut inflammation, by limiting the sustained pathogen assault on the epithelium in a critical window of the infection. Highlights{circ} After the first wave of mucosal invasion (day 1 p.i.), S. Tm maintains the assault from the lumen, triggering the continued expulsion of epithelial cells in antibiotic pre-treated mice. {circ}Neutrophil recruitment into the gut lumen is essential to limit this continued Salmonella attack on the epithelium. {circ}In antibiotic pre-treated SPF mice, neutrophil depletion exacerbates S. Tm invasion, causing excessive epithelial cell loss, which compromises epithelial barrier integrity at later time points (day 2-3 p.i.). {circ}In germ-free mice, neutrophil depletion exacerbates epithelial responses and epithelial barrier destruction even more strongly than in streptomycin pre-treated SPF mice. {circ}Gentamicin treatment and ssaV mutant infections indicate that neutrophils prevent epithelial damage by eliminating and physically blocking gut-luminal pathogens.