PAD4-dependent NETs generation are indispensable for intestinal clearance of Citrobacter rodentium

Peptidyl arginine deiminase-4 (PAD4) is indispensable for generation of neutrophil extracellular traps (NETs), which can provide antimicrobial effects during host innate immune response; however, the role of PAD4 against gastrointestinal infection is largely unknown. Herein, we challenged PAD4-deficient (Pad4(-/-)) mice and wild-type (WT) littermates with Citrobacter rodentium (CR), and investigated bacteria clearance and gut pathology. Luminal colonization of CR in Pad4(-/-) mice peaked between 11-14 days post-infection, whereas WT mice suppressed the infection by 14 days. We demonstrated that Pad4(-/-) mice were unable to form NETs, whereas WT mice showed increased NETs formation in the colon during infection. Pad4(-/-) mice showed aggravated CR-associated inflammation as indicated by elevated systemic and colonic pro-inflammatory markers. Histological analysis revealed that transmissible colonic hyperplasia, goblet cell depletion, and apoptotic cell death were more pronounced in the colon of CR-infected Pad4(-/-) mice. Treating WT mice with deoxyribonuclease I, which can disrupt NETs generation, recapitulated the exacerbated CR infection and gut pathology associated with the loss of PAD4. Administration of the PAD4 inhibitor, Cl-amidine also aggravated CR infection, but to a lesser extent. Taken together, our findings highlight the importance of PAD4 in the mucosal clearance of CR and in resolving gut-associated inflammation.