Impact of peptidylarginine deiminase 4 (PAD4) deficiency in a fecal-induced peritonitis model of sepsis

Abstract

Background: Peptidylarginine deiminase 4 (PAD4) citrullinates histones, enabling the release of neutrophil extracellular traps. While neutrophil extracellular traps capture and kill pathogens, they also drive immunothrombosis, potentially worsening sepsis outcomes. However, it remains unclear whether PAD4 deficiency is beneficial or harmful in sepsis.

Objectives: To evaluate the impact of PAD4 deficiency in a fecal-induced peritonitis sepsis model, with and without antibiotic treatment, and incorporating fluid resuscitation in both sexes.

Methods: Wild-type and PAD4 knockout (PAD4^-/-) C57Bl/6 mice received intraperitoneal injections of fecal slurry (0.6 mg/g). Mice received buprenorphine every 8 hours and antibiotics/fluids every 12 hours. Survival studies were also conducted without antibiotics at a reduced fecal dose (0.4 mg/g). Mice were culled at 8 hours or 48 hours after infection. Organs, blood, and peritoneal cavity fluid were collected. Plasma levels of interleukin (IL)-6, IL-10, cell-free DNA, and thrombin-antithrombin were quantified, as well as bacterial loads in blood and peritoneal cavity fluid. Organ histology/immunohistochemistry was performed.

Results: Female PAD4^-/- mice had worsened survival compared with female wild-type mice. Male mice exhibited worse survival than females in both strains. Antibiotics eliminated survival differences between strains and sexes. Septic PAD4^-/- mice had reduced IL-10 in the early phase of sepsis, increased lung myeloperoxidase, and exacerbated lung injury compared with septic wild-type mice.

Conclusion: PAD4 deficiency in female mice worsened survival in the fecal-induced peritonitis sepsis model. In both strains, male mice exhibited worse survival compared with their female counterparts. PAD4 deficiency is associated with reduced IL-10, increased neutrophil infiltration, and exacerbated lung injury. Antibiotics eliminated survival differences between strains and sexes.

Keywords: PAD4; mice; neutrophil extracellular traps (NETs); sepsis.