Vitamin C: a novel regulator of neutrophil extracellular trap formation.
BACKGROUND
Neutrophil extracellular trap (NET) formation was recently identified as a novel mechanism to kill pathogens. However, excessive NET formation in sepsis can injure host tissues. We have recently shown that parenteral vitamin C (VitC) is protective in sepsis. Whether VitC alters NETosis is unknown.


METHODS
We used Gulo-/- mice as they lack the ability to synthesize VitC. Sepsis was induced by intraperitoneal infusion of a fecal stem solution (abdominal peritonitis, FIP). Some VitC deficient Gulo-/- mice received an infusion of ascorbic acid (AscA, 200 mg/kg) 30 min after induction of FIP. NETosis was assessed histologically and by quantification for circulating free DNA (cf-DNA) in serum. Autophagy, histone citrullination, endoplasmic reticulum (ER) stress, NFκB activation and apoptosis were investigated in peritoneal PMNs.


RESULTS
Sepsis produced significant NETs in the lungs of VitC deficient Gulo-/- mice and increased circulating cf-DNA. This was attenuated in the VitC sufficient Gulo-/- mice and in VitC deficient Gulo-/- mice infused with AscA. Polymorphonuclear neutrophils (PMNs) from VitC deficient Gulo-/- mice demonstrated increased activation of ER stress, autophagy, histone citrullination, and NFκB activation, while apoptosis was inhibited. VitC also significantly attenuated PMA induced NETosis in PMNs from healthy human volunteers.


CONCLUSIONS
Our in vitro and in vivo findings identify VitC as a novel regulator of NET formation in sepsis. This study complements the notion that VitC is protective in sepsis settings.
Department of Pharmacotherapy and Outcomes Science, Virginia Commonwealth University, Richmond, VA 23298, USA.
Bassem M
BM
Mohammed
Bernard J
BJ
Fisher
Donatas
D
Kraskauskas
Daniela
D
Farkas
Donald F
DF
Brophy
Alpha A
AA
Fowler
Ramesh
R
Natarajan
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001205:Ascorbic Acid / Q000494:pharmacology*; D001343:Autophagy / Q000187:drug effects; D059865:Endoplasmic Reticulum Stress / Q000235:genetics; D006657:Histones / Q000378:metabolism; D006801:Humans; D006867:Hydrolases / Q000235:genetics / Q000378:metabolism; D008168:Lung / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D009504:Neutrophils / Q000166:cytology / Q000187:drug effects*; D010538:Peritonitis / Q000150:complications / Q000517:prevention & control; D000080002:Protein-Arginine Deiminase Type 4; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D018805:Sepsis / Q000209:etiology / Q000517:prevention & control; D015398:Signal Transduction; D015854:Up-Regulation
D006657:Histones; D016328:NF-kappa B; D012333:RNA, Messenger; D006867:Hydrolases; D000080002:Protein-Arginine Deiminase Type 4; C552151:peptidylarginine deiminase 4, mouse; D001205:Ascorbic Acid