Keratinocytes escape cell damage caused by neutrophil extracellular traps

Summary: Abstract Body: Neutrophil extracellular traps (NETs) are released by activated neutrophils and are defined as cytotoxic protein-modified DNA-based extracellular reticular structures. The major component of the extracellular reticular structure is decondensed DNA bound to a variety of proteins, including citrullinated histone H3 (cit-H3), neutrophil elastase, cathepsin G, and myeloperoxidase, which are stored in azurophilic granules under normal conditions. Therefore, when examining NETs, it is common to look for reticular structures and the presence of cit-H3. Indeed, reticular structures and cit-H3 were clearly observed when NETs were induced in vitro by exposure of circulating neutrophils from healthy volunteers to phorbol 12-myristate 13-acetate (PMA). In contrast, we found that cit-H3 was identified in neutrophils of pustules of neutrophilic pustular dermatoses such as generalized pustular psoriasis, acute generalized exanthematous pustulosis, and palmoplantar pustulosis, but the extracellular reticular structure was absent. Thus, it has been hypothesized that epidermal keratinocytes release molecules that degrade reticular structures or inhibit the formation of reticular structures. To test this hypothesis, neutrophil culture medium was replaced with HaCaT cell culture supernatant, and PMA was added for 5 hours, resulting in the identification of cit-H3 without extracellular reticular structure. Next, culture supernatants were recovered from sub confluent and full confluent normal human epidermal keratinocytes. As a result, disappearance of the extracellular reticular structure was observed only in the latter, suggesting that full confluent or differentiated keratinocytes release molecules that degrade reticular structures or inhibit the formation of reticular structures. Cit-H3-positive neutrophils without reticular structures were also observed in the section of squamous cell carcinoma. Considering that reticular structures of NETs contain many cytotoxic molecules and cause severe cell damage, this process may be a way for keratinocytes to escape cell damage caused by NETs. Lisa Minai¹, Youichi Ogawa¹, Tatsuyoshi Kawamura¹, Shinji Shimada¹ 1. Dermatology, Shimkato, Chuo, Japan. Cell Communication Networks and Stromal Biology