IL-15 constrains mast cell-dependent antibacterial defenses by suppressing chymase activities.

Sepsis remains a global clinical problem. By using the mouse cecal ligation and puncture model of sepsis, here we identify an important aspect of mast cell (MC)-dependent, innate immune defenses against Gram-negative bacteria by demonstrating that MC protease activity is regulated by interleukin-15 (IL-15). Mouse MCs express both constitutive and lipopolysaccharide-inducible IL-15 and store it intracellularly. Deletion of Il15 in mice markedly increases chymase activities, leading to greater MC bactericidal responses, increased processing and activation of neutrophil-recruiting chemokines, and significantly higher survival rates of mice after septic peritonitis. By showing that intracellular IL-15 acts as a specific negative transcriptional regulator of a mouse MC chymase (mast cell protease-2), we provide evidence that defined MC protease activity is transcriptionally regulated by an intracellularly retained cytokine. Our results identify an unexpected breach in MC-dependent innate immune defenses against sepsis and suggest that inhibiting intracellular IL-15 in MCs may improve survival from sepsis.

A monoclonal antibody and an enzyme immunoassay for human Ala-IL-8(77).

Interleukin-8 (IL-8) plays a central role in neutrophil chemotaxis and exerts a wide range of effects on various cells, ranging from tumor angiogenesis to impairment of neuronal signaling. Two main forms of IL-8 exist, one containing 77 amino acids (Ala-IL-8(77)) and a second containing 72 amino acids (Ser-IL-8(72)), which comprise more than 90% of IL-8 protein in cell cultures. IL-8(77) was reported to be produced predominantly by endothelial cells and is known as "endothelial" IL-8. IL-8(72) predominates in monocyte cultures and is known as "leukocyte" IL-8. While both forms have equal chemotactic activity in vivo, recent data suggest that their biological activities might be different. Here we describe the generation of a mouse monoclonal antibody (mAb) specific for IL-8(77) and the development of a corresponding immunoassay. Various immunization protocols were investigated. Immunization with conjugates of a peptide from the N-terminus of IL-8(77) (NTP(77)) resulted in the production of an IgG1 mAb (N11) that recognizes human IL-8(77) and neutralizes its chemotactic activity. A sensitive ELISA specific for IL-8(77) was developed using N11 for capture and a biotinylated mAb to IL-8(72) for detection. Using this immunoassay it was shown that the only form of IL-8 secreted in cell culture was IL-8(77) and that the IL-8(72) present was the result of proteolysis of IL-8(77). IL-8(77) was detected in plasma and cerebrospinal fluid (CSF) from patients with sepsis and meningitis.