Cold-inducible RNA-binding Protein Induces Neutrophil Extracellular Traps in the Lungs during Sepsis

Abstract

Extracellular cold-inducible RNA-binding protein (CIRP) exaggerates inflammation and tissue injury in sepsis. Neutrophil extracellular traps (NETs) are released by activated neutrophils during sepsis. NETs contribute to pathogen clearance, but excessive NET formation (NETosis) causes inflammation and tissue damage. Peptidylarginine deiminase 4 (PAD4) is associated with NETosis by increasing histone citrullination and chromatin decondensation. We hypothesized that CIRP induces NETosis in the lungs during sepsis via upregulating PAD4 expression. Sepsis was induced in C57BL/6 wild-type (WT) and CIRP −/− mice by cecal ligation and puncture (CLP). After 20 h of CLP induction, NETs in the lungs of WT and CIRP −/− mice were quantified by flow cytometry by staining the single cell suspensions with MPO and CitH3 Abs. PAD4 expression in the lungs of WT and CIRP −/− mice after sepsis was assessed by Western blotting. In vitro effects of recombinant mouse (rm) CIRP for NETosis and PAD4 expression in the bone marrow-derived neutrophils (BMDN) were assessed by flow cytometry and Western blotting, respectively. After 20 h of CLP, NETosis in the lungs was significantly decreased in CIRP −/− mice compared to WT mice, which also correlated with the decreased PAD4 expression. Intratracheal administration of rmCIRP into WT mice significantly increased NETosis and PAD4 expression in the lungs compared to vehicle-injected mice. In vitro culture of BMDN with rmCIRP significantly increased NETosis and PAD4 expression compared to PBS-treated control. Fluorescence microscopy revealed typical web-like structures consistent with NETs in rmCIRP-treated BMDN. Thus, CIRP serves as a novel inducer of NETosis via PAD4 during sepsis.

Extracellular CIRP as an endogenous TREM-1 ligand to fuel inflammation in sepsis

“…Similar to other DAMPs, eCIRP recognizes Toll-like receptor 4 (TLR4) (5,8) expressed in macrophages, lymphocytes, and neutrophils to increase cytokine production (5), T cell polarization (9), and neutrophil activation (10). Healthy mice injected with recombinant murine CIRP (rmCIRP) develop acute lung injury (ALI) via activation of macrophages, neutrophils, and endothelial cells and display increased vascular permeability in the lungs (11,12).…”

Cold-inducible RNA-binding protein might determine the severity and the presences of major/minor criteria for severe community-acquired pneumonia and best predicted mortality

“…Cold-inducible RNA-binding protein (CIRP) is a damage-associated molecular pattern molecule that plays a pivotal role in triggering inflammatory response, and antisera to CIRP attenuate shock-induced inflammation, tissue injury, and lethality [20]. CIRP induces excessive neutrophil extracellular traps, which cause inflammation and tissue damage, in the lungs during sepsis [21]. Intravenous injection of recombinant murine CIRP in C57BL/6 mice causes lung injury, evidenced by vascular leakage, edema, increased leukocyte infiltration and cytokine production in the lung tissue.…”

Exosomes in Sepsis

“…Although NETs may trap and kill bacteria, excessive NET formation in sepsis cause tissue damage ( 3 ). DAMPs like eCIRP directly induces NETosis in sepsis in lungs and blood ( 58 , 59 ). This implies that exosomes may induce NET formation as they contain DAMPs and other stimulants of NETs ( Table 1 ).…”