Extracellular CIRP (eCIRP) and inflammation

Abstract

Cold-inducible RNA-binding protein (CIRP) was discovered 2 decades ago while studying the mechanism of cold stress adaptation in mammals. Since then, the role of intracellular CIRP (iCIRP) as a stress-response protein has been extensively studied. Recently, extracellular CIRP (eCIRP) was discovered to also have an important role, acting as a damage-associated molecular pattern, raising critical implications for the pathobiology of inflammatory diseases. During hemorrhagic shock and sepsis, inflammation triggers the translocation of CIRP from the nucleus to the cytosol and its release to the extracellular space. eCIRP then induces inflammatory responses in macrophages, neutrophils, lymphocytes, and dendritic cells. eCIRP also induces endoplasmic reticulum stress and pyroptosis in endothelial cells by activating the NF-κB and inflammasome pathways, and necroptosis in macrophages via mitochondrial DNA damage. eCIRP works through the TLR4-MD2 receptors. Studies with CIRP^-/- mice reveal protection against inflammation, implicating eCIRP to be a novel drug target. Anti-CIRP Ab or CIRP-derived small peptide may have effective therapeutic potentials in sepsis, acute lung injury, and organ ischemia/reperfusion injuries. The current review focuses on the pathobiology of eCIRP by emphasizing on signal transduction machineries, leading to discovering novel therapeutic interventions targeting eCIRP in various inflammatory diseases.

Keywords: ALI; CIRP; DAMP; eCIRP; hemorrhage; inflammation; ischemia/reperfusion; macrophage; neutrophils; sepsis.

Figure 1:. Release of eCIRP.

Sepsis and sterile injury promote global or localized infectious and hypoxic environment which activate TLR4 and HIF-1α. Bacterial LPS binds to TLR4 and recruits MyD88 to transduce downstream signal for activating NF-κB. HIF-1α and NF-κB serve as potent transcription factors to increase the expression of CIRP at mRNA levels. During inflammation or hypoxia cytoplasmic CIRP protein is enriched in the lysosomes. The lysosomes containing CIRP are then excreted outside the cells through exocytosis. eCIRP, extracellular cold-inducible RNA-binding protein; I/R, ischemia/reperfusion; TLR4, Toll-like receptor 4; HIF-1α, hypoxia inducible factor-1α; LPS, lipopolysaccharide; NF-κB, nuclear factor-κB; MyD88, myeloid differentiation factor 88.

Figure 2:. Proinflammatory effects of eCIRP.

eCIRP binds to its receptor TLR4/MD2 complex expressed in various cell types. In macrophages and lymphocytes, eCIRP activates NF-κB through TLR4/MD2 complex and increases the expression of proinflammatory cytokines and chemokines, and T cell activation markers such as CD69 and CD25. eCIRP skews proinflammatory Th1 type T cells by activating the master transcription factor T-bet. In neutrophils, through the classical TLR4/MD2 and NF-κB pathways eCIRP upregulates the expression of ICAM-1 on their surface. ICAM-1 transduces downstream signaling to activate PAD4-dependent NET formation. Increased NETosis by ICAM-1⁺ neutrophils in turn causes acute lung injury. In the DCs, a CIRP fusion protein prepared by combining amino acids of CD8 T cell’s TCR specific epitope from OVA and three amino-acid flanking residues linked to the N-terminus of murine CIRP named SIIN-CIRP induces DC maturation, cytokine production and migration in a TLR4-dependent manner. SIIN-CIRP protein also enhances antigen presentation. In the ECs, eCIRP increases the expression of ICAM-1 through TLR4/MD2 and NF-κB pathways. In addition, eCIRP induces NLRP3 inflammasome through TLR4/MD2 and NF-κB-dependent pathways. Increased activation of inflammasome leads to caspase-1 activation and subsequently IL-1β and IL-18 production and pyroptosis. eCIRP, extracellular cold-inducible RNA-binding protein; TLR4, Toll-like receptor 4; NF-κB, nuclear factor-κB; MyD88, myeloid differentiation factor 88; ICAM-1, intercellular cell adhesion molecule-1; DCs, dendritic cells; ECs, endothelial cells; SMC, smooth muscle cells; PAD4, peptidyl arginase deaminase 4; NLRP3, NACHT, LRR and PYD domains-containing protein 3.

Figure 3:. Neutrophil reverse transendothelial migration: role of eCIRP.

During inflammation neutrophils migrate from the blood to the tissues across the EC barrier through transendothelial migration. During inflammation eCIRP is increased in the blood and tissues. The TM neutrophils in the tissues are activated by eCIRP-TLR4 axis to release NE. The EC barrier is maintained by the interaction between JAM-C with its counterpart JAM-B. NE cleaves the JAM-C expressed on the surface of EC and enables TM neutrophils to pass through the disrupted EC barrier towards luminal direction. This process of neutrophil migration from the tissues to the blood during inflammation is called rTEM. The neutrophils that undergo rTEM is called RM neutrophils which further fuel inflammation. eCIRP, extracellular cold-inducible RNA-binding protein; TLR4, Toll-like receptor 4; EC, endothelial cells; TEM, transendothelial migration; TM, transmigrated; NE, neutrophil elastase; JAM-B/C, junctional adhesion molecule-B/C; rTEM, reverse transendothelial migration; RM, reverse migrated.