Extracellular Cold-Inducible RNA-Binding Protein and Hemorrhagic Shock: Mechanisms and Therapeutics

Abstract

Hemorrhagic shock is a type of hypovolemic shock and a significant cause of trauma-related death worldwide. The innate immune system has been implicated as a key mediator in developing severe complications after shock. Inflammation from the innate immune system begins at the time of initial insult; however, its activation is exaggerated, resulting in early and late-stage complications. Hypoxia and hypoperfusion lead to the release of molecules that act as danger signals known as damage-associated molecular patterns (DAMPs). DAMPs continue to circulate after shock, resulting in excess inflammation and tissue damage. We recently discovered that cold-inducible RNA-binding protein released into the extracellular space acts as a DAMP. During hemorrhagic shock, hypoperfusion leads to cell necrosis and the release of CIRP into circulation, triggering both systemic inflammation and local tissue damage. In this review, we discuss extracellular cold-inducible RNA-binding protein (eCIRP)'s role in sterile inflammation, as well as its various mechanisms of action. We also share our more newly developed anti-eCIRP agents with the eventual goal of producing drug therapies to mitigate organ damage, reduce mortality, and improve patient outcomes related to hemorrhagic shock. Finally, we suggest that future preclinical studies are required to develop the listed therapeutics for hemorrhagic shock and related conditions. In addition, we emphasize on the challenges to the translational phase and caution that the therapy should allow the immune system to continue to function well against secondary infections during hospitalization.

Keywords: extracellular CIRP; inflammation; ischemia and reperfusion; organ damage; trauma hemorrhage.

Figure 1

Mechanisms of action of eCIRP in hemorrhagic shock. Damaged vessels are unable to deliver blood to tissue due to hemorrhage. The loss of perfusion and resultant hypoxemia result in ischemic damage to cells. Due to hypoxemia, cellular metabolism shifts to anaerobic metabolism via glycolysis which leads to increasing lactate production. The accumulation of lactate creates an acidotic environment, causing further cell damage. The accumulation of these factors results in cell death by varying mechanisms including apoptosis, necrosis, and necroptosis. As a result of cell death, the cell membrane is disrupted, and intracellular CIRP is released into the extracellular space, becoming eCIRP, as shown in the top right. During the shock state, blood flow is shunted towards vital organs. eCIRP, now in systemic circulation, will also be shunted towards these organs, causing increased inflammation. eCIRP will bind to TLR4 and triggering receptor expressed on myeloid cells (TREM1) receptors on both tissue and circulating macrophages, demonstrated in the bottom left. As a result of this binding, the macrophage will release pro-inflammatory cytokines into circulation, worsening inflammation in the already ischemic tissues. A detailed image shown in the bottom right illustrates eCIRP acting on TLR4/MyD88 and inducing damage to the mitochondria, causing an increase in cytosolic DNA. This cytosolic DNA via the cGAS pathway activates stimulator of interferon genes (STING) on the endoplasmic reticulum membrane. This results in an increase in pIRF, a transcription factor responsible for increasing expression of type I IFNs. The activation of this pathway increases the release of type I IFNs into the circulation. These pathways collectively lead to inflammation and injury in hemorrhagic shock.

Figure 2

eCIRP as a therapeutic target in hemorrhagic shock. Both C23 and M3 are small peptides derived from the human sequence of CIRP. C23 has high affinity to the TLR4/MD2 complex, and M3 has specific binding for the TREM1 receptor. Thus, both peptides block the binding of eCIRP to their respective receptors. In contrast, A12 is a synthetic oligonucleotide consisting of a poly(A) tail mimic that binds to eCIRP, blocking its binding site to the TLR4/MD2 complex. PS-OME miR 130 is a microRNA with stabilizing adjustments, including 3 phosphorothioate (PS) bonds at the 5′ and 3′ ends and 2′Omethyl (2′Ome) bases. It has a strong binding affinity to eCIRP, thereby preventing eCIRP from signaling via the TLR4/MD2 signaling pathway. As a result, these small molecule peptides and oligonucleotides prevent the interaction between endogenous eCIRP and its target receptors on macrophages. As a result, the inflammatory cascade is inhibited, and the hyperinflammatory response at the initial insult from hemorrhagic shock is prevented.