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Cold-inducible RNA-binding protein (CIRP) triggers inflammatory responses in hemorrhagic shock and sepsis

Nature Medicine volume 19, pages 1489–1495 (2013)Cite this article

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Abstract

A systemic inflammatory response is observed in patients undergoing hemorrhagic shock and sepsis. Here we report increased levels of cold-inducible RNA-binding protein (CIRP) in the blood of individuals admitted to the surgical intensive care unit with hemorrhagic shock. In animal models of hemorrhage and sepsis, CIRP is upregulated in the heart and liver and released into the circulation. In macrophages under hypoxic stress, CIRP translocates from the nucleus to the cytosol and is released. Recombinant CIRP stimulates the release of tumor necrosis factor-α (TNF-α) and HMGB1 from macrophages and induces inflammatory responses and causes tissue injury when injected in vivo. Hemorrhage-induced TNF-α and HMGB1 release and lethality were reduced in CIRP-deficient mice. Blockade of CIRP using antisera to CIRP attenuated inflammatory cytokine release and mortality after hemorrhage and sepsis. The activity of extracellular CIRP is mediated through the Toll-like receptor 4 (TLR4)–myeloid differentiation factor 2 (MD2) complex. Surface plasmon resonance analysis indicated that CIRP binds to the TLR4-MD2 complex, as well as to TLR4 and MD2 individually. In particular, human CIRP amino acid residues 106–125 bind to MD2 with high affinity. Thus, CIRP is a damage-associated molecular pattern molecule that promotes inflammatory responses in shock and sepsis.

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Figure 1: Increased expression and release of CIRP after hemorrhage.
Figure 2: Recombinant CIRP induces cytokine release in macrophages.
Figure 3: Blockade of CIRP reduces TNF-α and IL-6 production, hepatic injury and mortality after hemorrhage.
Figure 4: Role of CIRP in CLP sepsis.
Figure 5: The TLR4-MD2 complex mediates extracellular CIRP activity.

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References

  1. Holcomb, J.B. et al. Challenges to effective research in acute trauma resuscitation: consent and endpoints. Shock 35, 107–113 (2011).

    PubMed  Google Scholar 

  2. Bulger, E.M. et al. Hypertonic resuscitation of hypovolemic shock after blunt trauma: a randomized controlled trial. Arch. Surg. 143, 139–148 (2008).

    PubMed  Google Scholar 

  3. Kaczorowski, D.J., Mollen, K.P., Edmonds, R. & Billiar, T.R. Early events in the recognition of danger signals after tissue injury. J. Leukoc. Biol. 83, 546–552 (2008).

    CAS  PubMed  Google Scholar 

  4. Dombrovskiy, V.Y., Martin, A.A., Sunderram, J. & Paz, H.L. Rapid increase in hospitalization and mortality rates for severe sepsis in the United States: a trend analysis from 1993 to 2003. Crit. Care Med. 35, 1244–1250 (2007).

    PubMed  Google Scholar 

  5. Ward, P.A. New approaches to the study of sepsis. EMBO Mol. Med. 4, 1234–1243 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Medzhitov, R. Origin and physiological roles of inflammation. Nature 454, 428–435 (2008).

    CAS  PubMed  Google Scholar 

  7. Oppenheim, J.J. & Yang, D. Alarmins: chemotactic activators of immune responses. Curr. Opin. Immunol. 17, 359–365 (2005).

    CAS  PubMed  Google Scholar 

  8. Zhang, X. & Mosser, D.M. Macrophage activation by endogenous danger signals. J. Pathol. 214, 161–178 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Beutler, B.A. TLRs and innate immunity. Blood 113, 1399–1407 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  10. Ward, P.A. The sepsis seesaw: seeking a heart salve. Nat. Med. 15, 497–498 (2009).

    CAS  PubMed  Google Scholar 

  11. Kawai, T. & Akira, S. The role of pattern-recognition receptors in innate immunity: update on Toll-like receptors. Nat. Immunol. 11, 373–384 (2010).

    CAS  PubMed  Google Scholar 

  12. Chen, G.Y., Tang, J., Zheng, P. & Liu, Y. CD24 and Siglec-10 selectively repress tissue damage–induced immune responses. Science 323, 1722–1725 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  13. Wang, H. et al. HMG-1 as a late mediator of endotoxin lethality in mice. Science 285, 248–251 (1999).

    CAS  PubMed  Google Scholar 

  14. Tsung, A. et al. The nuclear factor HMGB1 mediates hepatic injury after murine liver ischemia-reperfusion. J. Exp. Med. 201, 1135–1143 (2005).

    CAS  PubMed  PubMed Central  Google Scholar 

  15. Quintana, F.J. & Cohen, I.R. Heat shock proteins as endogenous adjuvants in sterile and septic inflammation. J. Immunol. 175, 2777–2782 (2005).

    CAS  PubMed  Google Scholar 

  16. Martinon, F., Pétrilli, V., Mayor, A., Tardivel, A. & Tschopp, J. Gout-associated uric acid crystals activate the NALP3 inflammasome. Nature 440, 237–241 (2006).

    CAS  PubMed  Google Scholar 

  17. Vogl, T. et al. Mrp8 and Mrp14 are endogenous activators of Toll-like receptor 4, promoting lethal, endotoxin-induced shock. Nat. Med. 13, 1042–1049 (2007).

    CAS  PubMed  Google Scholar 

  18. Xu, J. et al. Extracellular histones are major mediators of death in sepsis. Nat. Med. 15, 1318–1321 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  19. Zhang, Q. et al. Circulating mitochondrial DAMPs cause inflammatory responses to injury. Nature 464, 104–107 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Nishiyama, H. et al. Cloning and characterization of human CIRP (cold-inducible RNA-binding protein) cDNA and chromosomal assignment of the gene. Gene 204, 115–120 (1997).

    CAS  PubMed  Google Scholar 

  21. Nishiyama, H. et al. A glycine-rich RNA-binding protein mediating cold-inducible suppression of mammalian cell growth. J. Cell Biol. 137, 899–908 (1997).

    CAS  PubMed  PubMed Central  Google Scholar 

  22. Xue, J.H. et al. Effects of ischemia and H2O2 on the cold stress protein CIRP expression in rat neuronal cells. Free Radic. Biol. Med. 27, 1238–1244 (1999).

    CAS  PubMed  Google Scholar 

  23. Nishiyama, H. et al. Decreased expression of cold-inducible RNA-binding protein (CIRP) in male germ cells at elevated temperature. Am. J. Pathol. 152, 289–296 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  24. Nishiyama, H. et al. Diurnal change of the cold-inducible RNA-binding protein (Cirp) expression in mouse brain. Biochem. Biophys. Res. Commun. 245, 534–538 (1998).

    CAS  PubMed  Google Scholar 

  25. Sheikh, M.S. et al. Identification of several human homologs of hamster DNA damage–inducible transcripts. Cloning and characterization of a novel UV-inducible cDNA that codes for a putative RNA-binding protein. J. Biol. Chem. 272, 26720–26726 (1997).

    CAS  PubMed  Google Scholar 

  26. Wellmann, S. et al. Oxygen-regulated expression of the RNA-binding proteins RBM3 and CIRP by a HIF-1–independent mechanism. J. Cell Sci. 117, 1785–1794 (2004).

    CAS  PubMed  Google Scholar 

  27. Qu, Y. & Dubyak, G.R. P2X7 receptors regulate multiple types of membrane trafficking responses and non-classical secretion pathways. Purinergic Signal. 5, 163–173 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  28. Aida, Y. & Pabst, M.J. Removal of endotoxin from protein solutions by phase separation using Triton X-114. J. Immunol. Methods 132, 191–195 (1990).

    CAS  PubMed  Google Scholar 

  29. Wang, Y. et al. Identification of stimulating and inhibitory epitopes within the heat shock protein 70 molecule that modulate cytokine production and maturation of dendritic cells. J. Immunol. 174, 3306–3316 (2005).

    CAS  PubMed  Google Scholar 

  30. Henderson, B. et al. Caught with their PAMPs down? The extracellular signalling actions of molecular chaperones are not due to microbial contaminants. Cell Stress Chaperones 15, 123–141 (2010).

    CAS  PubMed  Google Scholar 

  31. Andersson, U. et al. High mobility group 1 protein (HMG-1) stimulates proinflammatory cytokine synthesis in human monocytes. J. Exp. Med. 192, 565–570 (2000).

    CAS  PubMed  PubMed Central  Google Scholar 

  32. Yang, H. et al. Reversing established sepsis with antagonists of endogenous high-mobility group box 1. Proc. Natl. Acad. Sci. USA 101, 296–301 (2004).

    CAS  PubMed  Google Scholar 

  33. Yang, S., Zhou, M., Chaudry, I.H. & Wang, P. Novel approach to prevent the transition from the hyperdynamic phase to the hypodynamic phase of sepsis: role of adrenomedullin and adrenomedullin binding protein-1. Ann. Surg. 236, 625–633 (2002).

    PubMed  PubMed Central  Google Scholar 

  34. Nagai, Y. et al. Essential role of MD-2 in LPS responsiveness and TLR4 distribution. Nat. Immunol. 3, 667–672 (2002).

    CAS  PubMed  Google Scholar 

  35. Hyakushima, N. et al. Interaction of soluble form of recombinant extracellular TLR4 domain with MD-2 enables lipopolysaccharide binding and attenuates TLR4-mediated signaling. J. Immunol. 173, 6949–6954 (2004).

    CAS  PubMed  Google Scholar 

  36. Yang, C. & Carrier, F. The UV-inducible RNA-binding protein A18 (A18 hnRNP) plays a protective role in the genotoxic stress response. J. Biol. Chem. 276, 47277–47284 (2001).

    CAS  PubMed  Google Scholar 

  37. Cammas, A., Lewis, S.M., Vagner, S. & Holcik, M. Post-transcriptional control of gene expression through subcellular relocalization of mRNA binding proteins. Biochem. Pharmacol. 76, 1395–1403 (2008).

    CAS  PubMed  Google Scholar 

  38. De Leeuw, F. et al. The cold-inducible RNA-binding protein migrates from the nucleus to cytoplasmic stress granules by a methylation-dependent mechanism and acts as a translational repressor. Exp. Cell Res. 313, 4130–4144 (2007).

    CAS  PubMed  Google Scholar 

  39. Yang, R. et al. Functional significance for a heterogenous ribonucleoprotein A18 signature RNA motif in the 3′-untranslated region of ataxia telangiectasia mutated and Rad3-related (ATR) transcript. J. Biol. Chem. 285, 8887–8893 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  40. Qu, Y., Franchi, L., Nunez, G. & Dubyak, G.R. Nonclassical IL-1β secretion stimulated by P2X7 receptors is dependent on inflammasome activation and correlated with exosome release in murine macrophages. J. Immunol. 179, 1913–1925 (2007).

    CAS  PubMed  Google Scholar 

  41. Benhamou, Y. et al. Toll-like receptors 4 contribute to endothelial injury and inflammation in hemorrhagic shock in mice. Crit. Care Med. 37, 1724–1728 (2009).

    CAS  PubMed  Google Scholar 

  42. Wittebole, X., Castanares-Zapatero, D. & Laterre, P.F. Toll-like receptor 4 modulation as a strategy to treat sepsis. Mediators Inflamm. 2010, 568396 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  43. Park, J.S. et al. Involvement of toll-like receptors 2 and 4 in cellular activation by high mobility group box 1 protein. J. Biol. Chem. 279, 7370–7377 (2004).

    CAS  PubMed  Google Scholar 

  44. Ohashi, K., Burkart, V., Flohé, S. & Kolb, H. Cutting edge: heat shock protein 60 is a putative endogenous ligand of the toll-like receptor-4 complex. J. Immunol. 164, 558–561 (2000).

    CAS  PubMed  Google Scholar 

  45. Termeer, C. et al. Oligosaccharides of Hyaluronan activate dendritic cells via toll-like receptor 4. J. Exp. Med. 195, 99–111 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  46. Okamura, Y. et al. The extra domain A of fibronectin activates Toll-like receptor 4. J. Biol. Chem. 276, 10229–10233 (2001).

    CAS  PubMed  Google Scholar 

  47. Yang, H. et al. A critical cysteine is required for HMGB1 binding to Toll-like receptor 4 and activation of macrophage cytokine release. Proc. Natl. Acad. Sci. USA 107, 11942–11947 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  48. Yang, H., Antoine, D.J., Andersson, U. & Tracey, K.J. The many faces of HMGB1: molecular structure-functional activity in inflammation, apoptosis, and chemotaxis. J. Leukoc. Biol. 93, 865–873 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  49. Shin, H.J. et al. Kinetics of binding of LPS to recombinant CD14, TLR4, and MD-2 proteins. Mol. Cells 24, 119–124 (2007).

    CAS  PubMed  Google Scholar 

  50. Scaffidi, P., Misteli, T. & Bianchi, M.E. Release of chromatin protein HMGB1 by necrotic cells triggers inflammation. Nature 418, 191–195 (2002).

    CAS  PubMed  Google Scholar 

  51. Brochu, C. et al. NF-κB–dependent role for cold-inducible RNA binding protein in regulating interleukin 1β. PLoS ONE 8, e57426 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  52. Sakurai, T. et al. Cirp protects against tumor necrosis factor-α–induced apoptosis via activation of extracellular signal-regulated kinase. Biochim. Biophys. Acta 1763, 290–295 (2006).

    CAS  PubMed  Google Scholar 

  53. Wu, R. et al. Mechanisms responsible for vascular hyporesponsiveness to adrenomedullin after hemorrhage: the central role of adrenomedullin binding protein-1. Ann. Surg. 242, 115–123 (2005).

    PubMed  PubMed Central  Google Scholar 

  54. Miksa, M. et al. Immature dendritic cell–derived exosomes rescue septic animals via milk fat globule epidermal growth factor-factor VIII. J. Immunol. 183, 5983–5990 (2009).

    CAS  PubMed  Google Scholar 

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Acknowledgements

This work was supported by US National Institutes of Health (NIH) grants HL076179 and GM053008 (P.W.). We thank H. Erlandsson-Harris (Karolinska Institute) for providing Ager−/−, Tlr2−/− and Tlr4−/− mice, M. Hu, J.H. Li and L.M. Corbo for technical assistance, L. Caracappa for editorial assistance and Y. Al-Abed and A. Ragab for assistance with SPR analysis. The BIAcore instrument was supported by NIH grant S10OD012042. J.F. was partly supported by the Smoking Research Foundation of Japan.

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  1. Xiaoling Qiang and Weng-Lang Yang: These authors contributed equally to this work.

Authors and Affiliations

  1. Center for Translational Research, The Feinstein Institute for Medical Research, Manhasset, New York, USA

    Xiaoling Qiang, Weng-Lang Yang, Rongqian Wu, Mian Zhou, Asha Jacob, Weifeng Dong, Michael Kuncewitch, Youxin Ji, Jeffrey Nicastro, Gene F Coppa & Ping Wang

  2. Center for Biomedical Science, The Feinstein Institute for Medical Research, Manhasset, New York, USA

    Huan Yang, Haichao Wang & Kevin J Tracey

  3. Department of Clinical Molecular Biology, Kyoto University, Kyoto, Japan

    Jun Fujita

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Contributions

X.Q. and M.Z. performed the experiments and analyzed data. W.-L.Y. conducted the translocation study, designed and coordinated SPR analysis and wrote the manuscript. R.W. designed the experiments. A.J. assisted with the design of experiments and participated in manuscript editing. W.D. and Y.J. performed animal studies. M.K. collected the serum from patients admitted to the surgical ICU and analyzed human data. J.N. and G.F.C. analyzed animal studies. H.Y. and K.J.T. assisted in the knockout mice study and SPR analysis. J.F. assisted in the CIRP knockout mice and GFP-CIRP study and revised the manuscript. H.W. assisted with the design of the study and analyzed data. P.W. designed and supervised the study and revised the manuscript.

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Correspondence to Ping Wang.

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Competing interests

P.W. is an inventor of the pending PCT application WO 2010/120726 A1 entitled “Treatment of inflammatory diseases by inhibiting cold-inducible RNA-binding protein (CIRP).” This patent application covers the fundamental concept of using CIRP inhibitors for the treatment of inflammatory diseases.

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Qiang, X., Yang, WL., Wu, R. et al. Cold-inducible RNA-binding protein (CIRP) triggers inflammatory responses in hemorrhagic shock and sepsis. Nat Med 19, 1489–1495 (2013). https://doi.org/10.1038/nm.3368

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