Human CRP Defends against the Toxicity of Circulating Histones

Abstract

C-reactive protein (CRP) is an acute-phase protein that plays an important defensive role in innate immunity against bacterial infection, but it is also upregulated in many noninfectious diseases. The generic function of this highly conserved molecule in diseases that range from infection, inflammation, trauma, and malignancy is not well understood. In this article, we demonstrate that CRP defends the human body against the toxicity of histones released into the circulation after extensive cell death. In vitro, CRP significantly alleviates histone-induced endothelial cell damage, permeability increase, and platelet aggregation. In vivo, CRP rescues mice challenged with lethal doses of histones by inhibiting endothelial damage, vascular permeability, and coagulation activation, as reflected by significant reductions in lung edema, hemorrhage, and thrombosis. In patients, elevation of CRP significantly increases the capacity to neutralize extracellular histones in the circulation. We have also confirmed that CRP interacts with individual histones in vitro and forms CRP–histone complexes in serum from patients with both elevated CRP and histones. CRP is able to compete with phospholipid-containing liposomes for the binding to histones. This explains how CRP prevents histones from integrating into cell membranes, which would otherwise induce calcium influx as the major mechanism of cytotoxicity caused by extracellular histones. Because histone elevation occurs in the acute phase of numerous critical illnesses associated with extensive cell death, CRP detoxification of circulating histones would be a generic host defense mechanism in humans.

Histones as Mediators of Host Defense, Inflammation and Thrombosis

“…Histones bind to endothelial cells and cause cell permeabilization leading to calcium influx and cell death at concentrations in the range of 50 μg/ml [22,95,96]. Levels of free histones in this range or higher were found in serum of patients with severe trauma, pancreatitis or sepsis [95].…”

“…Extracellular histones have also been found to mediate liver injury [93,94]. Lung injury occurring during major trauma, after transfusions or after C5a generation, has also found to be mediated by histones [22,[95][96][97]. In the case of transfusion-related acute lung injury, platelet activation resulted in NET formation in blood and lungs and anti-histone antibody or DNase were protective [97].…”

Biology, role and therapeutic potential of circulating histones in acute inflammatory disorders

“…Indeed, the mechanism of cell necrosis has significant impact on the kinetics of nuclear DAMP release,11 and nuclear DAMPs acting as complexes have been reported to exert different activities compared to protein isolates 12. Furthermore, where purified histones injected into experimental animals are lethal within minutes,13 necrotic cell death releases nucleosomes (ie: histone‐DNA complexes) which overall appear to be less toxic 14. Indeed a study injecting similar doses of nucleosomes in mice makes no mention of toxicity,15 and others have demonstrated cofactors such as HMGB1 responsible for the immune‐stimulatory effects of nucleosomes 16.…”

“…Once integrated, histones induce permeabilization of membranes to cations, disruptions of cellular calcium signalling112 and cell death by necrosis. Negatively charged acute‐phase proteins (such as C‐reactive protein, CRP),14 DNA,128 innate polysaccharides (heparin)120 or synthetic macromolecules126 compete with membrane phospholipids and prevent histone integration and toxicity. Bactericidal properties of histone fragments are dependent on their ability to form amphipathic α‐helices—potentially membrane spanning domains—however no such structural analyses have been performed on mammalian cells to date 89…”

“…A number of innate and synthetic substances have demonstrated the ability to inhibit histone‐related toxicity based on surface charge alone, including plasma proteins (albumin,123 CRP14), polypeptides (polyglutamic acid126) and polysaccharides (heparin/heparanoids,111, 146 polysialic acid,147 bacterial O‐antigen148). Elevated histone‐degrading activated protein C (APC) levels are associated with better outcomes in sepsis115, 149 and trauma patients137; APC therapy is being evaluated for treatment of sepsis150 and pancreatitis 151.…”