Platelet-erythrocyte complex production is exacerbated by histones and protected by small polyanion histone neutralizing compounds

Abstract

Platelet-erythrocyte (PE) complexes, composed of single erythrocytes bound to platelets, are a physiological component of the bloodstream and contribute to erythrocyte turnover. Conditions in which PE complex production is increased or their clearance from the circulation is impeded are associated with hypercoagulable states and thrombocytopenia. Sepsis is a serious acute inflammatory condition that is often complicated by major hemostatic derangements, including thrombosis, anemia and thrombocytopenia, and an important driver of sepsis pathology are extracellular histones, which are released into the circulation from activated neutrophils. Histone-neutralizing small polyanions (SPAs) have been developed to block the cytotoxic effects of histones and are beneficial in treating sepsis. This study investigated whether histones may stimulate increased PE complex production as a potential cause of the hemostatic complications in disease, and whether SPAs can prevent complex generation. Treatment with histones in co-incubated human erythrocytes and platelets, or murine blood, increased PE complex production approximately 10-fold compared with control conditions. Mice administered histones by intravenous injection displayed reduced platelet counts but sustained complex levels, suggesting complex production was enhanced within the diminished platelet pool. The platelets in the histone-stimulated complexes were also more activated than cell-free platelets, according to increased surface expression of P-selectin and the fibrin-binding form of integrin αIIbβ3, determined using flow cytometry, and their altered shape and presence of filopodia observed by scanning electron microscopy. Inclusion of SPAs in the histone-treated cells and blood prevented histone-stimulated production of complexes, indicating the importance of the histone-mediated cell injury in their generation and identifying PE complexes as a treatment target. Why was the study done? Platelets can attach to red blood cells to form platelet–erythrocyte (PE) complexes, which normally help remove old or damaged red blood cells. However, excessive complex formation may contribute to blood clotting disorders and low platelet counts in diseases like sepsis. In these conditions, proteins called histones are released into the bloodstream and can damage cells. This study investigated whether histones, as a result of damaging erythrocytes, increase PE complex formation and whether drugs that neutralize histones can prevent this. What did the researchers do? Researchers examined human blood cells in the laboratory and mouse blood samples to measure PE complex formation after exposure to histones. They also assessed platelet activation and tested two histone-neutralizing compounds (small polyanions, SPAs) to see if they could block these effects. Additional experiments were conducted in mice given histones. What did the researchers find? Histones increased PE complex formation by about 10-fold and made platelets more activated in the blood cells and samples. In live mice, histone administration sustained complex formation despite the histones also reducing the number of available platelets. The SPA compounds prevented complex formation when added before or alongside histones but were ineffective after complexes had formed. What do the findings mean? Histones may drive harmful clotting and lead to low platelet counts by increasing PE complexes and platelet activation. Drugs that neutralize histones could help prevent these effects and represent a potential treatment approach.