Fig. 2: Histones promote RBC and platelet aggregation, reduce RBC deformability, and induce platelet degranulation, processes that are inhibited by SPAs.

a Human RBCs were incubated for 1 h at 37 °C alone (control) or in the presence of histones (400 μg ml⁻¹) with or without mCBS (200 μg ml⁻¹). Percentage of RBC aggregation measured by flow cytometry based on forward (FSC) and side (SSC) scatter parameters and an appropriate gating strategy to discriminate aggregated from normal (non-aggregated) RBC. b Scanning electron micrographs depicting the level of RBC aggregation at low and high magnification following the three treatments (n = 1) depicted in a and each representative of three fields per magnification. c Concentration-dependent inhibition of histone-mediated RBC aggregation by mCBS and MTS, in this case RBC aggregation being calculated by flow cytometry as fold change in RBC auto-fluorescence relative to RBC in the absence of histones. The fold change values also provide an estimate of the number of RBC in each aggregate. Data were analyzed by two-way ANOVA with Sidak’s correction for multiple comparisons (n = 3). d Retention of RBC in an artificial spleen that measures RBC deformability. RBCs were incubated with increasing concentrations of histones for 1 h and, at the highest concentration used (400 μg ml⁻¹), also incubated with either mCBS or MTS (100 and 200 μg ml⁻¹), prior to passage through the artificial spleen. Data are presented as mean ± s.e.m. (n = 2–3) and analyzed by one-way ANOVA with Dunnett’s multiple comparisons tests. e Histone-induced aggregation of isolated platelets. f SPA inhibition of histone (HIS) (150 μg ml⁻¹) induced platelet aggregation. Data were analyzed by two-way ANOVA with Dunnett’s correction for multiple comparisons (n = 6). g Histone-induced degranulation of platelets in whole blood, as measured by ATP release. Dotted line ATP release from thrombin-activated platelets (n = 1). h SPA inhibition of histone-induced platelet degranulation. Data in g, h are representative of one of two experiments. Source data are provided as a Source Data File.