Activated platelets release abundant and functional IL-27. (A) Platelets from apheresis platelet concentrates (n = 11) were stimulated with TRAP-SFFLRN peptide (50 μg/ml, 30 minutes; Sigma-Aldrich, Saint-Quentin Fallavier, France); negative controls were not stimulated. IL-27 concentration in platelet supernatants was determined using a commercial enzyme-linked immunosorbent assay kit (RnD Systems Europe, Lille, France). Thrombin receptor activating peptide (TRAP) stimulation significantly increased IL-27 release from 18.42 ± 2.28 ng/ml to 47.17 ± 2.99 ng/ml (***P < 0.0005, t-test). (B) Then, the influence of IL-27-rich platelet supernatants on the expression of the activation marker CD86 was assessed in vitro on five independent highly purified blood B lymphocyte sets by means of flow cytometry, in duplicate for each condition. As a control, each set of B cells was incubated in minimal medium for 48 h with recombinant IL-27 (10 ng/ml; RnD Systems). We found that, in contrast to supernatants of non-activated platelets, supernatants of activated platelets provoke a significant increase in CD86 expression on B cells from 21 to 47% (*P < 0.05, t-test). CD86 expression was restored to baseline when B cells were pre-incubated with an antibody blocking the gp130 subunit of the IL-27 receptor (0.5 μg/ml; clone 28126, RnD Systems; #
P < 0.05, t-test). Results are presented as mean values ± standard deviations.