Nilsson, Per H.

Introduction Platelets contain a wide range of bioactive mediators, many of which accumulate during storage of platelet concentrates and have immunomodulatory properties. Our objective was to investigate how mediators derived from stored platelet concentrates modulate the acute inflammatory response using an ex vivo model. Methods Platelet-depleted whole blood from healthy donors (n=8) was supplemented with pooled platelet concentrate supernatants generated from six platelet concentrates sampled on storage days 1, 3, 5, and 7, and incubated with or without heat-inactivated Escherichia coli for four hours at 37 °C. Inflammatory responses were assessed by measuring complement activation, platelet-derived mediators, and cytokines. Results In the absence of E. coli , addition of platelet concentrate supernatants increased the levels of 24 of 32 measured inflammatory mediators. In contrast, under E. coli -stimulated conditions, the supernatants increased 10 markers and attenuated 14 markers, including the key pro-inflammatory cytokines IL-1β, IL-6, and TNF. The attenuation of these cytokines ranged from 25-43%, whereas several other mediators were unchanged or increased. The majority of biomarkers showed limited differences across the investigated storage time points. Conclusions Our findings demonstrate that soluble factors released from platelet concentrates can modulate inflammatory responses in platelet-depleted blood in an ex vivo setting, with context-dependent, bidirectional, and selective immunomodulatory effects.

Platelet activation and the complement system are mutually dependent. Here, we investigated the effects of storage time on complement activation and platelet function in routinely produced platelet concentrates. The platelet concentrates (n = 10) were stored at 22 °C for seven days and assessed daily for complement and platelet activation markers. Additionally, platelet function was analyzed in terms of their responsiveness to protease-activated receptor-1 (PAR-1) and thromboxane A2 receptor (TXA2R) activation and their capacity to adhere to collagen. Complement activation increased over the storage period for all analyzed markers, including the C1rs/C1-INH complex (fold change (FC) = 1.9; p < 0.001), MASP-1/C1-INH complex (FC = 2.0; p < 0.001), C4c (FC = 1.8, p < 0.001), C3bc (FC = 4.0; p < 0.01), and soluble C5b-9 (FC = 1.7, p < 0.001). Furthermore, the levels of soluble platelet activation markers increased in the concentrates over the seven-day period, including neutrophil-activating peptide-2 (FC = 2.5; p < 0.0001), transforming growth factor beta 1 (FC = 1.9; p < 0.001) and platelet factor 4 (FC = 2.1; p < 0.0001). The ability of platelets to respond to activation, as measured by surface expression of CD62P and CD63, decreased by 19% and 24% (p < 0.05) for PAR-1 and 69–72% (p < 0.05) for TXA2R activation, respectively, on Day 7 compared to Day 1. The extent of platelet binding to collagen was not significantly impaired during storage. In conclusion, we demonstrated that complement activation increased during the storage of platelets, and this correlated with increased platelet activation and a reduced ability of the platelets to respond to, primarily, TXA2R activation.