Page 95 - Haematologica May 2020

Platelet vesicles and monocyte interaction
ity score [ISS] >8). Analysis by flow cytometry showed acquisition of CD41 by circulating leukocytes with prefer- ential binding to monocytes (Figure 8A-B and Online Supplementary Figure S11A-B). The CD41 measured on monocytes was likely derived from PEV, as the intensity of fluorescent staining at 4 hours post trauma (2,226±474) was substantially below that of a single platelet (13,702±964) (Online Supplementary Figure S11B). Both the number of CD41+ monocytes and the intensity of staining for CD41 on them (MFI), correlated significantly with the severity of trauma (Figure 8C-D). Lastly, there was a marked loss of CD41+ monocytes from the blood within 4 to 12 hours, which was sustained for up to 72 hours (Figure 8E) and a decrease in circulating platelet counts which reflects platelet activation and PEV generation (Online Supplementary Figure S11C).
Discussion
We have defined a new thrombo-inflammatory route of monocyte recruitment via an adhesion molecule trans- ferred from platelets. Recruitment is reliant upon platelet- derived GPIbα, which allows monocyte capture by VWF exposed on the vessel wall. Previous studies have indicat- ed that platelet-derived chemokines can then induce arrest and migration.45 Online Supplementary Figure S12 sum- marises the steps we propose in this thrombo-inflamma- tory cascade. Importantly, the cascade may diverge from the normal pathways of leukocyte trafficking in a manner that could contribute to disease, as plasma borne PEV preferentially deliver functional GPIbα to the monocyte surface. Transfer of GPIbα can support adhesion of mono- cytes in vitro and in vivo, in human and murine models of
A
B
CD
E
Figure 5. GPIbα derived from platelet-derived extracellular vesicles supports monocytes rolling on von Wlllebrand Factor. (A) Representative plots of CRP_XL (1 mg/mL) generated-GPIbα+ platelet- derived extracellular vesicles (PEV) bound to monocytes measured by flow cytometry. (B-D) Representative pictures of monocytes (B), mono- cytes bearing CRP_XL (1 mg/mL) generated-PEV (C) and monocytes bearing PEV with GPIbα block- ade (clone 6B4, 20 mg/mL) (D) recruited on von Willebrand Factor (VWF) under flow conditions. (E) Total adhesion of monocytes with or without PEV and GPIbα blockade on VWF in flow conditions, n>10. Data are mean ± standard error of the mean (SEM). **P≤ 0.01 by ANOVA and Bonferroni post-test.
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