PB0854 - The influence of platelets on monocyte inflammatory phenotype and function

Background: Increased numbers of platelet-monocyte complexes have been implicated in a range of thrombo-inflammatory diseases such as myocardial infarction, with higher numbers relating to worsened clinical outcomes. Several studies suggest that platelets may alter a monocyte’s inflammatory function. However, characterisation of these phenotypic changes and their functional implications following thrombotic stimuli is yet to be fully elucidated.

Aims: To investigate platelet-monocyte complex (PMC) formation and the resulting changes in phenotype and function.

Methods: Whole blood, or isolated monocytes/peripheral blood mononuclear cells (PBMCs) with washed platelets (1:100), were incubated with collagen related peptide (CRP; 1µg/ml) for up to 2 hours at 37ºC. Cell surface immunophenotyping (22 colour, 18 parameter panel), intracellular cytokine production (TNF-⍺ and MCP-1 in the presence of 3µM Monensin) and phagocytosis ability (using pHrodo bioparticles) were all assessed by flow cytometry. Data were analysed using FlowJo v.10 and Graphpad Prism.

Results: Platelet GPVI-specific stimulation results in rapid increase in platelet-monocyte complex (PMC) formation in both whole blood and isolated cells. These PMC’s have significantly higher surface expression of CD16, CCR2 and CXCR4 (p < 0.05) than unbound monocytes; consistent with adoption of a pro-inflammatory phenotype. PMCs produce higher levels of cytokines TNF-⍺ and MCP-1, as well as demonstrating increased phagocytic capabilities when compared to unbound monocytes. This correlates with increased surface expression of CD36 (p < 0.05) which is important in the phagocytosis process. They also show an upregulation of CD142 (tissue factor) suggesting a role for these complexes in thrombosis.

Conclusion(s): Our data suggests that in response to CRP stimulation the resulting platelet-monocyte complexes rapidly ( < 2 hours) adopt a more pro-inflammatory and pro-coagulative phenotype. Additional work is required to explore these changes and to further characterise platelet-monocyte complexes and their contribution to thrombo-inflammation.