P2Y12 receptor blockers are anti-inflammatory drugs inhibiting both circulating monocytes and macrophages including THP-1 cells

Patrick M Siegel,Alba Fricke,Ya-Lan Ying,Philipp Diehl,Prerna Sharma,Laura Sander,Steffen U Eisenhardt,Xiaowei Wang,Christoph B Olivier,Karlheinz Peter,Christoph Bode,Nicole Bassler,Ingo Ahrens,Johannes Stamm

doi:10.1038/s41598-021-95710-3

Abstract

P2Y12 blockade improves patient outcomes after myocardial infarction. As well as antithrombotic effects, anti-inflammatory effects may contribute to this beneficial clinical outcome. Here we aimed to identify potential anti-inflammatory effects of P2Y12 receptor blockers on monocytes and macrophages. Using flow cytometry, migration assays, flow chambers and RNA microarrays, we investigated the effects of adenosine diphosphate (ADP) and P2Y12 receptor blockers on blood monocytes, THP-1 monocytes and THP-1 monocytes after differentiation to macrophages. P2Y12 -expressing platelets can form aggregates with monocytes in circulating blood. Mediated by platelets, ADP results in activation of the integrin receptor Mac-1 on blood monocytes, as detected by the conformation-specific single-chain antibody MAN-1. Via the same association with platelets, THP-1 monocyte adhesion to the endothelial intercellular adhesion molecule 1 (ICAM-1) is induced by ADP. P2Y12 receptor blockers prevent these ADP effects on monocytes. Interestingly, in contrast to THP-1 monocytes, THP-1 monocytes, after differentiation to macrophages, directly expressed the P2Y12 receptor and consequently ADP was found to be a potent chemoattractant. Again, P2Y12 receptor blockers antagonised this effect. Accordingly, stimulation of THP-1 macrophages with ADP caused a substantial change in gene expression pattern and upregulation of several genes associated with inflammation and atherogenesis. These data establish novel anti-inflammatory effects of P2Y12 receptor blockers on monocytes and macrophages, which are expected to contribute to cardiovascular risk reduction.

Highlights

P2Y12 blockade improves patient outcomes after myocardial infarction
We hypothesised that monocyte activation is mediated by adenosine diphosphate (ADP) indirectly via platelets attached to monocytes
To investigate whether blood monocytes can be activated via a plateletdependent pathway, whole blood was stimulated with ADP (Fc 20 μM) and Mac-1 activation on CD14+ monocytes was assessed in flow cytometry using the activation-specific single-chain antibody MAN-1 (Fig. 1b)

Summary

Introduction

P2Y12 blockade improves patient outcomes after myocardial infarction. As well as antithrombotic effects, anti-inflammatory effects may contribute to this beneficial clinical outcome. Stimulation of THP-1 macrophages with ADP caused a substantial change in gene expression pattern and upregulation of several genes associated with inflammation and atherogenesis These data establish novel anti-inflammatory effects of P2Y12 receptor blockers on monocytes and macrophages, which are expected to contribute to cardiovascular risk reduction. ADP stimulation of the Gi-coupled P2Y12 receptor leads to activation of PI 3-kinase, Akt, ERK, Rap1b, Src family kinases and G protein-gated inwardly rectifying potassium channels resulting in activation of GPIIb/IIIa and platelet a ggregation[17] Due to their anti-platelet effect, P2Y12 receptor blockers (e.g. clopidogrel, prasugrel, ticagrelor and cangrelor) are recommended for use in patients after percutaneous coronary intervention and in ACS to avoid thrombotic complications, stent thrombosis[18]. The mechanisms, by which P 2Y12 receptor blockers mediate these anti-inflammatory effects are currently poorly u nderstood[18,23]

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