Alteration of platelet GPVI signaling in ST-elevation myocardial infarction patients demonstrated by a combination of proteomic, biochemical, and functional approaches.

Paula Vélez,José María García-Acuña,Lilian Grigorian-Shamagian,Esteban Guitián,Isaac Rosa,Raymundo Ocaranza-Sánchez,José Ramón González-Juanatey,Ángel García,Diego López-Otero

doi:10.1038/srep39603

Abstract

The platelet-specific collagen receptor glycoprotein VI (GPVI) is critical for the formation of arterial thrombosis in vivo. We analyzed GPVI-activated platelets from ST-elevation myocardial infarction (STEMI) patients and matched stable coronary artery disease (SCAD) controls in order to provide novel clues on the degree of involvement of GPVI signaling in the acute event. Firstly, platelets were isolated from systemic venous blood and activated with the GPVI specific agonist CRP (collagen-related peptide). STEMI and SCAD samples were compared by a phosphoproteomics approach. Validations were by immunoblotting in systemic and intracoronary blood from independent cohorts of patients. Twenty-six differentially regulated proteins were identified when comparing CRP-activated systemic platelets from STEMI and SCAD patients, 4 of which were selected for validation studies: PLCɣ2, G6f, SLP-76, and Dok-2. Immunoblot analyses showed these four proteins had higher tyrosine phosphorylation levels in response to CRP in platelets from STEMI patients, being these levels more pronounced at the culprit site of coronary artery occlusion. Moreover, platelet aggregation studies showed a higher response to GPVI agonists in STEMI patients compared to SCAD controls. In conclusion, we show an altered activation state of GPVI signaling in STEMI patients, confirming this receptor as a promising anti-thrombotic target for myocardial infarction.

Highlights

ST-elevation myocardial infarction (STEMI) is a particular type of acute coronary syndrome (ACS) that involves intracoronary artery occlusion by a growing thrombus
We focused on stained bands of higher intensity in the lane corresponding to collagen-related peptide (CRP)-stimulated platelets from STEMI patients, comparing to the equivalent stable coronary artery disease (SCAD) control (Fig. 1A); those bands seemed to correlate with increased tyrosine phosphorylated bands in the immunoblot run in parallel (Fig. 1B), such correlation is difficult to establish with total certainty due to intrinsic differences between both images
The principal findings of the present study are: (i) proteomic and biochemical identification of a panel of tyrosine-phosphorylated signaling proteins reflecting altered glycoprotein VI (GPVI) activation levels in STEMI patients compared to SCAD matched controls; (ii) increased aggregation levels of CRP, and collagen-activated platelets in STEMI

Summary

Introduction

ST-elevation myocardial infarction (STEMI) is a particular type of acute coronary syndrome (ACS) that involves intracoronary artery occlusion by a growing thrombus. Collagenous structures in the core region of human atheromatous plaque[5] The inhibition of this receptor results in protection against atherosclerosis in rabbit and mice models[6] and against thrombus formation on injured arteries[7,8,9]. GPVI signaling activation was studied in platelets from intracoronary culprit-site arterial blood taken from STEMI patients during PCI compared to platelets from peripheral blood taken from the radial artery of the same patients The latter was to test if platelets from blood in direct contact with the thrombus at the occluded coronary artery could reflect better the GPVI signaling activation levels compared to peripheral platelets; this would provide further clues on the relevance of this receptor in the pathological events associated with an acute myocardial infarction

Objectives

Methods

Results

Conclusion