PB0846 - ZO-2 Enriched Tight Junction-like Structures at Sites of Platelet-Platelet Contact

Monday, July 11, 2022

6:30 PM – 7:30 PM

Location: Exhibition

Presentation Format: In-Person

Co-Author(s)

Magdolna Nagy, PhD

Postdoctoral Researcher
Coagulation Profile, Maastricht, the Netherlands
Maastricht, Limburg, Netherlands
MB

Markus Bender

Principal Investigator
Institute of Experimental Biomedicine, University Hospital Würzburg and Rudolf Virchow Center for Integrative and Translational Bioimaging, University of Würzburg
Würzburg, Bayern, Germany
NP

Natalie Poulter

Research Fellow
Institute of Cardiovascular Sciences, College of Medical and Dental Sciences, University of Birmingham, Edgbaston, Birmingham, United Kingdom; Centre of Membrane Proteins and Receptors (COMPARE), Universities of Birmingham and Nottingham, Midlands, United Kingdom
Birmingham, England, United Kingdom
AS

Albert Sickmann

President
ISAS
Dortmund, Nordrhein-Westfalen, Germany
XS

Xavier Stéphenne

Professor, chef de clinique Saint-Luc
Laboratoire d'Hépatologie Pédiatrique et Thérapie Cellulaire, Unité PEDI, Institut de Recherche Expérimentale et Clinique, Université catholique de Louvain (UCLouvain), Brussels, Belgium
Woluwe-Saint-Lambert, Brussels Hoofdstedelijk Gewest, Belgium
SB

Sanne Brouns

-
Department of Biochemistry, Cardiovascular Research Institute Maastricht (CARIM), Maastricht University, Maastricht, The Netherlands
Maastricht, Limburg, Netherlands
Rory R. Koenen, PhD

Professor
Department of Biochemistry, Cardiovascular Research Institute Maastricht, Maastricht University
Maastricht, Limburg, Netherlands
Hugo ten Cate, MD, PhD, FAHA

Professor, MD, PhD
Thrombosis Expertise Center, Heart & Vascular Center, Maastricht University Medical Center, Maastricht, the Netherlands
Maastricht, Limburg, Netherlands
JW

Johan W. M. Heemskerk

Emeritus Professor
Synapse research insitute
MAASTRICHT, Limburg, Netherlands

Presenting Author(s)

Constance Baaten, PhD

postdoctoral researcher
Department of Biochemistry, Cardiovascular Research Institute Maastricht, Maastricht University, Maastricht, The Netherlands
Maastricht, Limburg, Netherlands

Background: Platelets express junction proteins that are known to regulate cell-cell adhesion, barrier formation and communication in tissue forming cells. Previous research has shown that the gap junction proteins expressed by platelets regulate platelet activation in an αIIbβ3-dependent manner. Also, platelet activation processes are negatively regulated by endothelial cell-selective adhesion molecule (ESAM) and junctional adhesion molecule A (JAM-A), proteins that are part of the tight junctions (TJ) of endothelial cells. However, the role of zonula occludens 2 (ZO-2) -an intracellular component of the endothelial TJs- in the formation of inter-platelet contacts has not yet been described.

Aims: To unravel the molecular composition, in particular ZO-2, of tight platelet-platelet contacts.

Methods: Using phosphoproteomics databases, ZO-2 phosphorylation in activated platelets was investigated. Isolated platelets were allowed to fully spread on a fibrinogen surface. Fixed samples were stained to assess F-actin polymerization in relation to ZO-2 and ESAM distribution using confocal and/or super-resolution fluorescence microscopy. Platinum replica electron microscopy was applied for ultrastructural visualization of the cytoskeleton at sites of platelet-platelet interaction.

Results: ZO-2 undergoes extensive phosphorylation changes upon agonist-induced platelet activation as well as cAMP-mediated platelet inhibition. Confocal and super-resolution fluorescence microscopy indicated a marked redistribution and clustering of ZO-2 molecules at sites of platelet-platelet contacts, in which ZO-2 colocalized with ESAM. Furthermore, platinum replica electron microscopy revealed that inter-platelet contacts resulted in the merging of the circumferential actin bundles between interacting platelets. These phenomena were antagonized by cAMP elevation.

Conclusion(s): Jointly, these data point to the assembly of tight junction-like structures, where intracellular components of tight junctions accumulate at sites of close platelet-platelet contact.

This work was supported by the Dutch Heart Foundation (2020T020 to C.B.)