“Next-generation Toolbox for Studying Platelet Activation: New Labels, Label-free Methods, and Single Cell Approaches”
Ilya Reviakine, Ph.D.
Department of Bioengineering
University of Washington
Cardiovascular disorders (CVDs) are currently the number one cause of death world-wide. A key role in their etiology is played by the platelets (thrombocytes)—anuclear 3 – 4 μm cell fragments circulating in the blood. Their function is to limit traumatic blood loss due to injury by controlling blood coagulation (hemostasis), but under pathological conditions their activation leads to thrombotic complications that cause heart attacks and stroke. Antiplatelet therapy is used to manage CVDs with considerable success, but existing approaches suffer from a number of limitations (e.g., bleeding complications and high patient monitoring costs). This drives the search for better, more selective antiplatelet agents. Furthermore, platelets perform numerous nonhemostatic functions in wound healing and tissue regeneration, innate and adaptive immune response, cancer metastasis, angiogenesis, and vascular remodeling. Important role in the regulation of these functions is thought to be played by the secretion reactions that accompany platelet activation. Current understanding of platelet physiology is rooted in the quantitative functional assays introduced in ~ 1960s and flow cytometry analysis using monoclonal antibodies against platelet surface activation markers. These assays are limited in their ability to disentangle individual activation pathways in the complex platelet signaling network, and in their focus on the hemostatic functions of the platelets. New assays are needed to catalyze advances in the understanding of platelet physiology so that treatment strategies could be developed based on their regenerative functions and other physiological roles. I will present several examples of new in vitro assays for studying platelet activation and secretion that are being developed in my group—based on studying changes in platelet surface glycosylation, single platelet microfluidics, and label-free techniques such as time-of-flight secondary ion mass spectrometry (ToF-SIMS).
Ilya Reviakine received his undergraduate degree in biochemistry from McMaster University (Hamilton, Canada) and his Ph.D. in Mathematics and Natural Sciences from the University of Groningen (the Netherlands), where he worked with Prof. Alain Brisson on atomic force microscopy of biological macromolecules and their assemblies. He studied the assembly annexin A5 (formerly annexin V) ordered arrays on supported lipid bilayers (SLBs), as well as the mechanism of SLB formation from liposomes. He did a postdoc in the area of biointerfaces at the Swiss Federal Institute of Technology (ETH) in Zürich, Switzerland, where he further worked on lipid interactions with artificial materials found in implants, in particular titania (TiO2), that is thought to confer favorable properties to the widely used titanium implants. That work led to the discovery of titania‐supported asymmetric SLBs containing phosphatidylserine (PS). PS is a phospholipid that is important in blood coagulation, making this phenomenon potentially interesting to the area of material biocompatibility. Indeed, the mechanism by which PS‐containing asymmetric SLBs formed on titaia pointed to the role of surface ion exchange in the interactions between artificial materials and biological model systems. (Native cell membranes are also asymmetric with respect to the distribution of this phospholipid, making titania‐supported SLBs interesting for cell membrane biophysics studies). After a postdoc on protein crystallization at the University of Houston (Houston, TX, USA), Reviakine returned to the work on biointerfaces as an Alexander von Humboldt Research fellow, hosted by Diethelm Johannsmann at the Clausthal University of Technology in Germany in 2005. There, he started to work on shear‐acoustic resonators that are used, for example, in the quartz crystal microbalance (QCM or QCM‐D). His work focused on the shear‐acoustic response from ultrathin, laterally heterogeneous films such as adsorbed liposomes, viruses, nanoparticles, and some proteins. In 2006, Reviakine joined CIC biomaGUNE, a new research institute in San Sebastian, Spain, as a groupleader. There, he had a chance to participate in the establishment of a new institute and continued his work on surface acoustic sensing and, more generally, on the interactions between artificial materials and biological systems, ranging from lipids to thrombocytes (platelets) to blood. Highlights of his work include the identification of a new (hydrodynamic) contrast mechanism by which shear acoustic resonators used in quartz crystal microbalances sensed adsorbed nanoparticles, which later allowed the deformation of surface‐adsorbed liposomes to be measured; design of a new platelet activation assay based on the analysis of platelet surface glycosylation; and the investigation of the role surface ion exchange played in the activation of platelets on TiO2. Platelets are key coagulation effectors (thus the connection with the earlier work on phosphatidylserine) responsible for pathological thrombosis occurring in cardiovascular disorders (CVDs) and adverse responses to biomaterials used to treat CVDs—responses such as material‐induced thrombosis and inflammation. Building on the understanding of platelets and platelet‐biomaterial interactions he developed, he moved to KIT in 2013 to start a group on blood compatibility. Achievements of his KIT group include the design of a single platelet secretion assay and a multiparametric whole blood assay to study the interaction between blood and metallic biomaterials (TiO2, CoCr, steel). The key idea driving his research is that platelets’ regenerative functions can be harnessed to foster implant integration through controlling their activation at biomaterial surfaces.
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