Eur J Anat, 20 (2): 121-130 (2016)

Morphological studies on platelet function using a microfluidic chip. A proof of concept study

Teresa Nuñez Lopez¹, Isabel Cortegano Jimeno², Laura Lopez Gomez1, Gilberto del Rosario³, Maria Luisa Gaspar Alonso-Vega²,Pedro Mestres-Ventura^4,1

¹Área de Histología. Departamento de Ciencias Básicas de la Salud. Universidad Rey Juan Carlos, Alcorcón (Madrid, Spain), ²Unidad de Inmunobiologia, Instituto Carlos III, Majadahonda (Madrid, Spain), ³Laboratorio de Microscopia Electrónica, Centro de Apoyo Tecnológico, Universidad Rey Juan Carlos, Móstoles (Madrid, Spain), ⁴Department of Anatomy and Cell Biology, Saarland University, Homburg (Germany)

ABSTRACT Platelets are blood cellular components involved in hemostatic processes and thrombus formation. Activation and inhibition of platelets result in an increase in morphological changes and a significant reduction in adhesion. There are several approaches towards the determination of the functional status of platelets, based on criteria such as cell adhesion, molecular changes at the cell surface, etc. In recent years, microfluidic devices have been introduced to mimic conditions proper to the vascular system, and so emulate thrombus formation in vivo. This study presents a microchip, the Thrombi Chip®, which is partially fitted with fluidic properties. This microchip has various types of micro-channels into which the platelets are inserted and, after drug treatment, the investigation is completed with the examination of the chip under an invert light microscope.For microscopy, cells were labeled with FCDA (human platelets) and Rho6G (mouse platelets). Counts and morphometric measurements of the adhered cells were carried out using digital images. To validate the results obtained with the microchip, the fractions of mice platelets were investigated with flow cytometry as well. Scanning electron microscopy was used to examine the morphological changes related to activation and inhibition in human platelets.The results show that, with this microchip, activation and inhibition of platelets can be detected. Flow cytometry studies largely confirm the microchip results. Certain variability in the results observed in human platelets is considered normal, as donors were randomized. In this respect the mouse platelets were much more uniform.Measurements with the microchip require that the sample be divided into three groups: control, activated and inhibited, resulting in a set of data, which, after respective evaluation, provides activity profiles, giving information on the status and response capacity of a sample. Such profiles could have diagnostic relevance and therefore be useful in a clinical context, for example in the monitoring of the effects of short- and long-term treatment of patients, as well as to test new drugs.

Keywords: Micro-fluidic chip, Platelet activation, Platelet inhibition, Cell adhesion, Cell shape, Thrombosis and hemostasis