MORPHOLOGICAL ANALYSIS OF GLYCOCALYX ON A HUMAN BLOOD-BRAIN BARRIER MODEL
Afnan Al Doghmi Mohammad Rashed1, 2, Ana Raquel Santa-Maria1, 3, Fruzsina R. Walter1, András Dér1, Mária A. Deli1
Blood brain barrier (BBB) is formed by capillary brain endothelial cells (BECs) which create a dynamic interface between the blood and the brain and have complex interaction with the neighboring cells. Microvessel forming BECs are constantly exposed to mechanical forces generated by blood flow. The surface glycocalyx is one of the mechano-sensors that have been identified on the endothelial surface. We found that the surface charge of primary isolated BECs is highly negative, due to the presence of the negatively charged lipid heads in the cell membrane and the surface glycocalyx. Several in vitro models have been developed to better characterize and understand functions of the BBB, but only few models are dynamic and use continuous fluid flow like our lab-on-a-chip device (Walter et al, 2016). During our experiments we used this chip system in combination with the stem cell derived CD34+ human endothelial cells in co-culture with bovine pericytes (Cecchelli et al, 2014). Our goal was to characterize the glycocalyx in this human BBB model in mono- and co-culture, and after fluid flow. The glycocalyx was stained with wheat germ agglutinin lectin binding to N-acetyl glucosamine and sialic acid. The surface charge of the stem cell derived CD34+ human endothelial cells was measured using the Zetasizer Nano (Malvern) instrument. We could observe a difference in the intensity of the glycocalyx staining between mono- and co-culture of the model. Cell surface zeta values were also lower in cells derived from the co-culture compared to the mono-culture. Our study helps to further characterize this widely used BBB model.