An orbital shear platform for real‐time, in vitro endothelium characterization

Abstract

Electrical impedance techniques have been used to characterize endothelium morphology, permeability, and motility in vitro. However, these impedance platforms have been limited to either static endothelium studies and/or induced laminar fluid flow at a constant, single shear stress value. In this work, we present a microfabricated impedance sensor for real-time, in vitro characterization of human umbilical vein endothelial cells (HUVECs) undergoing oscillatory hydrodynamic shear. Oscillatory shear was applied with an orbital shaker and the electrical impedance was measured by a microfabricated impedance chip with discrete electrodes positioned at radial locations of 0, 2.5, 5.0, 7.5, 10.0, and 12.5 mm from the center of the chip. Depending on their radial position within the circular orbital platform, HUVECs were exposed to shear values ranging between 0.6 and 6.71 dyne/cm(2) (according to numerical simulations) for 22 h. Impedance spectra were fit to an equivalent circuit model and the trans-endothelial resistance and monolayer's capacitance were extracted. Results demonstrated that, compared to measurements acquired before the onset of shear, cells at the center of the platform that experienced low steady shear stress (∼2.2 dyne/cm(2) ) had an average change in trans-endothelial resistance of 6.99 ± 4.06% and 1.78 ± 2.40% change in cell capacitance after 22 hours of shear exposure; cells near the periphery of the well (r = 12.5 mm) experienced transient shears (2.5-6.7 dyne/cm(2) ) and exhibited a greater change in trans-endothelial resistance (24.2 ± 10.8%) and cell capacitance (4.57 ± 5.39%). This study, demonstrates that the orbital shear platform provides a simple system that can capture and quantify the real-time cellular morphology as a result of induced shear stress. The orbital shear platform presented in this work, compared to traditional laminar platforms, subjects cells to more physiologically relevant oscillatory shear as well as exposes the sample to several shear values simultaneously. Biotechnol. Bioeng. 2016;113: 1336-1344. © 2015 Wiley Periodicals, Inc.

Understanding mechanobiology in cultured endothelium: A review of the orbital shaker method

“…Starting with the work of Berson et al [ 35 ], numerical models of the orbital shaker system [ [30] , [31] , [32] , 34 , [36] , [37] , [38] , [39] , [40] , [41] , [42] , [43] , [44] , [45] , [46] ] have shown that WSS magnitude fluctuates with time, more so towards the edge of the well than at its centre, and that TAWSS is also greater towards the edge. Additionally, the direction of the instantaneous WSS vector rotates evenly through 360° at the centre of the well during each orbit whereas at the edge, vectors tend only to switch between forward and backward orientations along a line that is approximately parallel to the wall.…”

Investigation of Wall Shear Stress in Cardiovascular Research and in Clinical Practice—From Bench to Bedside

“…It can be used to measure a variety of cellular properties, e.g., behavior under flow [ 52 ]. Another setup is the combination of a small impedance chip with an orbital shaker to produce oscillatory shear, which allows the measurement of trans-endothelial resistance against varying shear stress conditions [ 53 ]. Furthermore, microfluidic chips, also known as “organ-on-a-chip” technology, is a further advanced form of in vitro systems, which may contribute to our understanding of EC responses to hemodynamics [ 54 ] (reviewed in [ 55 , 56 ]).…”

Detection of frequency-dependent endothelial response to oscillatory shear stress using a microfluidic transcellular monitor

“…Moreover, extrapolation of animal data to human conditions has been frustratingly controversial, particularly for complex diseases afflicting dynamic microenvironments prevalent with mechanochemical stimuli such as blood flow and inflammatory cytokines 13 . Overcoming traditional in vitro models that predominantly rely on static culture of vascular endothelial cells, current advanced in vitro studies on endothelial mechanotransduction have taken the magnitude and direction of OSS into consideration to complement existing animal model research 3 , 4 , 8 , 9 , 14 – 16 . The frequencies typically associated to be characteristic of disease-causing OSS are based on measured physiological values in vivo ; this value is about 5 Hz in mice, whereas in humans this level drops to approximately 1 Hz 3 , 10 , 17 .…”