Improvement and validation of a computational model of flow in the swirling well cell culture model

Abstract

Effects of fluid dynamics on cells are often studied by growing the cells on the base of cylindrical wells or dishes that are swirled on the horizontal platform of an orbital shaker.The swirling culture medium applies a shear stress to the cells that varies in magnitude and directionality from the center to the edge of the vessel. Computational fluid dynamics methods are used to simulate the flow and hence calculate shear stresses at the base of the well. The shear characteristics at each radial location are then compared with cell behavior at the same position. Previous simulations have generally ignored effects of surface tension and wetting, and results have only occasionally been experimentally validated. We investigated whether such idealized simulations are sufficiently accurate, examining a commonly-used swirling well configuration. The breaking wave predicted by earlier simulations was not seen, and the edge-to-center difference in shear magnitude (but not directionality) almost disappeared, when surface tension and wetting were included. Optical measurements of fluid height and velocity agreed well only with the computational model that incorporated surface tension and wetting. These results demonstrate the importance of including accurate fluid properties in computational models of the swirling well method.

NO Synthesis but Not Apoptosis, Mitosis or Inflammation Can Explain Correlations between Flow Directionality and Paracellular Permeability of Cultured Endothelium

“…The rate at which permeability fell with increasing distance from the centre was initially slow but increased after 2 mm; permeability reached a plateau by 5 mm that was maintained out to 8 mm, the maximum distance examined. As previously discussed [ 23 ], this pattern of permeability does not strongly resemble the trend in either the time-averaged WSS, which is approximately constant, or the oscillatory shear index (OSI), which decreases in a nearly linear fashion [ 33 ]. There is also a poor correlation with transverse WSS (transWSS, a measure of multidirectionality which averages those components of WSS during one cycle that act at right angles to the mean WSS direction [ 35 ]): it, like the time-averaged WSS, is approximately constant over the 0–8 mm range.…”

“…Here, we sought to investigate the mechanisms underlying these phenomena, and specifically the roles of apoptosis, proliferation, mitosis, proinflammatory transcription factors and NO production, all of which have been implicated in determining endothelial permeability. The study employed arterial endothelial cells of low passage number [ 23 ], swirling well configurations for which shear stresses have been characterised by computational fluid dynamics [ 33 ], chronic rather than acute exposure to flow [ 20 , 23 ], staining for both early- and late-stage apoptosis, and an avidin-based tracer that binds to the biotinylated endothelial substrate and whose accumulation can consequently be compared to the state of the individual cells immediately overlying it [ 23 , 34 ].…”

“…The platform translated with an orbital diameter of 10 mm in the horizontal plane and a rotation rate of 150 rpm. Flow characteristics were obtained using computational simulations that included surface tension and wetting [ 33 ]. Static controls for each experiment were carried out using cells from the same aorta.…”

Haemodynamic Wall Shear Stress, Endothelial Permeability and Atherosclerosis—A Triad of Controversy

“…The flow can be altered by changing the orbital radius and velocity, the dish radius and the depth of medium, but it is difficult to separate the effects of different shear metrics, several of which change with increasing radial distance from the centre. Furthermore, although different flow regimes have been identified and approximate analytical methods devised ( Alpresa et al, 2018a ; 2018b ), accurate description of the shear acting on the base of the well requires CFD models that include the orbital forcing, the free surface of the gravitational wave ( Berson et al, 2008 ) and, in some cases, the surface tension ( Arshad et al, 2022 ).…”

“…The swirling well method was used to investigate effects of different types of flow on transendothelial transport of the three avidin-based tracers. The choice of culture plate, medium depth and orbital parameters gave a large difference in the directionality of WSS from the centre to the edge of the well, but a negligible change in time average WSS, according to numerical simulations that incorporated surface tension ( Arshad et al, 2022 ). Transport of the small and medium-sized macromolecular tracers through intercellular junctions was increased in the centre of the well (multidirectional flow) and decreased at its edge (uniaxial flow) compared to static controls ( Ghim et al, 2017 ).…”