(455j) Endothelial Cell Migratory Response to Simple and Spatial Gradients in Wall Shear Stress

Surya, V., Stanford University
Ostrowski, M. A., Stanford University
Dunn, A. R., Stanford University
Fuller, G. G., Stanford University

AIChE: Annual Meeting, Salt Lake City, UT. November 8-13,
Session: Bio-Fluid Dynamics
Talk Title: Endothelial cell migratory response to simple and spatial
gradients in wall shear stress
Keywords: Endothelial cell, cell migration, wall shear stress, fluid
Vinay N. Surya, Maggie A. Ostrowski, Eleftheria Michalaki, Alexander R.
Dunn, Gerald G. Fuller

Endothelial cells (ECs) line the inner surface of
blood and lymphatic vessels and are highly sensitive to fluid flow as part of
their physiological function. EC orientation with flow, migration and vessel
development are profoundly influenced by hydrodynamic stresses, affecting the
ability to develop capillaries through angiogenesis and formation of valves
through valvulogenesis. How ECs sense fluid flow is a central and unanswered
question in cardiovascular research with important implications in
cardiovascular disease and tumor metastasis.

Previously, we developed a high-throughput flow
chamber which applies low Reynolds number impinging flow to a monolayer of ECs.
This flow profile is physiologically relevant to vessel bifurcations, where the
spatial gradient in wall shear stress may mechanically trigger angiogenic and
valvulogenic cues (Figure 1). Using this device, we previously found that human
lymphatic microvascular endothelial cells migrate against the flow direction,
towards regions of higher shear stress. The gradient in wall shear stress also
triggers the same physiological response as was observed in in vivo
mouse embryonic studies, including increased expression of the two
transcription factors involved in valve formation,  prospero-homeobox protein 1
and forkhead box protein C2, indicating a plausible connection between mechanically
triggered cell migration and vessel structure formation.

We have used the high-throughput flow chamber to probe
how endothelial cells from different regions of the human vasculature respond
to spatial gradients in wall shear stress. We have examined how coronary
artery, aortic, umbilical vein, and blood and lymphatic microvascular ECs respond
to both simple shear and gradients in shear stress in an effort to determine
similarities and differences in the migratory responses (Figure 2). We have
found that while the lymphatic microvascular ECs uniquely migrate against the
flow, large and small blood vessel ECs display very different migratory
responses to the same shear stress gradient. Large vessel blood ECs align and
move with the flow direction, while blood microvascular ECs adopt an azimuthal
orientation to the flow direction with no preference for migration direction.

Ongoing work aims to develop microfluidic channels
with physiologically relevant geometries that mimic the complex flows found in
vivo. We are currently pursuing constricting geometries such as those found at
the sites of valve formation in the venous and lymphatic systems, and U-shaped
geometries, which model the low and high curvature geometries characteristic of
Dean flows in the descending aorta. In both cases, we hypothesize that the
cellular response to fluid flow will depend on the absolute magnitude and
spatial gradient in wall shear stress, a topic which to our knowledge is
relatively unexplored despite its physiological significance.

Figure 1. A conceptual schematic of the relevant
vessel fluid flows experienced at a vessel bifurcation, which can be broken
down into two simple flow profiles: Poiseuille and impinging flow. A monolayer
of endothelial cells (red) are seeded on a gelatin-coated coverslip, submerged
in cell culture medium (light blue) and experience fluid flow of the cell
culture medium. Brightfield and fluorescent microscopy are used to capture
migratory response over time.

Figure 2. (A) The impinging flow device uses a
submerged nozzle to flow media (light blue) over ECs (red) seeded on a glass
substrate (dark blue). Six identical jets provide ease of use with standard
6-well tissue culture plates. (B) Finite element simulation (COMSOL) shows the
cross-sectional fluid velocity profile. Fluid streamline arrows (red) indicate
that the cells experience a spatially varying, axisymmetric flow. (C) An impinging
flow chamber produces a spatially conserved, highly tunable wall shear stress
profile (controlled by pump flow rate) to produce physiologically relevant
gradients in wall shear stress. Wall shear stress (blue) increases to a maximum
at ~400 microns from the stagnation point, then decays to zero. (D) Endothelial
cells from different vessel regions display widely varying responses to the
same spatial gradient in wall shear stress after 20 hours of exposure to impinging
flow. The total distance a cell travels while exposed to impinging flow is
plotted against its radial displacement, i.e. radial movement towards the jet
center (negative value) or movement away from the jet center (positive value).
N = 100 cells per cell type. Of the cell types tested, only lymphatic
endothelial cells collectively migrate upstream, against the direction of fluid