Spatial Shear Stress Gradient Research Articles

Disturbed fluid flow reinforces endothelial tractions and intercellular stresses

Disturbed fluid flow is well understood to have significant ramifications on endothelial function, but the impact disturbed flow has on endothelial biomechanics is not well understood. In this study, we measured tractions, intercellular stresses, and cell velocity of endothelial cells exposed to disturbed flow using a custom-fabricated flow chamber. Our flow chamber exposed cells to disturbed fluid flow within the following spatial zones: zone 1 (inlet; length 0.676–2.027 cm): 0.0037 ± 0.0001 Pa; zone 2 (middle; length 2.027–3.716 cm): 0.0059 ± 0.0005 Pa; and zone 3 (outlet; length 3.716–5.405 cm): 0.0051 ± 0.0025 Pa. Tractions and intercellular stresses were observed to be highest in the middle of the chamber (zone 2) and lowest at the chamber outlet (zone 3), while cell velocity was highest near the chamber inlet (zone 1), and lowest near the middle of the chamber (zone 2). Our findings suggest endothelial biomechanical response to disturbed fluid flow to be dependent on not only shear stress magnitude, but the spatial shear stress gradient as well. We believe our results will be useful to a host of fields including endothelial cell biology, the cardiovascular field, and cellular biomechanics in general.

Association of Shear Stress with Subsequent Lumen Remodeling in Hemodialysis Arteriovenous Fistulas.

Blood flow-induced wall shear stress is a strong local regulator of vascular remodeling, but its effects on arteriovenous fistula (AVF) remodeling are unclear. In this prospective cohort study, we used computational fluid dynamics simulations and statistical mixed-effects modeling to investigate the associations between wall shear stress and AVF remodeling in 120 participants undergoing AVF creation surgery. Postoperative magnetic resonance imaging data at 1 day, 6 weeks, and 6 months were used to derive current wall shear stress by computational fluid dynamic simulations and to quantify subsequent changes in AVF lumen cross-sectional area at 1-mm intervals along the proximal artery and AVF vein. Combining artery and vein data, prior mean wall shear stress was significantly associated with lumen area expansion. Mean wall shear stress at day 1 was significantly associated with change in lumen area from day 1 to week 6 (11% larger area per interquartile range [IQR] higher mean wall shear stress, 95% confidence interval [95% CI], 5% to 18%; n =101), and mean wall shear stress at 6 weeks was significantly associated with change in lumen area from 6 weeks to month 6 (14% larger area per IQR higher, 95% CI, 3% to 28%; n =52). The association of mean wall shear stress at day 1 with lumen area expansion from day 1 to week 6 differed significantly by diabetes ( P =0.009): 27% (95% CI, 17% to 37%) larger area per IQR higher mean wall shear stress without diabetes and 9% (95% CI, -1% to 19%) with diabetes. Oscillatory shear index at day 1 was significantly associated with change in lumen area from day 1 to week 6 (5% smaller area per IQR higher oscillatory shear index, 95% CI, 3% to 7%), and oscillatory shear index at 6 weeks was significantly associated with change in lumen from 6 weeks to month 6 (7% smaller area per IQR higher oscillatory shear index, 95% CI, 2% to 11%). Wall shear stress spatial gradient was not significantly associated with subsequent remodeling. In a joint model, wall shear stress and oscillatory shear index statistically significantly interacted in their associations with lumen area expansion in a complex nonlinear fashion. Higher wall shear stress and lower oscillatory shear index were associated with greater lumen expansion after AVF creation surgery.

Computational fluid dynamics of a novel perfusion strategy during hybrid thoracic aortic repair.

To mitigate the risk of perioperative neurological complications during frozen elephant trunk procedures, we aimed to computationally evaluate the effects of direct cerebral perfusion strategy through a left carotid-subclavian bypass on hemodynamics in a patient-specific thoracic aorta model. Between July 2016 and March 2019, 11 consecutive patients underwent frozen elephant trunk operation using the left carotid-subclavian bypass with a side graft anastomosis and right-axillary cannulation for systemic and brain perfusion. A multiscale model realized coupling three-dimensional computational fluid dynamics was developed and validated with in vivo data. Model comparison with direct antegrade cannulation of all epiaortic vessels was performed. Wall shear stress, wall shear stress spatial gradient, and localized normalized helicity were selected as hemodynamic indicators. Four cerebral perfusion flows were tested (6 to 15 mL/kg/min). Direct cerebral perfusion of the left subclavian bypass resulted in higher flow rates with augmented speeds in all epiaortic vessels in comparison with traditional perfusion model. At the level of the left vertebral artery (LVA), a speed of 22.5 vs 21 mL/min and mean velocity of 3.07 vs 2.93 cm/s were registered, respectively. With a cerebral perfusion flow of 15 mL/kg, lower LVA wall shear stress (1.596 vs 2.030 N/m2 ), and wall shear stress gradient (1445 vs 5882 N/m3 ) were observed. A less disturbed flow considering the localized normalized helicity was documented. No patients experienced neurological/spinal cord damages. Direct perfusion of a left carotid bypass proved to be cerebroprotective, resulting in a more physiological and stable anterior and posterior cerebral perfusion.

Lateral trapezoid microfluidic platform for investigating mechanotransduction of cells to spatial shear stress gradients

Microfluidic platforms enable the influence of a variety of chemical and physical gradients on single or multiple cells to be examined and monitored in real-time. Research into chemical gradients has been more prevalent in literature; however, with the interest in mechanotransduction, research investigating the influence of physical gradients, such as force from shear stress, are appearing. Shear stress studies, to date have been focused on using parallel plate flow chambers, or shear stress gradients spanning the length of the microchannel. In this paper, we designed a Trapezoid microchannel, which enables production of a unique lateral spatial shear stress gradient, in order to examine the responses it evokes in cells, using microscopy. We investigated the effect of a lateral spatial shear stress gradient on intracellular calcium signalling in HEK-293-TRPV4 cells. The Trapezoid microchannel was designed to produce a spatial shear stress gradient, spanning the width of the microchannel. Additionally, we investigated the effect of temporal shear stress using the microfluidic platform, to further examine the complexity of the transduction of shear stress forces, which mimic pathological and physiological conditions into cellular signalling. We show that the temporal pattern of shear stress, modulated intracellular calcium signalling, with slower response times (responding to stimulus and reaching maximum Calcium influx), compared to those observed in the absence of temporal shear stress. Our experiments highlight the importance of considering the nature of shear stress under physiological and pathological conditions – with microfluidic platforms providing a useful means to do so.

High resolution hemodynamic profiling of murine arteriovenous fistula using magnetic resonance imaging and computational fluid dynamics

BackgroundArteriovenous fistula (AVF) maturation failure remains a major cause of morbidity and mortality in hemodialysis patients. The two major etiologies of AVF maturation failure are early neointimal hyperplasia development and persistent inadequate outward remodeling. Although hemodynamic changes following AVF creation may impact AVF remodeling and contribute to neointimal hyperplasia development and impaired outward remodeling, detailed AVF hemodynamics are not yet fully known. Since murine AVF models are valuable tools for investigating the pathophysiology of AVF maturation failure, there is a need for a new approach that allows the hemodynamic characterization of murine AVF at high resolutions.MethodsThis methods paper presents a magnetic resonance imaging (MRI)-based computational fluid dynamic (CFD) method that we developed to rigorously quantify the evolving hemodynamic environment in murine AVF. The lumen geometry of the entire murine AVF was reconstructed from high resolution, non-contrast 2D T2-weighted fast spin echo MRI sequence, and the flow rates of the AVF inflow and outflow were extracted from a gradient echo velocity mapping sequence. Using these MRI-obtained lumen geometry and inflow information, CFD modeling was performed and used to calculate blood flow velocity and hemodynamic factors at high resolutions (on the order of 0.5 μm spatially and 0.1 ms temporally) throughout the entire AVF lumen. We investigated both the wall properties (including wall shear stress (WSS), wall shear stress spatial gradient, and oscillatory shear index (OSI)) and the volumetric properties (including vorticity, helicity, and Q-criterion).ResultsOur results demonstrate increases in AVF flow velocity, WSS, spatial WSS gradient, and OSI within 3 weeks post-AVF creation when compared to pre-surgery. We also observed post-operative increases in flow disturbances and vortices, as indicated by increased vorticity, helicity, and Q-criterion.ConclusionsThis novel protocol will enable us to undertake future mechanistic studies to delineate the relationship between hemodynamics and AVF development and characterize biological mechanisms that regulate local hemodynamic factors in transgenic murine AVF models.

Development of Novel Flow Chamber to Study Endothelial Cell Morphology: Effects of Shear Flow with Uniform Spatial Gradient on Distribution of Focal Adhesion

Fluid shear stress (SS) is well known to cause morphological changes in vascular endothelial cells (ECs) accompanied by alteration in actin cytoskeletal structure and distribution of focal adhesions. Recent studies have shown that spatial SS gradient also has effects on EC morphology, but the detailed mechanisms of EC responses to SSG remain unclear. In the present study, we sought morphological responses of ECs under SS and uniform SSG condition using a newly developed flow chamber. Confluent ECs were exposed to SS with SSG for 24 hours. Focal adhesions of the EC under SS without SSG were localized in the cell periphery. In contrast, focal adhesions were expressed not only in the periphery but also in interior portion of cells after exposure to SS with SSG. Unlike ECs exposure to SS developed thick actin filaments aligned to the direction of flow no development of thick actin filaments but thin and short filaments were observed in ECs after 24-hour exposure to SS with SSG. Since the distribution of focal adhesion is of critical importance for development of actin filaments and cell morphological changes, these results suggest that SSG suppresses redistribution of focal adhesions, resulting in the inhibition of EC morphological changes and development of thick actin filaments in response to flow.

High Fluid Shear Stress and Spatial Shear Stress Gradients Affect Endothelial Proliferation, Survival, and Alignment

Cerebral aneurysms develop near bifurcation apices, where complex hemodynamics occur: Flow impinges on the apex, accelerates into branches, then slows again distally, creating high wall shear stress (WSS) and positive and negative spatial gradients in WSS (WSSG). Endothelial responses to these kinds of high WSS hemodynamic environments are not well characterized. We examined endothelial cells (ECs) under elevated WSS and positive and negative WSSG using a flow chamber with constant-height channels to create regions of uniform WSS and converging and diverging channels to create positive and negative WSSG, respectively. Cultured bovine aortic ECs were subjected to 3.5 and 28.4Pa with and without WSSG for 24 and 36h. High WSS inhibited EC alignment to flow, increased EC proliferation assessed by bromodeoxyuridine incorporation, and increased apoptosis determined by terminal deoxynucleotidyl transferase dUTP-mediated nick-end labeling. These responses to high WSS were either accentuated or ameliorated by WSSG: Positive WSSG (+980Pa/m) inhibited alignment and stimulated proliferation and apoptosis, whereas negative WSSG (-1120Pa/m) promoted alignment and suppressed proliferation and apoptosis. These results demonstrate that ECs discriminate between positive and negative WSSG under high WSS conditions. EC responses to positive WSSG may contribute to pathogenic remodeling that occurs at bifurcations preceding aneurysm formation.

Application of a locally conservative Galerkin (LCG) method for modelling blood flow through a patient‐specific carotid bifurcation

AbstractIn the present work, blood flow through a patient‐specific carotid bifurcation is thoroughly analysed. A locally conservative Galerkin spatial discretization is applied along with an artificial compressibility and characteristic‐based split scheme to solve the 3D incompressible Navier–Stokes equations. Boundary layer meshes are introduced to accurately resolve high near‐wall velocity gradients inside a patient‐specific carotid bifurcation. A total of six haemodynamic wall parameters have been brought together to analyse the regions of possible atherogenesis within the domain. The results show that wall shear stress (WSS) is high (6–15 Pa) near the apex, along with a small region where WSS exceeded 20 Pa. This peak WSS region is close to the inner wall of the external carotid artery (ECA). It is also clear from the results that low WSS occurred in the common carotid artery (CCA). High oscillatory shear occurred in the CCA distal to a local narrowing of the internal carotid artery and along the outer wall, indicating a potential region of atherogenesis. The highest values of wall shear stress angle deviation and wall shear stress angle gradient occur at the apex. The wall shear stress temporal gradient reached 4000 Pa/s near the apex, closer to the ECA. Wall shear stress spatial gradient distribution corresponded with the WSS distribution, with a maximum occurring at the apex. Copyright © 2010 John Wiley & Sons, Ltd.

Wall Shear Stress Distribution in Patient Specific Coronary Artery Bifurcation

Problem statement: Atherogenesis is affected by hemodynamic parameters, such as wall shear stress and wall shear stress spatial gradient. These parameters are largely dependent on the geometry of arterial tree. Arterial bifurcations contain significant flow disturbances. Approach: The effects of branch angle and vessel diameter ratio at the bifurcations on the wall shear stress distribution in the coronary arterial tree based on CT images were studied. CT images were digitally processed to extract geometrical contours representing the coronary vessel walls. The lumen of the coronary arteries of the patients was segmented using the open source software package (VMTK). The resulting lumens of coronary arteries were fed into a commercial mesh generator (GAMBIT, Fluent Inc.) to generate a volume that was filled with tetrahedral elements. The FIDAP software (Fluent Corp.) was used to carry out the simulation by solving Navier-Stokes equations. The FIELDVIEW software (Version 10.0, Intelligent Light, Lyndhurst, NJ) was used for the visualization of flow patterns and the quantification of wall shear stress. Post processing was done with VMTK and MATLAB. A parabolic velocity profile was prescribed at the inlets and outlets, except for 1. Stress free outlet was assigned to the remaining outlet. Results: The results show that for angle lower than 90°, low shear stress regions are observed at the non-flow divider and the apex. For angle larger than 90°, low shear stress regions only at the non-flow divider. By increasing of diameter of side branch ratio, low shear stress regions in the side branch appear at the non-flow divider. Conclusion: It is concluded that not only angle and diameter are important, but also the overall 3D shape of the artery. More research is required to further quantify the effects angle and diameter on shear stress patterns in coronaries.

Effect of spatial gradient in fluid shear stress on morphological changes in endothelial cells in response to flow

Arterial bifurcations are common sites for development of cerebral aneurysms. Although this localization of aneurysms suggests that high shear stress (SS) and high spatial SS gradient (SSG) occurring at the bifurcations may be crucial factors for endothelial dysfunction involved in aneurysm formation, the details of the relationship between the hemodynamic environment and endothelial cell (EC) responses remain unclear. In the present study, we sought morphological responses of ECs under high-SS and high-SSG conditions using a T-shaped flow chamber. Confluent ECs were exposed to SS of 2–10 Pa with SSG of up to 34 Pa/mm for 24 and 72 h. ECs exposed to SS without spatial gradient elongated and oriented to the direction of flow at 72 h through different processes depending on the magnitude of SS. In contrast, cells did not exhibit preferred orientation and elongation under the combination of SS and SSG. Unlike cells aligned to the flow by exposure to only SS, development of actin stress fibers was not observed in ECs exposed to SS with SSG. These results indicate that SSG suppresses morphological changes of ECs in response to flow.

Bubble Motion in a Blood Vessel: Shear Stress Induced Endothelial Cell Injury

Mechanisms governing endothelial cell (EC) injury during arterial gas embolism have been investigated. Such mechanisms involve multiple scales. We have numerically investigated the macroscale flow dynamics due to the motion of a nearly occluding finite-sized air bubble in blood vessels of various sizes. Non-Newtonian behavior due to both the shear-thinning rheology of the blood and the Fahraeus-Lindqvist effect has been considered. The occluding bubble dynamics lends itself for an axisymmetric treatment. The numerical solutions have revealed several hydrodynamic features in the vicinity of the bubble. Large temporal and spatial shear stress gradients occur on the EC surface. The stress variations manifest in the form of a traveling wave. The gradients are accompanied by rapid sign changes. These features are ascribable to the development of a region of recirculation (vortex ring) in the proximity of the bubble. The shear stress gradients together with sign reversals may partially act as potential causes in the disruption of endothelial cell membrane integrity and functionality.

Correlations Among Indicators of Disturbed Flow at the Normal Carotid Bifurcation

A variety of hemodynamic wall parameters (HWP) has been proposed over the years to quantify hemodynamic disturbances as potential predictors or indicators of vascular wall dysfunction. The aim of this study was to determine whether some of these might, for practical purposes, be considered redundant. Image-based computational fluid dynamics simulations were carried out for N=50 normal carotid bifurcations reconstructed from magnetic resonance imaging. Pairwise Spearman correlation analysis was performed for HWP quantifying wall shear stress magnitudes, spatial and temporal gradients, and harmonic contents. These were based on the spatial distributions of each HWP and, separately, the amount of the surface exposed to each HWP beyond an objectively-defined threshold. Strong and significant correlations were found among the related trio of time-averaged wall shear stress magnitude (TAWSS), oscillatory shear index (OSI), and relative residence time (RRT). Wall shear stress spatial gradient (WSSG) was strongly and positively correlated with TAWSS. Correlations with Himburg and Friedman's dominant harmonic (DH) parameter were found to depend on how the wall shear stress magnitude was defined in the presence of flow reversals. Many of the proposed HWP were found to provide essentially the same information about disturbed flow at the normal carotid bifurcation. RRT is recommended as a robust single metric of low and oscillating shear. On the other hand, gradient-based HWP may be of limited utility in light of possible redundancies with other HWP, and practical challenges in their measurement. Further investigations are encouraged before these findings should be extrapolated to other vascular territories.

Atheroprotective Signaling Mechanisms Activated by Steady Laminar Flow in Endothelial Cells

Many studies of the human vascular tree have shown that atherosclerosis develops at an early age, with areas of the posterior aorta and branch sites showing the earliest fatty streaks. Closer analysis of the branch points showed that the flow dividers, regions of high shear stress with laminar flow, were relatively protected from fatty streak formation. In contrast, there was a predilection for atherosclerosis at the curvatures and lateral walls of branch points.1 These are regions where disturbed flow occurs; importantly, when the time-averaged shear stress is low, the spatial shear stress gradient is very high.1,2 Both monocyte adhesion and endothelial cell (EC) apoptosis also are highest in these areas.3–6 Recently, Cheng et al7 developed a vascular occluder that induced 3 different regions of altered flow: low, high, and low with oscillatory flow. They observed that lesions always developed in regions of low compared with high shear stress. However, the low shear stress was more likely to develop larger lesions with more lipids, expression of inflammatory mediators, and matrix metalloprotease activity than oscillatory flow. When viewed in concert with a study that showed that regions of low shear stress already had a greater number of inflammatory cells resident in the artery wall as a result of increased trafficking,8 it is clear that low flow is proinflammatory and atherogenic. More sophisticated 3-dimensional analysis of flow patterns in human carotid bifurcations has identified prototypic arterial waveforms, “atheroprone” and “atheroprotective,” representing the wall shear stresses in the carotid sinus, which is susceptible to atherosclerotic lesion development, and the distal internal carotid artery, which is resistant.9 Characteristics of the atheroprone waveform include a high oscillatory shear index (≈0.45) and low time-averaged shear stress amplitude (≈0 dyne/cm2). Thus, both the amplitude and flow pattern are key determinants of …

Individual and combined effects of shear stress magnitude and spatial gradient on endothelial cell gene expression

The apparent tendency of atherosclerotic lesions to form in complex blood flow environments has led to many theories regarding the importance of hemodynamic forces in endothelium-mediated atherosusceptibility. The effects of shear stress magnitude and spatial shear stress gradient on endothelial cell gene expression in vitro were examined in this study. Converging-width flow channels were designed to impose physiological ranges of shear stress gradient and magnitude on porcine aortic endothelial cells, and real-time quantitative PCR was performed to evaluate their expression of five genes of interest. Although vascular cell adhesion molecule-1 expression was insensitive to either variable, each of the remaining genes exhibited a unique dependence on shear stress magnitude and gradient. Endothelial nitric oxide synthase showed a strong positive dependence on magnitude but was insensitive to gradient. The expression of c-jun was weakly correlated with magnitude and gradient, without an interaction effect. Monocyte chemoattractant protein-1 expression varied inversely with gradient and also depended on the interaction of gradient with magnitude. Intercellular adhesion molecule-1 expression also exhibited an interaction effect, and increased with shear magnitude. These results support the notion that vascular endothelial cells are able to sense shear gradient and magnitude independently.

Analysis of pulsatile flow in the parallel-plate flow chamber with spatial shear stress gradient

The Parallel-Plate Flow Chamber (PPFC), of which the height is far smaller than its own length and width, is one of the main apparatus for the in-vitro study of the mechanical behavior of cultured vascular Endothelical Cells (ECs) exposed to fluid shear stress. The steady flow in different kinds of PPFC has been extensively investigated, whereas, the pulsatile flow in the PPFC has received little attention. In consideration of the characteristics of geometrical size and pulsatile flow in the PPFC, the 3-D pulsatile flow was decomposed into a 2-D pulsatile flow in the vertical plane, and an incompressible plane potential flow in the horizontal plane. A simple method was then proposed to analyze the pulsatile flow in the PPFC with spatial shear stress gradient. On the basis of the method, the pulsatile fluid shear stresses in several reported PPFCs with spatial shear stress gradients were calculated. The results were theoretically meaningful for applying the PPFCs in-vitro, to simulate the pulsatile fluid shear stress environment, to which cultured ECs were exposed.

The Effect of Spatial Shear Stress Gradient on Endothelial Elongation and eNOS Expression

Objective As large spatial shear stress gradients are found in regions of the vasculature susceptible to atherosclerosis, this study examines the effect of such a gradient on flow-dependent endothelial cell behavior. Methods A flow chamber was designed that exposed porcine aortic endothelial cells to a nearly constant gradient over a physiologically relevant range of shear stress magnitude. The cells were analyzed with respect to elongation and protein expression level of endothelial nitric oxide synthase (eNOS). The latter, measured via quantitative immunofluorescence microscopy, was compared to eNOS expression by cells exposed to similar levels of shear stress in the absence of a gradient. Results Cell elongation increased with shear stress along the channel and was unaffected by gradient. Shear gradient suppressed the shear sensitivity of eNOS expression (p=0.002). Conclusion The protective role of shear stress, as manifested by increased eNOS expression, is diminished in the presence of a shear stress gradient, suggesting a role for this hemodynamic factor in atherosusceptibility. Supported by NIH grant H50442 and a Whitaker Foundation Graduate Research Fellowship.

Interaction of Wall Shear Stress Magnitude and Gradient in the Prediction of Arterial Macromolecular Permeability

Large spatial shear stress gradients have anecdotally been associated with early atherosclerotic lesion susceptibility in vivo and have been proposed as promoters of endothelial cell dysfunction in vitro. Here, experiments are presented in which several measures of the fluid dynamic shear stress, including its gradient, at the walls of in vivo porcine iliac arteries, are correlated against the transendothelial macromolecular permeability of the vessels. The fluid dynamic measurements are based on postmortem vascular casts, and permeability is measured from Evans blue dye (EBD) uptake. Time-averaged wall shear stress (WSS), as well as a new parameter termed maximum gradient stress (MGS) that describes the spatial shear stress gradient due to flow acceleration at a given point, are mapped for each artery and compared on a point-by-point basis to the corresponding EBD patterns. While there was no apparent relation between MGS and EBD uptake, a composite parameter, WSS(-0.11) MGS(0.044), was highly correlated with permeability. Notwithstanding the small exponents, the parameter varied widely within the region of interest. The results suggest that sites exposed to low wall shear stresses are more likely to exhibit elevated permeability, and that this increase is exacerbated in the presence of large spatial shear stress gradients.

Endothelial cell dynamics under pulsating flows: significance of high versus low shear stress slew rates (d(tau)/dt).

Shear stress modulates endothelial cell (EC) remodeling via realignment and elongation. We provide the first evidence that the upstroke slopes of pulsatile flow, defined as shear stress slew rates (positive d(tau)/dt), affect significantly the rates at which ECs remodel. We designed a novel flow system to isolate various shear stress slew rates by precisely controlling the frequency, amplitude, and time-averaged shear stress (tau(ave)) of pulsatile flow. Bovine aortic endothelial cell (BAEC) monolayers were exposed to three conditions: (1) pulsatile flow (1 Hz) at high slew rate (293 dyn/cm2 s), (2) pulsatile flow (1 Hz) at low slew rate (71 dyn/cm2s), and (3) steady laminar flow at d(tau)/dt = 0. All of the three conditions were operated at tau(ave) = 50 dyn/cm2. BAEC elongation and alignment were measured over 17 h. We were able to demonstrate the effects of shear stress slew rates ((tau)/dt) on EC remodeling at a fixed spatial shear stress gradient (d(tau)/dx). We found that pulsatile flow significantly increased the rates at which EC elongated and realigned, compared to steady flow at d(tau)/dt = 0. Furthermore, EC remodeling was faster in response to high than to low slew rates at a given tau(ave).

Vascular endothelial cells respond to spatial gradients in fluid shear stress by enhanced activation of transcription factors.

The vascular endothelium is exposed to a spectrum of fluid mechanical forces generated by blood flow; some of these, such as fluid shear stress, can directly modulate endothelial gene expression. Previous work by others and in our laboratory, using an in vitro uniform laminar shear stress model, has identified various shear stress response elements (SSREs) within the promoters of certain endothelial genes that regulate their expression by interacting with various transcription factors, including nuclear factor-kappaB (NF-kappaB), early growth response-1 (Egr-1), and activator protein-1 (AP-1, composed of c-Jun/c-Jun and c-Jun/c-Fos protein dimers). In the current study, we have examined the topographical patterns of NF-kappaB, Egr-1, c-Jun, and c-Fos activation in a specially designed in vitro disturbed laminar shear stress model, which incorporates regions of significant spatial shear stress gradients similar to those found in atherosclerosis-prone arterial geometries in vivo (eg, arterial bifurcations, curvatures, ostial openings). Using newly developed quantitative image analysis techniques, we demonstrate that endothelial cells subjected to disturbed laminar shear stress exhibit increased levels of nuclear localized NF-kappaB, Egr-1, c-Jun, and c-Fos, compared with cells exposed to uniform laminar shear stress or maintained under static conditions. In addition, individual cells display a heterogeneity in responsiveness to disturbed flow, as measured by the amount of NF-kappaB, Egr-1, c-Jun, and c-Fos in their nuclei. This differential regulation of transcription factor expression by disturbed versus uniform laminar shear stress indicates that regional differences in blood flow patterns in vivo-in particular, the occurrence of spatial shear stress gradients-may represent important local modulators of endothelial gene expression at anatomic sites predisposed for atherosclerotic development.