Cytoskeletal response to

A pulsatile flow through a healthy carotid bifurcation is simulated. Several particles (in green) are lingering in the recirculation zone in the carotid bulb as shown on the picture on the left. In the picture, the particles are released from the common carotid, and they are travelling into either the internal (left) or external (right) carotid.

Adopted from http://itsa.ucsf.edu/~ldjou/carotid.html

The pulsatile nature of blood flow generates a complex interplay of mechanical forces against the inner wall of the vasculature. All three types of hemodynamic forces, namely shear stress, cyclic strain and hydrostatic pressure, greatly influence the physiology of the vessel wall. Among these, fluid shear stress is the predominant tangential force acting on the vessel wall. Shear stress has been implicated in mediating not only normal vascular tone and remodeling, but more importantly in the pathophysiology of atherosclerosis, reperfusion, and wound healing. In fact, there is a direct correlation between aberrant shear stress and atherogenesis. The earliest atherosclerotic lesions typically develop in area of high flow disturbances, such as non-laminar flow, flow reversal and roaming stagnation points.

The endothelial cells (ECs) therefore forms a physiologically dynamic interface with the bloodstream, acting as an integrator and transducer of both biomechanical and humoral signals. Fluid shear stress regulates endothelial phenotype not only by changing its gene expression profile, but on a long term basis by altering the EC morphology. Vascular regions with turbulent flow and low shear stress (high risk of atherogenesis) contain ECs that are polygonal, resembling those grown in static culture. In contrast, ECs in regions of unobstructed flow (relatively resistant to atherosclerosis) elongate in the direction of flow. These cell shape changes represent critical adaptive processes with which ECs cope with local mechanical load and subsequent injury.

Cell shape is directly mediated in large part by the actomyosin contractile system. A distinct response to increased shear stress is the transformation of the EC actin cytoskeleton, resulting in the alignment of F-actin bundles in the direction of flow, attenuation of cortical F-actin and enhanced central stress fibers. Although this phenomenon is thought to be partly the result of increased myosin-mediated tension, it can be readily distinguished from the myosin-based contractility in response to inflammatory stimuli that results in EC contraction, and not directional elongation. This distinction thus suggests a more controlled and elaborate cytoskeletal remodeling. Actin organization is mediated by a plethora of actin-binding proteins under the control of various upstream signals ranging from phospholipids, Ca²⁺, to the Rho family GTPases, all of which have been shown to respond directly with altered shear stress. Although the reorganization of the actin cytoskeleton is well documented, mechanistically we do not fully understand how the cortical actin bands are replaced by thick and flow-aligned stress fibers.

The shear stress-induced EC elongation inevitably necessitates the redistribution of cell-cell contacts to accommodate the changes in interaction between neighboring cells. The EC-specific adherens junction is composed of vascular endothelial (VE)-cadherin, which is linked to the actin cytoskeleton by catenin proteins. The flow-induced actin rearrangement thus dictates an accompanying change in the adherens junctions. It is unclear if adherens junctions are disassembled prior to its remodeling to accommodate the shear stress-related cell shape changes. This poorly understood event carries significant clinical implications. The adherens junction directly influences the endothelial permeability, and vascular regions exposed to extreme or fluctuating shear stresses display aberrantly high permeability. The disruption of cell-cell junction has been postulated to contribute to the pathogenesis of atherosclerosis, probably due to elevated leukocyte diapedesis and molecular extravasation as a direct result of shear-sensitive breach of the endothelial permeability barrier.

To fulfill the function of transducing biomechanical stimuli into biochemical responses during hemodynamic regulation, ECS must be equipped with mechanisms for flow sensing. We are interested therefore in how the cytoskeletal network and the cell adhesion systems respond to shear stress either in smooth laminar or turbulent flows. Numerous probes such as fluorescently tagged myosin, paxillin, vinculin, a-catenin, MLCK-FIP, tubulin and actin are used to decipher the dynamic interaction among these structural network under fluid shear stress. Listed below are snapshots of results obtained thus far.

	Focal contact elongates and increases in size in response to elevated laminar flow

	Reorganization of myosin II network as monitored by GFP-RLC in bovine pulmonary artery endothelial cells. Compare the stability of stress fibers parallel to the direction of flow and the rapid remodeling of those perpendicular to the fluid shear stress

	GFP-tagged a-catenin demonstrates how the integrity of cell-cell contact can be compromised by excessive shear stress