Rho-associated kinase-dependent contraction of stress fibres and the organization of focal adhesions.

Stress fibres and associated focal adhesions in cells constitute a contractile apparatus that regulates cell motility and contraction. Rho-kinase, an effector molecule of small GTPases, regulates non-muscle cell motility and contractility. Rho-kinase mediates the contraction of stress fibres in a Ca(2+)-independent manner, and is responsible for slower and more finely tuned contraction of stress fibres than that regulated by myosin light chain kinase activity in living cells. The specific inhibition of the Rho-kinase activity causes cells to not only lose their stress fibres and focal adhesions, but also to appear to lose their cytoplasmic tension. Activated Rho-kinase is also involved in the organization of newly formed stress fibres and focal adhesions in living cells.

Role of stress fibers and focal adhesions as a mediator for mechano-signal transduction in endothelial cells in situ.

Fluid shear stress is the mechanical force generated by the blood flow which is applied over the apical surface of endothelial cells in situ. The findings of a recent study suggest that stress fibers and its associated focal adhesions play roles in mechano-signal transduction mechanism. Stress fibers are present along the apical and the basal portion of the endothelial cells. Endothelial cells respond to fluid shear stress and change their morphological characteristics in both their cell shape and cytoskeletal organization. Atherosclerosis is a common disease of the arteries and it occurs in areas around the branching site of blood vessels where the cells are exposed to low fluid shear stress. The organization of stress fibers and focal adhesions are strongly influenced by shear stress, and therefore the generation of atherosclerotic lesions seem to be associated with the cytoskeletal components of endothelial cells. This review describes the possible role of the cytoskeleton as a mechano-transducer in endothelial cells in situ.

Rho-kinase dependent organization of stress fibers and focal adhesions in cultured fibroblasts.

The activation of Rho-kinase is known to modulate the organization of the actin-based cytoskeletal systems, including the formation of stress fibers and focal adhesions. Rho-kinase likely plays a more crucial and complex role in the organization of actin-based cytoskeletal systems than in that of myosin light chain kinase (MLCK). In order to understand the role of Rho-kinase in the organization of stress fibers and focal adhesions, we treated cultured fibroblasts with a Rho-kinase inhibitor and analyzed the stress fiber and focal adhesion organization under conventional fluorescence microscopy and replica electron microscopy. Some of the cells were transfected with GFP-labeled paxillin, actin or alpha-actinin, and the effects of the inhibitor were monitored in the living cells. The Rho-kinase inhibitor caused disassembly of the stress fibers and focal adhesions in the central portion of the cell within 1 h. However, the stress fibers and focal adhesions located in the cell periphery were not as severely affected by the Rho-kinase inhibitor. The time-lapse video recording revealed that when these cells were washed with a fresh medium in order to remove the Rho-kinase inhibitor, the stress fibers and focal adhesions located in the center of the cell gradually reorganized and, within 1.5-2 h, the cells completely recovered. This observation strongly suggests that the activation of Rho-kinase plays an important role in the organization of the central stress fibers and focal adhesions.

Morphological differences between guinea pig aortic and venous endothelial cells in situ.

Endothelial cells (ECs) respond to fluid shear stress. They reveal shear stress related morphological changes in both their cell shape and cytoskeletal organization. Little is known about the cytoskeletal organization of ECs in situ. We studied, together with the living ultrasound high resolution imaging system, the distribution of stress fibers (SFs), certain focal adhesion (FA) and signal transduction associated proteins in guinea pig aortic and venous ECs. Although SFs present in the basal portion of venous ECs ran along the direction of the blood flow, their size was smaller and their number was fewer than those of aortic ECs. Venous ECs were elongated to the direction of flow than in aortic ECs exposed over normal shear stress (SS). Since fluid SS in the vein is low, a sustained and uni-directional low SS over a long period might thus cause these structural features observed in venous ECs.