The arterial system is subjected to cyclic strain because of periodic alterations in blood pressure, but the effects of frequency of cyclic strain on arterial smooth muscle cells (SMCs) remain unclear. Here, we investigated the potential role of the cyclic strain frequency in regulating SMC alignment using an in vitro model. Aortic SMCs were subject to cyclic strain at one elongation but at various frequencies using a Flexercell Tension Plus system. It was found that the angle information entropy, the activation of integrin-beta1, p38 MAPK, and F/G actin ratio of filaments were all changed in a frequency-dependent manner, which was consistent with SMC alignment under cyclic strain with various frequencies. A treatment with anti-integrin-beta1 antibody, SB202190, or cytochalasin D inhibited the cyclic strain frequency-dependent SMC alignment. These observations suggested that the frequency of cyclic strain plays a role in regulating the alignment of vascular SMCs in an intact actin filament-dependent manner, and cyclic strain at 1.25 Hz was the most effective frequency influencing SMC alignment. Furthermore, integrin-beta1 and p38 MAPK possibly mediated cyclic strain frequency-dependent SMC alignment.

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This study evaluated the possibility of differentiation from embryonic stem (ES) cells to vascular wall cells by physical (mechanical) stress loading in vitro. A cell mixture containing Flk1-positive cells (ca. 30%) derived from murine ES cells was added to a compliant microporous tube made of segmented polyurethane. The compliance of the tube was close to that of the human artery [the stiffness parameter (beta) = 57.2 (n = 5, SD < 5%)]. The luminal surface of the tube was fully covered with the cells by preincubation for two days in the presence of vascular endothelial growth factor (VEGF). After 2 days of additional incubation without VEGF under static conditions, layering of the grown cells, mostly smooth muscle actin (SMA)-positive cells, was observed only on the luminal surface of the tube. The cells were flat, polygonal, and randomly oriented. On the other hand, after a 2-day incubation under a weak pulsatile flow simulating the human venous systems [wall shear stress (WSS) from -0.98 to 2.2 dyn/cm(2); circumferential strain (CS) 4.6-9.6 x 10(4) dyn/cm(2)] without VEGF, cells in the superficial layer were regularly oriented in the direction of the pulsatile flow. The oriented cells exhibited endothelial-like appearance, indicating that they were platelet endothelial cell adhesion molecule 1 (PECAM1)-positive. In addition, the cells growing into the interstices in the deeper layer showed smooth muscle-like appearance, indicating that they were SMA-positive. Differentiation to two different cell types and segregation of incorporated ES cells may be simultaneously encouraged by the combination of WSS and CS. It is expected that the monobloc building of hierarchically structured hybrid vascular prostheses composed of several vascular wall cell types is possible by physically synchronized differentiation of ES cells.