Effects of derection and pulsatility of flow on morphology and functions of cultured endothelial cells

研究了血流方向对体外培养的牛主动脉内皮细胞形态和细胞骨架结构的影响。在玻璃上培养的完全融合的内皮细胞，使用低截面0.5 mm高，宽60mm的平行板流室，承受20达因/cm^2的流体施加剪切应力24小时。剪切流暴露实验进行了(i)单向流动和（ii）往复流动，间隔30分钟。在往复流动实验中，通过滚轮泵和带定时器控制的夹管阀交替改变介质的反向流动。血流暴露后，固定内皮细胞，用罗丹明phalloidin染色Ff-actin丝。用带有会聚荧光光学装置的显微镜观察和拍摄内皮细胞，并在打印的显微照片上测量每个细胞的形状指数。观察到在两种流动条件下，细胞与流动方向和应力纤维排列一致。单向流条件下的细胞排列度比往复流条件下的细胞排列度高。往复流动条件下内皮细胞的形状指数值（0.68(]SY.+-.[)0.12）介于无流动条件下的形状指数值（0.87(]SY.+-.[)0.06）和单向流动条件下的形状指数值（0.48(]SY.+-.[)0.15）之间。在静态培养的细胞中，f -肌动蛋白丝的密集外周带呈蜂窝状。单向流动条件下，应力纤维主要形成于细胞的中心区域，并沿流动方向排列。每个细胞上游的f -肌动蛋白丝被强烈染色。相反，暴露于往复流动的细胞中的f -肌动蛋白丝几乎均匀染色。众所周知，内皮细胞对剪切应力有反应。综上所述，我们的实验结果表明内皮细胞

Effects of flow direction on shape and cytoskeletal structure of cultured bovine aortic endothelial cells were studied. Fully confluent endothelial cells cultured on glass were subjected to fluid-imposed shear stress of 20 dyne/cm^2 for 24 hours using a parallel plate flow chamber which has a low section 0.5 mm in height and 60 mm in width. Experiments on shear flow exposure were performed for (i) one-way flow, and (ii) reciprocating flow with a 30 minute interval. In the reciprocating flow experiment, medium flow was alternatively changed in the oposite direction by a roller pump and pinch valves controlled with a timer. After the flow exposure endothelial cells were fixed and Ff-actin filaments were stained with rhodamine phalloidin. Endothelial cells were observed and photographed by means of a microscope equipped with epiflluorescence optics, and the shape index of each individual cell was measured on the printed photomicrographs. Cells aligned with the flow direction and stress fibers were observed to be developed under the both flow conditions. The degrees of cell alignment under the one-way flow conditon became higher than those under reciprocating flow. Shape index value of endothelial cells exposed to the reciprocating flow (0.68(〕SY.+-.〔)0.12) was between that of the no-flow (0.87(〕SY.+-.〔)0.06)and the one-way flow conditions (0.48(〕SY.+-.〔)0.15). Dense peripheral bands of F-actin filaments were characteristically formed in honeycomb shape in the statically cultured cells. Under the one-way flow conditions, stress fibers were mainly formed in the central area of the cells and aligned with the flow direction. And the F-actin filaments around the upstream side of each cell were observed to be strongly stained. In contrast, F-actin filaments in the cells exposed to reciprocating flow almost stained homogeneously. Endothelial cells are well known to respond to shear stress. Furthermore, in conclusion, our experimental results show that endothelial cell