Mechanisms of morphological and functional changes of endothelial cells in atherogesesis.

1.为了揭示恒定层流切应力抗动脉粥样硬化的机制，我们分析了人凋亡抑制蛋白-2(HIAP-2)在血管内皮细胞中的表达。在切应力作用下，HIAP-2的表达呈剂量和时间依赖性。体内也检测到HIAP-2的表达。剪切力抑制内皮细胞凋亡与抑制caspase-3活性有关。Smac是一种caspase激活剂，通过与HIAP结合并去除其抑制活性，可减弱切应力对caspase-3活性的抑制作用。这些数据表明，剪切力通过增加HIAP-2对caspase-3的负调控来阻止EC的凋亡。2.层流剪切力可完全阻断内皮细胞分泌MMP2，但细胞内MMP2蛋白水平及MMP2mRNA表达水平无明显变化。为了阐明这一机制，我们利用GFP标记的融合基质金属蛋白酶-2分析了基质金属蛋白酶-2在细胞内的转运。B…层流切应力越大，基质金属蛋白酶-2/绿色荧光蛋白颗粒体积增大，进入细胞膜而无分泌物。我们的数据表明，层流切应力通过调节IV型胶原代谢促进了EC在基质上的黏附。3.为了在活体内分析动脉粥样硬化的初始事件，我们建立了一种改进的EN Face方法，使我们能够获得包括动脉分叉口在内的整个内皮表面的清晰图像，动脉粥样硬化就是在这里开始的。在SD大鼠的主动脉分支区，大量增殖的内皮细胞和巨噬细胞黏附在内皮细胞上，经特异性抗体染色，分别是非分支区的2倍和3倍。综上所述，通过调节细胞死亡、细胞生长、内皮细胞与基质的黏附来控制胶原代谢，以及通过层流切应力抑制单核细胞与内皮细胞的黏附来稳定内皮细胞，可能有助于血管内抗动脉粥样硬化微环境的形成。较少

1.To disclose the anti-atherosclerotic mechanisms of steady laminar shear stress, we analyzed the expression of human inhibitor of apoptosis protein-2(HIAP-2) in endothelial cells(ECs). HIAP-2 was dose-dependently and time-dependently induced in ECs by shear stress. HIAP-2 expression also identified in vivo. Shear stress-mediated inhibition of EC apoptosis was associated with inhibition of caspase-3 activity. Transfection of Smac, a caspase activator by binding HIAPs and removing their inhibitory activity, reduced the suppression effect of shear stress on caspase-3 activity. These data suggest that shear stress prevents EC apoptosis via negative regulation of caspase-3 by increments of HIAP-2. 2.The MMP2 secretion from ECs into medium was interrupted completely by laminar shear stress, although level of this intracellular protein and mRNA expression of MMP2 were unchanged. To clarify this mechanism, we analyzed intracellular transportation of MMP-2, using fused MMP-2 tagged with GFP. B … More y laminar shear stress, MMP-2/GFP granules increased their size and transported into cell membrane without secretion. Our data suggest that laminar shear stress contribute to EC adhesion on matrix through the regulation of collagen type IV metabolism. 3.To analyze a initial events of atherosclerosis, in vivo, we established a modified en face method that enabled us to obtain clear images of the entire surface of endothelium including a bifurcation orifices of aorta, where atherosclerosis initiates. A number of proliferating ECs and macrophages adhered on ECs, stained with specific antibodies, at the branching area of aorta from SD rats were about 2 and 3 times higher than that of non-branching area, respectively. In conclusion, stabilization of ECs through the regulation of cell death, cell growth, EC adhesion on matrix with control of collagen metabolism and inhibition of monocyte adhesion on ECs by the laminar shear stress may contribute to the formation of the anti-atherosclerotic microenvironment in vessels. Less