Rizzo Victor, Morton Christine, DePaola Natacha, Schnitzer Jan E, Davies Peter F
Center for Cardiovascular Science, Albany Medical College, 47 New Scotland Ave., Albany, NY 12208, USA.
Am J Physiol Heart Circ Physiol. 2003 Oct;285(4):H1720-9. doi: 10.1152/ajpheart.00344.2002. Epub 2003 Jun 19.
The luminal surface of rat lung microvascular endothelial cells in situ is sensitive to changing hemodynamic parameters. Acute mechanosignaling events initiated in response to flow changes in perfused lung microvessels are localized within specialized invaginated microdomains called caveolae. Here we report that chronic exposure to shear stress alters caveolin expression and distribution, increases caveolae density, and leads to enhanced mechanosensitivity to subsequent changes in hemodynamic forces within cultured endothelial cells. Flow-preconditioned cells expressed a fivefold increase in caveolin (and other caveolar-residing proteins) at the luminal surface compared with no-flow controls. The density of morphologically identifiable caveolae was enhanced sixfold at the luminal cell surface of flow-conditioned cells. Laminar shear stress applied to static endothelial cultures (flow step of 5 dyn/cm2), enhanced the tyrosine phosphorylation of luminal surface proteins by 1.7-fold, including caveolin-1 by 1.3-fold, increased Ser1179 phosphorylation of endothelial nitric oxide synthase (eNOS) by 2.6-fold, and induced a 1.4-fold activation of mitogen-activated protein kinases (ERK1/2) over no-flow controls. The same shear step applied to endothelial cells preconditioned under 10 dyn/cm2 of laminar shear stress for 6 h and induced a sevenfold increase of total phosphotyrosine signal at the luminal endothelial cell surface enhanced caveolin-1 tyrosine phosphorylation 5.8-fold and eNOS phosphorylation by 3.3-fold over static control values. In addition, phosphorylated caveolin-1 and eNOS proteins were preferentially localized to caveolar microdomains. In contrast, ERK1/2 activation was not detected in conditioned cells after acute shear challenge. These data suggest that cultured endothelial cells respond to a sustained flow environment by directing caveolae to the cell surface where they serve to mediate, at least in part, mechanotransduction responses.
大鼠肺微血管内皮细胞原位的管腔表面对血流动力学参数的变化敏感。因灌注肺微血管内血流变化引发的急性机械信号转导事件定位于称为小窝的特殊内陷微区。在此，我们报告长期暴露于剪切应力会改变小窝蛋白的表达和分布，增加小窝密度，并导致培养的内皮细胞对随后血流动力学力变化的机械敏感性增强。与无血流对照相比，经血流预处理的细胞在管腔表面的小窝蛋白（以及其他定位于小窝的蛋白）表达增加了五倍。在经血流预处理的细胞的管腔细胞表面，形态上可识别的小窝密度增加了六倍。施加于静态内皮细胞培养物的层流剪切应力（5达因/平方厘米的血流阶跃）使管腔表面蛋白的酪氨酸磷酸化增加了1.7倍，包括小窝蛋白-1增加了1.3倍，使内皮型一氧化氮合酶（eNOS）的Ser1179磷酸化增加了2.6倍，并诱导丝裂原活化蛋白激酶（ERK1/2）的激活比无血流对照增加了1.4倍。将相同的剪切阶跃施加于在10达因/平方厘米的层流剪切应力下预处理6小时的内皮细胞，与静态对照值相比，诱导管腔内皮细胞表面的总磷酸酪氨酸信号增加了七倍，小窝蛋白-1酪氨酸磷酸化增强了5.8倍，eNOS磷酸化增强了3.3倍。此外，磷酸化的小窝蛋白-1和eNOS蛋白优先定位于小窝微区。相反，在急性剪切刺激后，在预处理细胞中未检测到ERK1/2激活。这些数据表明，培养的内皮细胞通过将小窝引导至细胞表面来响应持续的血流环境，在那里小窝至少部分地介导机械转导反应。
