Gloe T, Riedmayr S, Sohn H Y, Pohl U
Institute of Physiology, Ludwig Maximilians University, Schillerstrasse 44, D 80336 Munich, Germany.
J Biol Chem. 1999 Jun 4;274(23):15996-6002. doi: 10.1074/jbc.274.23.15996.
It has been suggested that the mechanical forces acting on endothelial cells may be sensed in part by cell-matrix connections. We therefore studied the role of different matrix proteins, in particular laminin I, on a shear stress-dependent endothelial response, namely nitric-oxide synthase (eNOS) expression. Primary porcine aortic endothelial cells were seeded onto glass plates either noncoated (NC cells) or precoated with fibronectin (FN cells), laminin (LN cells), or collagen I (CN cells). Western blots were used to detect differences in the final matrix composition of these cells. A shear stress of 16 dyn/cm2 was applied for 6 h. Only LN cells showed detectable amounts of laminin I in their underlying matrix when they reached confluence. They reacted with a 2-fold increase of eNOS expression (n = 16, p < 0.001) to the exposure of shear stress, which went along with enhanced eNOS protein and NO release. In contrast, neither FN cells (n = 9) nor NC cells (n = 13) showed a significant increase of eNOS expression under shear stress. The increase in CN cells was borderline (1.4-fold; n = 9, p < 0.05) and was not associated with an increase of eNOS protein. The shear-induced increase in eNOS expression of LN cells was abolished by the peptide YIGSR, which blocks the cellular binding to laminin I via a 67-kDa laminin-binding protein, whereas a control peptide (YIGSK) had no effect. The induction of eNOS expression by shear stress is stimulated by an interaction of endothelial cells with laminin which is, at least in part, mediated by a 67-kDa laminin-binding protein.
有人提出，作用于内皮细胞的机械力可能部分通过细胞与基质的连接来感知。因此，我们研究了不同基质蛋白，特别是层粘连蛋白I，对剪切应力依赖性内皮反应，即一氧化氮合酶（eNOS）表达的作用。将原代猪主动脉内皮细胞接种到未包被的玻璃板（NC细胞）或预先包被有纤连蛋白（FN细胞）、层粘连蛋白（LN细胞）或I型胶原（CN细胞）的玻璃板上。采用蛋白质免疫印迹法检测这些细胞最终基质组成的差异。施加16 dyn/cm2的剪切应力6小时。只有LN细胞在汇合时其下层基质中显示出可检测到的层粘连蛋白I量。它们在受到剪切应力作用后，eNOS表达增加了2倍（n = 16，p < 0.001），同时eNOS蛋白和NO释放增强。相比之下，FN细胞（n = 9）和NC细胞（n = 13）在剪切应力下均未显示eNOS表达有显著增加。CN细胞的增加处于临界水平（1.4倍；n = 9，p < 0.05），且与eNOS蛋白增加无关。肽YIGSR可消除剪切诱导的LN细胞eNOS表达增加，该肽通过一种67 kDa的层粘连蛋白结合蛋白阻断细胞与层粘连蛋白I的结合，而对照肽（YIGSK）则无作用。剪切应力诱导的eNOS表达受内皮细胞与层粘连蛋白相互作用的刺激，这种相互作用至少部分由一种67 kDa的层粘连蛋白结合蛋白介导。
