From the Departments of Medicine (O.N.S., R.M.B., C.Y., B.C.B.) and Pharmacology and Physiology (O.N.S., C.Y., B.C.B.), University of Rochester School of Medicine and Dentistry, Aab Cardiovascular Research Institute, NY.
Circ Res. 2014 Mar 28;114(7):1125-32. doi: 10.1161/CIRCRESAHA.114.301315. Epub 2014 Feb 10.
Fluid shear stress differentially regulates endothelial cell stress fiber formation with decreased stress fibers in areas of disturbed flow compared with steady flow areas. Importantly, stress fibers are critical for several endothelial cell functions including cell shape, mechano-signal transduction, and endothelial cell-cell junction integrity. A key mediator of steady flow-induced stress fiber formation is Src that regulates downstream signaling mediators such as phosphorylation of cortactin, activity of focal adhesion kinase, and small GTPases.
Previously, we showed that thioredoxin-interacting protein (TXNIP, also VDUP1 [vitamin D upregulated protein 1] and TBP-2 [thioredoxin binding protein 2]) was regulated by fluid shear stress; TXNIP expression was increased in disturbed flow compared with steady flow areas. Although TXNIP was originally characterized for its role in redox and metabolic cellular functions, recent reports show important scaffold functions related to its α-arrestin structure. Based on these findings, we hypothesized that TXNIP acts as a biomechanical sensor that regulates Src kinase activity and stress fiber formation.
Using en face immunohistochemistry of the aorta and cultured endothelial cells, we show inverse relationship between TXNIP expression and Src activity. Specifically, steady flow increased Src activity and stress fiber formation, whereas it decreased TXNIP expression. In contrast, disturbed flow had opposite effects. We studied the role of TXNIP in regulating Src homology phosphatase-2 plasma membrane localization and vascular endothelial cadherin binding because Src homology phosphatase-2 indirectly regulates dephosphorylation of Src tyrosine 527 that inhibits Src activity. Using immunohistochemistry and immunoprecipitation, we found that TXNIP prevented Src homology phosphatase-2-vascular endothelial cadherin interaction.
In summary, these data characterize a fluid shear stress-mediated mechanism for stress fiber formation that involves a TXNIP-dependent vascular endothelial cadherin-Src homology phosphatase-2-Src pathway.
与稳定流区域相比，流动切应力可使内皮细胞的应力纤维形成减少，从而使紊乱流区域的应力纤维减少。重要的是，应力纤维对于内皮细胞的几种功能至关重要，包括细胞形状、力学信号转导和内皮细胞-细胞连接完整性。稳定流诱导的应力纤维形成的关键介质是Src，它调节下游信号转导介质，如 cortactin 的磷酸化、粘着斑激酶的活性和小 GTP 酶。
先前，我们表明硫氧还蛋白相互作用蛋白（TXNIP，也称为 VDUP1[维生素 D 上调蛋白 1]和 TBP-2[硫氧还蛋白结合蛋白 2]）受流切应力调节；与稳定流区域相比，TXNIP 在紊乱流中表达增加。尽管 TXNIP 最初因其在氧化还原和代谢细胞功能中的作用而被描述，但最近的报道显示了与其 α-抑制蛋白结构相关的重要支架功能。基于这些发现，我们假设 TXNIP 作为一种生物力学传感器，调节 Src 激酶活性和应力纤维形成。
我们通过主动脉和培养的内皮细胞的正面免疫组织化学，显示 TXNIP 表达与 Src 活性之间呈反比关系。具体而言，稳定流增加了 Src 活性和应力纤维形成，而降低了 TXNIP 的表达。相反，紊乱流则产生相反的效果。我们研究了 TXNIP 在调节 Src 同源磷酸酶-2 质膜定位和血管内皮钙黏蛋白结合中的作用，因为 Src 同源磷酸酶-2 间接调节抑制 Src 活性的 Src 酪氨酸 527 的去磷酸化。通过免疫组织化学和免疫沉淀，我们发现 TXNIP 阻止了 Src 同源磷酸酶-2-血管内皮钙黏蛋白的相互作用。
总之，这些数据描述了一种由流切应力介导的应力纤维形成机制，涉及 TXNIP 依赖性血管内皮钙黏蛋白-Src 同源磷酸酶-2-Src 途径。
