Effect of L-homocysteine on endothelial cell-cell junctions following F-actin stabilization through tropomyosin-1 overexpression
Effect of L-homocysteine on endothelial cell-cell junctions following F-actin stabilization through tropomyosin-1 overexpression
复制标题
DOI：
10.3892/ijmm.2013.1357
复制
发表时间：
2013-07-01
影响因子：
5.4
通讯作者：
Grzanka, Alina
中科院分区：
文献类型：
--
作者：
Gagat, Maciej;Grzanka, Dariusz;Grzanka, Alina
自从肌动蛋白在非肌肉细胞中被发现以来，人们认为肌动蛋白细胞骨架的力学行为的调节调节了细胞的形状变化和细胞迁移和分裂过程中力的产生。细胞形状和极性的维持在细胞-细胞连接的形成中很重要。本研究旨在探讨L-同型半胱氨酸硫内酯盐酸盐通过稳定微丝肌动蛋白细胞骨架(F-Actin)维持细胞间连接对EA.hy926血管内皮细胞的影响。原肌球蛋白-1的过表达能够稳定肌肉和非肌肉细胞中的肌动蛋白细丝，从而稳定肌动蛋白细丝。在L-同型半胱氨酸作用下，F-肌动蛋白的稳定可显著降低晚期细胞凋亡率和坏死率。此外，高表达原肌球蛋白-1的细胞经L-同型半胱氨酸处理后，内皮细胞的迁移能力更强。此外，我们的结果表明，与不过表达原肌球蛋白-1的细胞相比，F-肌动蛋白在EA.hy926细胞中的稳定显著增加了连接β-连环蛋白的表达。同样，在F-肌动蛋白细胞骨架稳定的细胞中，连接α-连环蛋白的荧光强度也增加。然而，这一增加仅略高于未过表达原肌球蛋白-1的EA.hy926细胞。此外，闭锁带(ZO)-1的相对荧光分析表明，与不过表达原肌球蛋白-1的细胞相比，具有稳定的F-肌动蛋白细胞骨架的细胞-细胞连接面积显著减少。我们的结果表明，F-肌动蛋白的稳定不影响细胞的迁移能力，从而保护EA.hy926细胞免受L同型半胱氨酸诱导的细胞迁移率的下降。此外，α-连环蛋白可能参与了细胞-细胞连接区肌动蛋白聚合的抑制。可以假设，由于β-连环蛋白和ZO-1荧光信号的放大，F-肌动蛋白的稳定通过增加细胞-细胞连接的数量来加强内皮细胞的黏附和紧密连接。然而，ZO-1通过稳定F-肌动蛋白来稳定内皮屏障功能，而F-肌动蛋白本身又稳定了ZO-1的定位。
Since the identification of actin in non-muscle cells, it has been suggested that the regulation of the mechanical behaviors of the actin cytoskeleton regulates cellular shape changes and the generation of forces during cell migration and division. The maintenance of cell shape and polarity are important in the formation of cell-cell junctions. The aim of the present study was to determine the effect of L-homocysteine thiolactone hydrochloride on EA.hy926 endothelial cells in the context of the maintenance cell-cell junctions through the stabilization of filamentous actin cytoskeleton (F-actin). The actin filaments were stabilized by the overexpression of tropomyosin-1, which has the ability to stabilize actin filaments in muscle and nonmuscle cells. The stabilization of F-actin induced a significant decrease in the percentage of late apoptotic and necrotic cells following treatment with L-homocysteine. Moreover, the migratory potential of the endothelial cells was greater in the cells overexpressing tropomyosin-1 treated with L-homocysteine. Additionally, our results indicated that the stabilization of F-actin in the EA.hy926 cells significantly increased the expression of junctional beta-catenin, as compared to the cells not overexpressing tropomyosin-1. Similarly, the fluorescence intensity of junctional alpha-catenin was also increased in the cells with stabilized F-actin cytoskeleton. However, this increase was only slightly higher than that observed in the EA.hy926 cells not overexpressing tropomyosin-1. Furthermore, the analysis of Zonula occludens (ZO)-1 relative fluorescence demonstrated a statistically significant decrease in the cell-cell junction areas among the cells with stabilized F-actin cytoskeleton in comparison to the cells not overexpressing tropomyosin-1. Our results indicate that the stabilization of F-actin does not affect the migratory potential of cells, and consequently protects the EA.hy926 cells against the L-homocysteine-induced decrease in cell mobility. Moreover, it is suggested that alpha-catenin may participate id the suppression of actin polymerization in the area of cell-cell junctions. It can be hypothesized that the stabilization of F-actin strengthens endothelial adherens and tight junctions by increasing the number of cell-cell junctions due to the amplification of (beta-catenin and the ZO-1 fluorescence signal. However, ZO-1 stabilizes the endothelial barrier function through the stabilization of F-actin and F-actin itself stabilizes the localization of ZO-1.