The CellBorderTracker, a novel tool to quantitatively analyze spatiotemporal endothelial junction dynamics at the subcellular level
The CellBorderTracker, a novel tool to quantitatively analyze spatiotemporal endothelial junction dynamics at the subcellular level
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CellBorderTracker，一种在亚细胞水平定量分析时空内皮连接动态的新工具
DOI：
10.1007/s00418-015-1357-8
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发表时间：
2015
影响因子：
2.3
通讯作者：
Schnittler
中科院分区：
文献类型：
--
作者：
Seebach;Lindemann;Brinkmann;Bogdan;Schnittler
内皮连接是由多蛋白复合物组织的动态结构，其控制单层完整性、稳态、炎症、细胞迁移和血管生成。新开发的内皮细胞遗传操作和显微镜检查方法允许长时间的荧光蛋白延时记录。定量数据分析需要自动化方法。我们开发了一个软件包，CellBorderTracker，允许定量分析荧光标记的细胞连接蛋白动态的时间推移序列。CellBorderTracker由CellBorderExtractor和映射工具组成，CellBorderExtractor分割单元格并识别单元格边界，用于数据提取。该工具是通过分析荧光标记的VE-钙粘蛋白的骨干内皮细胞粘附连接。VE-钙粘蛋白显示高动力学，其由连接相关的间歇性板状伪足（JAIL）强制，所述连接相关的间歇性板状伪足是肌动蛋白驱动的并且是WASP/ARP 2/3复合物控制的。手动分割和自动分割的一致性达到90%，这一值表明可靠性高。基于分割，生成了不同的地图，从而可以提取更详细的数据。这包括蛋白质分布模式的定量、感兴趣区域的生成、连接位移、细胞形状变化、迁移速度和连接动态的可视化。此外，我们展示了一个先进的记波仪，J-记波仪，稳定地遵循不规则的细胞连接动态的延时序列在亚细胞水平的个别路口。通过使用CellBorderTracker，我们证明了VE-钙粘蛋白动力学在凝血酶刺激后迅速被捕，这种现象主要是由于JAIL的瞬时抑制，并在细胞间间隙形成和重新密封期间显示非常异质的亚细胞和潜水员VE-钙粘蛋白动力学。
Endothelial junctions are dynamic structures organized by multi-protein complexes that control monolayer integrity, homeostasis, inflammation, cell migration and angiogenesis. Newly developed methods for both the genetic manipulation of endothelium and microscopy permit time-lapse recordings of fluorescent proteins over long periods of time. Quantitative data analyses require automated methods. We developed a software package, the CellBorderTracker, allowing quantitative analysis of fluorescent-tagged cell junction protein dynamics in time-lapse sequences. The CellBorderTracker consists of the CellBorderExtractor that segments cells and identifies cell boundaries and mapping tools for data extraction. The tool is illustrated by analyzing fluorescent-tagged VE-cadherin the backbone of adherence junctions in endothelium. VE-cadherin displays high dynamics that is forced by junction-associated intermittent lamellipodia (JAIL) that are actin driven and WASP/ARP2/3 complex controlled. The manual segmentation and the automatic one agree to 90 %, a value that indicates high reliability. Based on segmentations, different maps were generated allowing more detailed data extraction. This includes the quantification of protein distribution pattern, the generation of regions of interest, junction displacements, cell shape changes, migration velocities and the visualization of junction dynamics over many hours. Furthermore, we demonstrate an advanced kymograph, the J-kymograph that steadily follows irregular cell junction dynamics in time-lapse sequences for individual junctions at the subcellular level. By using the CellBorderTracker, we demonstrate that VE-cadherin dynamics is quickly arrested upon thrombin stimulation, a phenomenon that was largely due to transient inhibition of JAIL and display a very heterogeneous subcellular and divers VE-cadherin dynamics during intercellular gap formation and resealing.
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影响因子：
7.5
作者：
Harris ES;Nelson WJ
通讯作者：
Nelson WJ
影响因子：
6.7
作者：
J. Seebach;H. Mädler;B. Wojciak-Stothard;H. Schnittler
通讯作者：
J. Seebach;H. Mädler;B. Wojciak-Stothard;H. Schnittler
影响因子：
21.3
作者：
Kametani, Yoshiko;Takeichi, Masatoshi
通讯作者：
Takeichi, Masatoshi
两种 F-BAR 蛋白 Cip4 和 Nostrin 在上皮形态发生中 E-钙粘蛋白调节中的协同功能
影响因子：
4
作者：
Zobel T;Brinkmann K;Koch N;Schneider K;Seemann E;Fleige A;Qualmann B;Kessels MM;Bogdan S
通讯作者：
Bogdan S
影响因子：
7.8
作者：
LAMPUGNANI, MG;CORADA, M;CAVEDA, L;BREVIARIO, F;AYALON, O;GEIGER, B;DEJANA, E
通讯作者：
DEJANA, E