Advanced Methods for the Investigation of Cell Contact Dynamics in Endothelial Cells Using Florescence-Based Live Cell Imaging
| العنوان: | Advanced Methods for the Investigation of Cell Contact Dynamics in Endothelial Cells Using Florescence-Based Live Cell Imaging |
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| المؤلفون: | Mohammed Aldirawi, Quentin Leidl, Christine Schimp, Jiahui Cao, Irene Hofer, Jochen Seebach, Muna Taha, Annelie Ahle, Hans J. Schnittler |
| المصدر: | Journal of Vascular Research. 55:350-364 |
| بيانات النشر: | S. Karger AG, 2018. |
| سنة النشر: | 2018 |
| مصطلحات موضوعية: | 0301 basic medicine, Gibson assembly, Endothelium, Physiology, Genetic Vectors, Green Fluorescent Proteins, Immunoblotting, Gene Expression, Cell junction, Antibodies, Viral vector, Mice, 03 medical and health sciences, Live cell imaging, Human Umbilical Vein Endothelial Cells, medicine, Animals, Humans, Cloning, Molecular, beta Catenin, Fluorescent Dyes, Chemistry, Goats, Antibodies, Monoclonal, Endothelial Cells, Cadherins, Cell biology, Endothelial stem cell, Intercellular Junctions, 030104 developmental biology, medicine.anatomical_structure, Rabbits, gamma Catenin, Lamellipodium, Cardiology and Cardiovascular Medicine, mCherry |
| الوصف: | Endothelial cells of the vascular system are dynamic cells whose molecular adaptability is decisive for the adjustment of homeostasis and organ perfusion. Advanced microscopic techniques, automation processing, and image analysis software was shown to improve the understanding of vascular biology. In this work, we describe advanced methods that allow investigating the dynamics of endothelial cell contacts. The development of viral vectors has contributed significantly to the genetic manipulation of endothelial cells. We used the Gibson assembly as a quick and cheap cloning system for introducing sequences into the lentiviral-based pFUGW vector. Furthermore, classical fluorescence tags such as mCherry and EGFP were compared with self-labeling tags such as Halo and SNAP for their suitability to study junction dynamics in cultured endothelium, and found the self-labeling tags as useful tools. Using such combinations, we found maintained cell junction integrity during shear stress-induced junction remodeling using VE-cadherin-EGFP. Remodeling was accompanied by VE-cadherin plaque formation, indicating that this process is mediated by the formation of the actin-driven junction-associated intermittent lamellipodia, JAIL. The combined methods including the Gibson assembly, lentiviral mediated gene transfer, spinning disk-based live cell imaging, and software for quantification allow analyses of the endothelial cell junction dynamics under static and under shear stress conditions. |
| تدمد: | 1423-0135 1018-1172 |
| DOI: | 10.1159/000494933 |
| URL الوصول: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::eaa0024376da1f972b6799414ed2363f https://doi.org/10.1159/000494933 |
| Rights: | CLOSED |
| رقم الانضمام: | edsair.doi.dedup.....eaa0024376da1f972b6799414ed2363f |
| قاعدة البيانات: | OpenAIRE |
| تدمد: | 14230135 10181172 |
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| DOI: | 10.1159/000494933 |