Abstract
Semaphorins (SEMA) are chemorepulsive guidance cues that, acting through plexin receptors, inhibit integrin-mediated cell adhesion to the extracellular matrix. The ensuing cell retraction and collapse is a key biological event downstream of SEMA/plexin signaling that is however hard to precisely quantify. Here, we describe a quantitative approach that allows monitoring over time the evolution of SEMA3E/plexin D1-elicited endothelial cell collapse. This method exploits the xCELLigence platform, an electrical impedance-based system in which microelectronic sensor arrays are integrated into the bottom of microplate wells. Measuring electrical impedance allows real-time monitoring of changes in endothelial cell morphology and adhesion induced by SEMA3E via plexin D1. Afterwards, analogic electrical impedance measurements are converted into digital numeric signals that can then be analyzed by mathematical and statistical methods.
Federico Bussolino and Guido Serini share the senior authorship of this manuscript.
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Acknowledgments
C. Camillo and G. Serini conceived the project and designed the experiments in collaboration with N. Gioelli. C. Camillo and N. Gioelli performed the experiments. C. Camillo, N. Gioelli, and G. Serini analyzed the data. C. Camillo, N. Gioelli, and G. Serini wrote the manuscript. F. Bussolino provided the XCELLigence system. G. Serini oversaw the project. G. Serini and F. Bussolino share the senior authorship of this manuscript. The authors acknowledge financial support from Associazione Italiana per la Ricerca sul Cancro (AIRC) (to G.S., IG #9211, to F.B., IG #10133); AIRC 2010 Special Program in Molecular Clinical Oncology 5 × 1000 Project no. 9970 (to F.B.), Associazione Augusto per la Vita (to G.S.); Fondazione Piemontese per la Ricerca sul Cancro, MIUR 2010—Vaschetto 5 × 1000 2010 (to G.S.); the University of Torino-Compagnia di San Paolo: RETHE grant-ORTO11RKTW (to F.B.); and Telethon, Italy (to G.S.).
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Camillo, C., Gioelli, N., Bussolino, F., Serini, G. (2017). An Electrical Impedance-Based Method for Quantitative Real-Time Analysis of Semaphorin-Elicited Endothelial Cell Collapse. In: Terman, J. (eds) Semaphorin Signaling. Methods in Molecular Biology, vol 1493. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6448-2_14
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DOI: https://doi.org/10.1007/978-1-4939-6448-2_14
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