Critchley, William R; Fearnley, Gareth WF; Abdul-Zani, Izma; Molina-Paris, Carmen; Bendtsen, Claus; Zachary, Ian C; Harrison, Michael A; Critchley, William R; Fearnley, Gareth WF; Abdul-Zani, Izma; Molina-Paris, Carmen; Bendtsen, Claus; Zachary, Ian C; Harrison, Michael A; Ponnambalam, Sreenivasan; - view fewer (2022) Monitoring VEGF-Stimulated Calcium Ion Flux in Endothelial Cells. Methods in Molecular Biology , 2475 pp. 113-124. 10.1007/978-1-0716-2217-9_7.

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Abstract

The endothelial response to vascular endothelial growth factor A (VEGF-A) regulates many aspects of animal physiology in health and disease. Such VEGF-A-regulated phenomena include vasculogenesis, angiogenesis, tumor growth and progression. VEGF-A binding to receptor tyrosine kinases such as vascular endothelial growth factor receptor 2 (VEGFR2 ) activates multiple signal transduction pathways and changes in homeostasis, metabolism, gene expression, cell proliferation, migration, and survival. One such VEGF-A-regulated response is a rapid rise in cytosolic calcium ion levels which modulates different biochemical events and impacts on endothelial-specific responses. Here, we present a series of detailed and robust protocols for evaluating ligand-stimulated cytosolic calcium ion flux in endothelial cells. By monitoring an endogenous endothelial transcription factor (NFATc2 ) which displays calcium-sensitive redistribution, we can assess the relevance of cytosolic calcium to protein function. This protocol can be easily applied to both adherent and non-adherent cultured cells to evaluate calcium ion flux in response to exogenous stimuli such as VEGF-A.

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