Main description:

This volume provides an updated collection of protocols for manipulating and studying VEGF signaling pathways in vitro and in vivo and aims to present a range of both firmly established and newly emerging technologies. Covering multiple model species, from mouse to zebrafish to human, the book explores the role of VEGF and VEGFR isoforms in exosomes, cultured cells, or in tissues, as well as robust cell assays for the investigation of basic angiogenic mechanisms and VEGF signaling in more complex cellular systems, amongst other subjects. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.
Authoritative and up-to-date, VEGF Signaling: Methods and Protocols, Second Edition provides a useful tool for researchers in the vascular biology community and beyond in understanding the basic biology of VEGF signaling and in translating this research into the clinic.

Contents:

1. Opposing Roles of Vascular Endothelial Growth Factor C in Metastatic Dissemination and Resistance to Radio/Chemotherapy: Discussion of Mechanisms and Therapeutic Strategies

Christopher Montemagno, Frederic Luciano, and Gilles Pages

2. Identification of VEGF Isoforms in Mouse, Rat, and Zebrafish Using RT-qPCR

Patricia de Winter

3. Multi-Parameter Fluorescence-Activated Cell Sorting of Human Lymphatic Endothelial Cells

Theresa Connor, Nerida Sleebs, and Zerina Lokmic-Tomkins

4. Absolute Quantification of Plasma Membrane Receptors via Quantitative Flow Cytometry

Yingye Fang, Manasi Malik, Sarah K. England, and P.I. Imoukhuede

5. Measurement of VEGF Content in Exosomes and Subsequent Tumor Tubulogenesis and In Vivo Angiogenesis Functional Assays

Ye Zeng, Xiaoqiang Du, Wenli Jiang, and Yan Qiu

6. SH2-Domain Protein Isolation Using Synthetic Phosphorylated Peptides to Study VEGFR2 Signaling

Chiara Testini

7. Monitoring VEGF-Stimulated Calcium Ion Flux in Endothelial Cells

William R. Critchley, Gareth W.F. Fearnley, Izma Abdul-Zani, Carmen Molina-Paris, Claus Bendtsen, Ian C. Zachary, Michael A. Harrison, and Sreenivasan Ponnambalam

8. Co-Immunoprecipitation Assays

Ian M. Evans and Ketevan Paliashvili

9. Using Immortalized Endothelial Cells to Study the Roles of Adhesion Molecules in VEGF-Induced Signaling

James A.G.E. Taylor, Christopher J. Benwell, and Stephen D. Robinson

10. RNAscope for VEGF-A Detection in Human Tumor Bioptic Specimens

Tiziana Annese, Roberto Tamma, and Domenico Ribatti

11. Global MicroRNA Profiling of Vascular Endothelial Cells

Eloi Schmauch, Anna-Liisa Levonen, and Suvi Linna-Kuosmanen

12. Endothelial Cell Tube Formation Assay: An In Vitro Model for Angiogenesis

Mary Kelley, Sara Fierstein, Laura Purkey, and Kathleen DeCicco-Skinner

13. Use of a Thin Layer Assay for Assessing the Angiogenic Potential of Endothelial Cells In Vitro

James A.E. Lane, Ashton Faulkner, Elizabeth J.T. Finding, Eleanor G. Lynam, and Caroline P.D. Wheeler-Jones

14. VEGF-A165 -Induced Endothelial Cells Chemotactic Migration and Invasion Assays

Caroline Pellet-Many

15. Measuring Mitochondrial Calcium Fluxes in Cardiomyocyte upon Mechanical Stretch-Induced Hypertrophy

Daniela Ramaccini, Carlotta Giorgi, and Michelle L. Matter

16. Simultaneous Measurement of Endothelial Cell Proliferation and Cell Cycle Stage Using Flow Cytometry

Eleanor G. Lynam, James A.E. Lane, Elizabeth J.T. Finding, and Caroline P.D. Wheeler-Jones

17. Ex Vivo Mouse Aortic Ring Angiogenesis Assay

Vedanta Mehta and Marwa Mahmoud

18. Retinal Microvasculature-on-a-Chip for Modeling VEGF-Induced Permeability

Heloise Ragelle, Karen Dernick, Peter D. Westenskow, and Stefan Kustermann

19. Preventing VEGF-Mediated Vascular Permeability by Experimentally Potentiating BBB Characteristics in Endothelial Cells

Bo Kyoung Kim, Jeremie Canonica, Filip Roudnicky, and Peter D. Westenskow

20. The Embryonic Mouse Hindbrain and Postnatal Retina as In Vivo Models to Study Angiogenesis

Alessandro Fantin and Christiana Ruhrberg

21. Evaluating VEGF-Induced Vascular Leakage Using the Miles Assay

James T. Brash, Christiana Ruhrberg, and Alessandro Fantin

22. Modulation of VEGFA Signaling during Heart Regeneration in Zebrafish

Kaushik Chowdhury, Shih-Lei Lai, and Ruben Marin-Juez

23. Three-Dimensional Visualization of Blood and Lymphatic Vessels in the Adult Zebrafish Heart by Chemical Clearing

Lorna R. Fiedler, Paul R. Riley, and Roger Patient

24. Fluorescence-Activated Cell Sorting and Quantitative Real-Time PCR to Reveal VEGF Expressing Macrophage Populations in the Zebrafish Larvae

Andrew Herman, Alexander Greenhough, and David B. Gurevich

25. Assessing Molecular Regulation of Vascular Permeability Using a VEGF-Inducible Zebrafish Model

Luke H. Hoeppner