VEGF Signaling

This volume provides a collection of protocols for studying and manipulating VEGF signaling pathways in vitro and in vivo, and in particular, aims to present a range of both firmly established and newly emerging technologies, including those that are amenable to aiding in drug discovery or pre-clinical investigations. Each of the three sections begins with an introductory overview followed by supporting relevant methods. A major advantage of this book is that each chapter not only contains methodological detail rarely seen in other literature, but also contains a section on key notes and troubleshooting advice. Part I focuses on quantification of specific VEGF and VEGF receptor isoforms, beginning with a summary on VEGF splice variants, their function and their regulation. Part II relates to the study of VEGF dependent signaling in vitro, starting with a comprehensive overview of VEGF-mediated signal transduction in the cardiovascular system that highlights recent discoveries in the field. Part III first details genetic mouse models, then discusses techniques for studying physiological angiogenesis in the developing mouse embryo. Written in the highly successful Methods in Molecular Biology series format, 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.Cutting-edge and thorough, VEFG Signaling: Methods and Protocols is a useful tool for researching and understanding the basic biology of VEGF signaling and translating it into the clinic.

Includes cutting-edge methods and protocols Provides step-by-step detail essential for reproducible results Contains key notes and implementation advice from the experts

Auteur

Lorna R. Fielder, Program in cardiovascular and Metabolic Disorders, Duke-NUS, Graduate Medical School Singapore, 8 College Road, Singapore 169857, Telephone: +65-6516-7666, Email: lorna.fiedler@gmail.com or l.fiedler@imperial.ac.uk.

Contenu

VEGF Splicing and the Role of VEGF Splice Variants: From Physiological-Pathological Conditions to Specific Pre-mRNA Splicing.- Detection and Quantification of VEGF Isoforms by ELISA.- Quantitation of Circulating Neuropilin-1 in Human, Monkey, Mouse and Rat Sera by ELISA.- Detection and Quantification of Vascular Endothelial Growth Factor Receptor Tyrosine Kinases in Primary Human Endothelial Cells.- Induction of VEGF Secretion in Cardiomyocytes by Mechanical Stretch.- Chromatin Immunoprecipitation Assay: Examining the Interaction of NFkB with the VEGF Promoter.- An Overview of VEGF-Mediated Signal Transduction.- Identification of Receptor Tyrosine Kinase Inhibitors using Cell Surface Biotinylation and Affinity Isolation.- Analysis of VEGF-Mediated ERK5 Activity in Endothelial Cells.- In Vitro Angiogenesis Assays.- Chemotactic Migration of Endothelial Cells Towards VEGF-A165.- Vasculogenesis and Angiogenesis in VEGF Receptor-1 Deficient Mice.- The Embryonic Mouse Hindbrain and Postnatal Retina as In Vivo Models to Study Angiogenesis.- VEGF Gene Transfer to the Utero-Placental Circulation of Pregnant Guinea Pigs to Enhance Fetal Growth.- VEGF Gene Transfer to the Utero-placental Circulation of Pregnant Sheep to Enhance Fetal Growth.- Generation of Targeted Mutations in Zebrafish Using the CRISPR/Cas System.