Quantitative Study of In Vivo Angiogenesis and Vasculogenesis Using Matrigel-Based Assays

Abstract

Neovascularization, the formation of new blood vessels, is critical for various physiological and pathological events such as wound repair and tumor growth as well as for functional tissue engineering as it ensures proper nutrient and oxygen supply to newly forming tissues. Neovascularization can occur via angiogenesis, where new capillaries sprout from pre-existing vessels, and/or vasculogenesis, where endothelial cells spontaneously self-assemble into vascular structures. Because neovascularization is the rate-limiting step in new tissue formation, there is great interest in developing robust quantitative approaches that facilitate greater understanding of this complex process. Matrigel-based assays are particularly useful for studying neovascularization as it provides a more natural environment for endothelial cell recruitment and capillary formation. In addition, Matrigel-based assays are reproducible and easy to perform when compared to other available in vivo angiogenesis assays. We have recently developed a novel matrigel assay that permits quantification of the endogenous angiogenic response in vivo by determining the number of recruited endothelial cells using fluorescence-activated cell sorting (FACS). We have also developed a technique to quantitatively evaluate vasculogenesis where endothelial cells and mesenchymal stem cells are suspended together in Matrigel prior to subcutaneous injection and vascular network formation is quantitatively analyzed from whole-mount confocal images of lectin-perfused Matrigel implants. Here, we outline the utility of these Matrigel assays for quantitative analyses of both angiogenic response and vasculogenesis and provide a detailed description of the methodology involved.