Quantification of microvascular remodeling as a meaningful discovery tool requires mapping and measurement of site-specific changes within vascular trees and networks. Vessel density and other critical vascular parameters are often modulated by molecular regulators as determined by local vascular architecture. For example, enlargement of vessel diameter by vascular endothelial growth factor (VEGF) is restricted to specific generations of vessel branching (Parsons-Wingerter et al., Microvascular Research72: 91, 2006). The averaging of vessel diameter over many successively smaller generations is therefore not particularly useful. The newly automated, user-interactive software VESsel GENeration Analysis (VESGEN) quantifies major vessel parameters within two-dimensional (2D) vascular trees, networks, and tree-network composites. This report reviews application of VESGEN 2D to angiogenic and lymphangiogenic tissues that includes the human and murine retina, embryonic coronary vessels, and avian chorioallantoic membrane. Software output includes colorized image maps with quantification of local vessel diameter, fractal dimension, tortuosity, and avascular spacing. The density of parameters such as vessel area, length, number, and branch point are quantified according to site-specific generational branching within vascular trees. The sole user input requirement is a binary (black/white) vascular image. Future applications of VESGEN will include analysis of 3D vascular architecture and bioinformatic dimensions such as blood flow and receptor localization. Branching analysis by VESGEN has demonstrated that numerous regulators including VEGF 165 , basic fibroblast growth factor, transforming growth factor b-1, angiostatin and the clinical steroid tri- Microvascular remodeling is now widely acknowledged as fundamental to normal physiological processes that include embryonic development, reproductive biology and healthy wound-healing, and the progressive pathologies of neovascular diseases such as cancer, diabetes, and heart disease (Folkman, 2007). However, angiogenesis, lymphangiogenesis, and other microvascular remodeling processes are difficult to map and quantify because of the morphological complexity of branching vascular trees and their associated capillary networks. Furthermore, the architecture of a vascular tree or network is locally adapted to specific needs of the host tissue or organ. In general, blood and lymphatic vascular structures can be classified as (1) heterogeneous, asymmetric trees of vessels that branch and taper, (2) relatively homogeneous, symmetric networks or plexuses, or (3) tree-network composites. Mature vascular trees typically develop from immature, capillary-like vasculogenic networks. Within a mature organ or tissue, capillary networks are necessarily continuous with their arterial and venous trees.The computer software VESsel GENeration Analysis (VESGEN) maps and quantifies major parameters of angiogenesis and lymphangiogenesis in vascular trees and networks. This review descr...

Pseudocolor view of vascular branching generations in the chorioallantoic membrane (CAM) of quail. Vascular architecture was analyzed using the automated, user‐interactive software, VESsel GENeration Analysis (VESGEN). See Vickerman, et al., on page 320, in this issue.

Development of effective vascular therapies requires the understanding of all modes of vessel formation involved in angiogenesis (here termed "hemangiogenesis") and lymphangiogenesis. Two major modes of vessel morphogenesis include sprouting of a new vessel from a preexisting vessel and splitting of a preexisting parent vessel into two offspring vessels. In the quail chorioallantoic membrane (CAM) during mid-development (embryonic days E6 -E9), lymphangiogenesis progressed primarily via blind-ended vessel sprouting. Isolated lymphatic endothelial progenitor cells were recruited to the tips of growing vessels. During concurrent hemangiogenesis, parent blood vessels expanded from the capillary network and split into offspring vessels, accompanied by transient capillary expression of alpha smooth muscle actin (␣SMA) and recruitment of polarized mural progenitor cells. Lymphatics and blood vessels were identified by confocal/fluorescence microscopy of vascular endothelial growth factor (VEGF) receptor VEGFR-2, ␣SMA (specific to CAM blood vessels), homeobox transcription factor Prox1 (specific to lymphatics), and the quail hematopoetic marker, QH-1. VEGFR-2 was expressed intensely in isolated cells and lymphatics, and moderately in blood vessels. Prox1 was absent from isolated progenitor cells prior to lymphatic recruitment. Exogenous vascular endothelial growth factor-165 (VEGF 165 ) increased blood vessel density and anastomotic frequency without changing endogenous modes of vascular/lymphatic vessel formation or marker expression. Although VEGF 165 is a key cellular regulator of hemangiogenesis and vasculogenesis, the role of VEGF 165 in lymphangiogenesis is less clear. Interestingly, VEGF 165 increased lymphatic vessel diameter and density as measured by novel Euclidean distance mapping, and the antimaturational dissociation of lymphatics from blood vessels, accompanied by lymphatic reassociation into homogeneous networks. Published 2006 Wiley-Liss, Inc. †

Vessel density oscillated with the progression of DR. Alternating phases of angiogenesis/neovascularization and vascular dropout were dominated first by remodeling of arteries and subsequently by veins.