Novel three-dimensional analysis tool for vascular trees indicates complete micro-networks, not single capillaries, as the angiogenic endpoint in mice overexpressing human VEGF(165) in the brain

To adequately supply tissues with oxygen and nutrients, the formation of functional vascular networks requires generation of normal, healthy vessels and their arrangement into an effective network architecture. While our knowledge about the development of single vessels significantly increased during the last years, mechanisms responsible for network formation are still poorly understood. This is probably due to the lack of suitable methods for quantification of structural properties of microvascular networks. Previously we showed that cerebral blood flow is not increased in mice exhibiting a 2- to 3-fold higher density of normal and perfused capillaries as a result of transgenic overexpression of the human vascular endothelial growth factor (VEGF(165)). Here we used vascular corrosion casting and hierarchical micro-computed tomography combined with a new network analysis tool to characterize the vascular architecture in gray and white matter of these mice. Our results indicate that VEGF overexpression leads to formation of additional micro-networks connected to higher order vessels rather than insertion of individual capillaries into the existing vessel structure. This implies that the smallest "angiogenic quantum", i.e. the final, stable result of angiogenesis and subsequent remodeling, is not a single microvessel, but a complete micro-network. In conclusion, high-resolution 3D imaging combined with network analysis can substantially improve our understanding of vascular architecture, beneficial for the development of therapeutic angiogenesis as a clinical tool for applications such as wound healing or treatment of ischemic diseases.