Selective induction of sprouting and intussusception is associated with the concentration distributions of oxygen and hypoxia-induced VEGF.

Sprouting and intussusception are two important modes of capillary angiogenesis, the mechanisms of selective induction of which remain unclear. In this study, we focus on the two developing tissues of yolk sac and skeletal muscle of 2-week-old rat and try to explain the mechanisms to induce selectively sprouting and intussusception in a new way to combine numerical calculation, experimental observations and schematic simulation. We propose the concept of capillary network unit and show that the concentration and gradient of oxygen/hypoxia-induced VEGF around straight segments are lower/higher than that around vascular bifurcations; sprouting mainly occurs at straight segments and intussusception at vascular bifurcations. The results indicate that the locations susceptible to sprouting and intussusception are determined by the distribution characteristics of oxygen/hypoxia-induced VEGF in the capillary network unit. Furthermore, it is considered that the flow dynamics at these locations also play important roles, namely laminar flow at straight segments promotes sprouting, and flow disruption at bifurcations promotes intussusception. Our work suggests the presence of the location preference for sprouting and intussusception, and provides a new research perspective to reveal its core mechanisms.

Different angiogenesis modes and endothelial responses in implanted porous biomaterials.

An in vivo experimental model based on implanting porous biomaterials to study angiogenesis was proposed. In the implanted porous polyvinyl alcohol, three major modes of angiogenesis, sprouting, intussusception and splitting, were found. By electron microscopy and three-dimensional simulation of the angiogenic vessels, we investigated the morphological characteristics of the three modes and paid special attention to the initial morphological difference between intussusception and splitting, and it was confirmed that the endothelial abluminal invagination and intraluminal protrusion are pre-representations of intussusception and splitting, respectively. Based on immunohistochemical analysis of HIF-1α, VEGF and Flt-1 expressions, it was demonstrated that the dominant mode of angiogenesis is related to the local hypoxic condition, and that there is difference in the response of endothelial cells to hypoxia-induced VEGF between sprouting and splitting. Specifically, in the biomaterials implanted for 3 days, the higher expression and gradient of VEGF induced by severe hypoxia in the avascular area caused sprouting of the peripheral capillaries, and in the biomaterial implanted for 9 days, with moderate hypoxia, splitting became a dominant mode. Whether on day 3 or day 9, Flt-1 expression in sprouting endothelia was significantly higher than that in splitting endothelia, which indicates that sprouting is caused by the strong response of endothelial cells to VEGF, while splitting is associated with their weaker response. As a typical experimental example, these results show the effectiveness of the porous biomaterial implantation model for studying angiogenesis, which is expected to become a new general model.

Fractal characteristics of the microvascular network: A useful index to assess vascularization level of porous silk fibroin biomaterial.

The neovascularization of biomaterials for tissue engineering is not only related to growth of capillaries but also involves appropriate hierarchy distribution of the microvessels. In this study, we proposed hierarchy distribution contrast method which can assess vascular transport capacity, in order to examine the hierarchy distribution of the neovessels during vascularization of the porous silk fibroin biomaterials implanted into rats and its evolution. The results showed that the fractal characteristics appeared toward the end of the vascularization stages, and the structure of the microvascular network after 3 weeks of implantation was similar to the fractal microvascular tree with bifurcation exponent x = 3 and fractal dimension D = 1.46, which became a sign of maturation of the regenerative vasculature. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2276-2290, 2017.

Endothelial tubes form from intracellular vacuoles in implanted biomaterial in vivo of rat.

When the porous biomaterials are used to implant in vivo of animal for repairing wound, the angiogenesis in microenvironment of porous biomaterial is a key process in order to achieve the goal of treatment. While clarifying the process of vascularization and its mechanism is of great significance for design and development of medical biomaterials. In this area, it is noted that the endothelial tubes of new capillaries are formed by intracellular vacuoles, which has been proved in vitro model of angiogenesis. However, there is still no conclusive evidence in vivo model for mammals. By experimental tracking and observation of angiogenesis in the biomaterials implanted in rats, the angiogenesis process and the characteristics were explored. This study focused on the behavior of endothelial cell (EC)s and the capillary lumen formatting from EC cord in sprouting. Through marking and observing the ECs, the experimental evidences of angiogenesis after implanted materials into rats were obtained, which including various stages, such as rapidly proliferating of ECs, assembling of ECs to build up cell cord and vacuoles formation in ECs. An important mechanism of lumen formation for mammal in vivo was proved, which complemented the experimental results of the assembly of endothelial tubes in vivo through the formation and fusion of vacuoles for transgenic zebrafish. Our results provide support for the model of lumen formation of new capillary in mammal.

Role of vascular endothelial progenitor cells in construction of new vascular loop.

Since bone marrow-derived endothelial progenitor cells (EPCs) have been detected in adult peripheral blood, the mode of vasculogenesis in the adult tissue has caught attention in field of vascularization research. To confirm the role of EPCs in construction of new vascular loop, we took the biomaterial scaffold implanted into adult rat as an experimental model to observe and examine the actions of the EPCs in neovascularization of the material by immunohistochemistry and transmission electron microscopy. Additionally, by establishing a chemotactic migration model for vascular endothelial cells (ECs) and EPCs, the migrations of ECs and EPCs were explored in simulations. The results of 20,000 simulations showed that the number of the vascular loops assisted by the EPCs was 2-5 times that of the vascular sprouts being naturally joined. Based on the results of experiments and simulations, we conclude that the EPCs are able to assist the angiogenic sprouts in joining under the condition of plenty of the EPCs being mobilized, which aggregate at sites close to sprout tips, forming a cell cord and differentiating to ECs in situ, and become vessel segments between neighboring sprouts. This suggests that there is a difference between the adult and embryo in the manner of vasculogenesis and that a small number of EPCs can play an important role to make the new blood vessels achieve rapid functionalization.