ABSTRACT
A major problem for the in vitro engineering of larger tissue equivalents like those required in reconstructive surgery is the lack of solutions for sufficient nutrition and oxygenation. The starting point of our investigation was the question of whether the principles of in vitro angiogenesis can be applied and utilized for tissue engineering. A soft tissue model was developed, consisting of human adipose tissue stromal cells and umbilical vein endothelial cells in a fibrin-microcarrier scaffold. Capillary-like structures were visualized using UEA-I-lectin labelling and confocal laser scanning microscopy. Length of capillary-like structures was measured in an image analysis system. Under serum-free culture conditions, maintenance of capillary-like structures was significantly increased in comparison to serum-containing cultures. The application of vascular endothelial growth factor (VEGF) resulted in a high initial angiogenic response; long-term stabilization of capillary-like structures could not be achieved, however supplementation with IGF-1 resulted in the highest values and the slightest decrease in length of capillary-like structures, so that the results could be interpreted as an improved stabilization.
Subject(s)
Adipose Tissue/blood supply , Endothelium, Vascular/cytology , Neovascularization, Physiologic , Tissue Engineering/methods , Capillaries/growth & development , Cell Culture Techniques/methods , Cells, Cultured , Culture Media, Serum-Free , Endothelial Growth Factors/pharmacology , Humans , Image Processing, Computer-Assisted , Insulin-Like Growth Factor I/pharmacology , Lymphokines/pharmacology , Microscopy, Confocal , Neovascularization, Physiologic/drug effects , Statistics, Nonparametric , Umbilical Veins/cytology , Vascular Endothelial Growth Factor A , Vascular Endothelial Growth FactorsABSTRACT
The utilization of in vitro angiogenesis in tissue engineering might be useful in order to establish an artificial vascular network. However, it remains unclear how far the in vitro preformation of vascular structures may contribute to the perfusion of larger artificial tissue aggregates regarding the improvement of oxygenation and nutrition. In an in vitro study, we developed a model of a vascularized tissue. Stromal cells of a target tissue, e.g., adipose tissue or bone tissue, were expanded in vitro and seeded onto microcarriers or microparticles. Densely covered microcarriers were brought into a fibrin matrix together with endothelial cells. In order to demonstrate the formation and stabilization of capillary-like structures, UEA-I labeled specimens were evaluated using laser scanning microscopy and digital image analysis. The stabilization of capillary-like structures was better with stromal cells from bone marrow than from adipose tissue. In one of the culture aggregates, the total length of capillary-like structures increased after 6 weeks of cultivation to up to 140 mm/mm3. Additional tests were performed utilizing hyperbaric oxygenation. In the oxygenation group, a significant increase in the length of capillary-like structures was found. The method implies the option of coculturing different tissue elements and of an in vitro preformation of vascularized tissues.
