Abstract
A challenge in tissue engineering biomimetic models for studying angiogenesis is building the physiological complexity of real microvascular networks. Our laboratory recently introduced the rat mesentery culture model as an ex vivo experimental platform for investigating multicellular dynamics involved in angiogenesis within intact microvascular networks. The objective of this study was to compare endothelial cell phenotypes along capillary sprouts in cultured ex vivo rat mesentery microvascular networks to in vivo endothelial cell phenotypes. For Day 3 (Ex Vivo) tissues, adult rat mesentery tissues were cultured for three days in media supplemented with 10% serum. For Day 3 (In Vivo) tissues, adult rats were anesthetized and the mesentery was exteriorized for twenty minutes to induce angiogenesis. Microvascular networks from Day 3 (Ex Vivo) and Day 3 (In Vivo) groups were angiogenic, characterized by an increase in vessel density, capillary sprouting, and identification of similar BrdU-positive endothelial cell distributions along sprouts. Endothelial cells in both groups extended pseudopodia at the distal edge of capillary sprouts and displayed similar endothelial cell UNC5b, VEGFR-2, and CD36 labeling patterns. The results from this study support the physiological relevance of the rat mesentery culture model and highlight its novelty as a biomimetic tool for angiogenesis research.
Highlights
Common bottom-up tissue engineering models attempt to recapitulate the physiological process associated with angiogenesis through the combination of multiple cell types, soluble growth factors, and extracellular matrix proteins
Our results suggest that endothelial cells along capillary sprouts in cultured mesentery microvascular networks 1) maintain an angiogenic phenotype similar to in vivo characterized by VEGFR-2 and UNC5b labeling patterns, 2) extend actin-positive pseudopodia at the distal edges, and 3) display a CD36-low phenotype that diminishes along the length of the capillary
The main contribution of this study is the demonstration that angiogenic ex vivo endothelial cells in cultured rat mesentery microvascular networks maintain a phenotype similar to angiogenic in vivo endothelial cells. This is the first demonstration of an ex vivo model that maintains endothelial cell phenotypes during culture, which is the goal for angiogenesis biomimetic models
Summary
Common bottom-up tissue engineering models attempt to recapitulate the physiological process associated with angiogenesis through the combination of multiple cell types, soluble growth factors, and extracellular matrix proteins. The innate complexity of the mesentery tissue, which includes blood vessels, lymphatic vessels, interstitial cells, and nerves, combined with the time-lapse capability highlights the advantages of the rat mesentery culture model as an alternative strategy to tissue engineering approaches, which often lack the diversity of cell types and signaling cues that collectively influence endothelial cell behavior during angiogenesis. Previous work from our laboratory focused on demonstrating the feasibility of stimulating angiogenesis and time-lapse imaging with the rat mesentery culture model[14,16] The findings from this current study confirm that ex vivo cultured endothelial cells in rat mesentery tissue maintain a phenotype similar to in vivo endothelial cells and support the physiological relevance of the rat mesentery culture model as a tool to investigate endothelial cell dynamics during angiogenesis within an intact microvascular network. As this type of phenotypic characterization is surprisingly atypical of biomimetic model development, our results offer an important comparative description and suggest a new benchmark for model validation studies
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