Bioreactor System to Perfuse Mesentery Microvascular Networks and Study Flow Effects During Angiogenesis

Abstract

A challenge for engineering models of angiogenesis is mimicking the physiological complexity of real microvascular networks. Utilizing an alternative top-down tissue culture approach, our laboratory developed the rat mesentery culture model as an ex vivo platform for investigating the multicellular dynamics involved during angiogenesis within an intact microvascular network. The objective of this study was to introduce physiologically relevant microvascular perfusion in cultured rat mesentery tissues and demonstrate its effect on angiogenesis. Adult male Wistar rat mesenteric tissues were harvested along with the main feeding artery and vein, and then transferred to a custom-designed biochamber for perfusion. The main feeding artery was cannulated with a 30G needle and secured in place with 7–0 suture. Single-pass perfusion was accomplished using a peristaltic pump in series with the biochamber placed inside an incubator set to standard culture conditions (37°C and 5% CO₂). Flow passed through the vasculature and drained out of the venous side to be collected in a waste reservoir. Tissues were cultured for 48 h with perfusion in the biochamber (Perfused) in serum-supplemented media to stimulate angiogenesis. Control tissues were cultured in biochambers without perfusion (Static). Injection with FITC-albumin through the cannulated artery identified the lumens of vessels across the hierarchy of intact microvascular networks and confirmed successful perfusion. Labeling with BSI-lectin identified endothelial cells along microvascular networks and confirmed perfused tissues undergo angiogenesis after 48 h in culture, characterized by an increase in capillary sprouting. The presence of physiologic levels of capillary fluid velocities and associated shear stresses attenuated the angiogenic response compared to static controls. These results demonstrate the effect of perfusion on angiogenesis and establishes the novelty of the rat mesentery culture model as an experimental platform that incorporates perfusion with real microvascular networks in an ex vivo environment.

Impact Statement

Microvascular remodeling, or angiogenesis, plays a central role in multiple pathological conditions, including cancer, diabetes, and ischemia. Tissue-engineered in vitro models have emerged as tools to elucidate the mechanisms that drive the angiogenic process. However, a major challenge with model development is recapitulating the physiological complexity of real microvascular networks, including incorporation of the entire vascular tree and hemodynamics. This study establishes a bioreactor system that incorporates real microvascular networks with physiological flow as a novel ex vivo tissue culture model, thereby providing a platform to evaluate angiogenesis in a physiologically relevant environment.