Shira Landau

Macrophages are key contributors to vascularization, but the mechanisms behind their actions are not understood. Here, we show that diverse macrophage phenotypes have distinct effects on endothelial cell behavior, with resulting effects on vascularization of engineered tissues. In Transwell coculture, proinflammatory M1 macrophages caused endothelial cells to up-regulate genes associated with sprouting angiogenesis, whereas prohealing (M2a), proremodeling (M2c), and anti-inflammatory (M2f) macrophages promoted up-regulation of genes associated with pericyte cell differentiation. In 3D tissue-engineered human blood vessel networks in vitro, short-term exposure (1 day) to M1 macrophages increased vessel formation, while long-term exposure (3 days) caused regression. When human tissue-engineered blood vessel networks were implanted into athymic mice, macrophages expressing markers of both M1 and M2 phenotypes wrapped around and bridged adjacent vessels and formed vessel-like structures themselves. Last, depletion of host macrophages inhibited remodeling of engineered vessels, infiltration of host vessels, and anastomosis with host vessels.

Graft vascularization remains one of the most critical challenges facing tissue-engineering experts in their attempt to create thick transplantable tissues and organs. In vitro prevascularization of engineered tissues has been suggested to promote rapid anastomosis between the graft and host vasculatures; however, thrombotic events have been reported upon graft implantation. Here, we aimed to determine whether in vitro vessel maturation in transplantable grafts can accelerate vascular integration and graft perfusion and prevent thrombotic events in the grafts. To this end, endothelial cells and fibroblasts were cocultured on 3D scaffolds for 1, 7, or 14 d to form vasculature with different maturation degrees. Monitoring graft-host interactions postimplantation demonstrated that the 14-d in vitro-cultured grafts, bearing more mature and complex vessel networks as indicated by elongated and branched vessel structures, had increased graft-host vessel anastomosis; host vessel penetration into the graft increased approximately eightfold, and graft perfusion increased sixfold. The presence of developed vessel networks prevented clot accumulation in the grafts. Conversely, short-term cultured constructs demonstrated poor vascularization and increased thrombus formation. Elevated expression levels of coagulation factors, von Willebrand factor (vWF), and tissue factor (TF), were demonstrated in constructs bearing less mature vasculature. To conclude, these findings demonstrate the importance of establishing mature and complex vessel networks in engineered tissues before implantation to promote anastomosis with the host and accelerate graft perfusion.

Cultured meat can provide a sustainable and more ethical alternative to conventional meat. Most of the research in this field has been focused on developing muscle tissue, as it is the main component of meat products, while very few studies address cultured fat tissue, an essential component in the human diet and determinant of meat quality, flavor, juiciness, and tenderness. Here, we engineered bovine fat tissue for cultured meat and incorporated it within engineered bovine muscle tissue. Mesenchymal stem cells (MSCs) were derived from bovine adipose tissue and exhibited the typical phenotypic profile of adipose-derived MSCs. MSC adipogenic differentiation and maturation within alginate-based three-dimensional constructs were optimized to yield a fat-rich edible engineered tissue. Subsequently, a marble-like construct, composed of engineered bovine adipose and muscle tissues, was fabricated, mimicking inter- and intra-muscular fat structures.