| Adipose tissue resides as a vital endocrine organ of the human body, and plays a critical role in energy storage, metabolism, and whole body homeostasis. There is currently a great clinical need for both adipose tissue substitutes as well as in vitro models to further understand adipose tissue pathologies. Adipose tissue engineering strategies provide great potential in the development of both soft tissue reconstruction therapies and in vitro adipose models. In this dissertation, we discuss our work in adipose tissue engineering using three-dimensional (3D) aqueous silk fibroin porous scaffolds and both human adipose-derived stem cells (ASCs) and human umbilical vein endothelial cells (HUVECs). We developed a 3D co-culture adipose construct of differentiated adipocytes and endothelial cells, to incorporate a vascular component, in which both adipose tissue structure and function were demonstrated. Specifically, endothelial cells aligned and organized to form lumen-like structures within differentiated adipocytes. The use of lentivirus transduction techniques to fluorescently label each cell source provided advantages in imaging co-cultures within 3D constructs. Additionally, we demonstrated that lentivirus transduction does not affect differentiation potential of ASCs. Following the development of 3D adipose constructs, we measured metabolic activity of both adipocyte monocultures and adipocyte/endothelial cocultures with respect to both lipogenic and lipolytic function. We found that the addition of endothelial cells in co-culture did not significantly impact the ability of adipocyte monocultures to undergo basal metabolic activity. However, upon exposure to hyperinsulinemic conditions, we found that endothelial cells in coculture were necessary to observe a physiological metabolic response with respect to lipogenesis and lipolysis. In summary, we developed a 3D in vitro adipose tissue construct that incorporated endothelial co-culture, exhibited metabolic function, and responded to hyperinsulinemia in a physiological manner. The strategies developed in this thesis offer new options for adipose tissue engineering and provides a basis for future work in soft tissue regeneration and 3D sustainable models. |
