(496e) Human Skeletal Muscle Growth and Maturation in 3-Dimensional Silk-Extracellular Matrix Scaffolds

Injuries resulting in volumetric skeletal muscle loss (VML) are unable to heal and properly regenerate, resulting in a permanent functional deficit. Advances in tissue engineering have created new opportunities for treatments that use engineered muscle to replace the lost tissue. However, a thorough understanding of the material properties that lead to healthy skeletal muscle tissue regeneration as opposed to scar tissue formation is needed. The process of fibrosis and scar tissue formation following muscle injury relies on many variables, some of which are dependent upon the patient’s own wound healing rates and some which are dependent upon the formulation of the biomaterial implanted. Furthermore, previous work in our lab has demonstrated that the addition of bioactive components, such as extracellular matrix (ECM) and growth factors, to silk scaffolds alters in vivo degradation and resorption. Therefore, we aimed to understand the role of biomaterial formulation on muscle, vascular, and stromal cell behavior in vitro to guide future in vivo applications for repair of skeletal muscle defects. Specifically, we developed three-dimensional silk fibroin-based tissue constructs containing skeletal muscle derived ECM and growth factors (vascular endothelial growth factor VEGF, epidermal growth factor EGF). Engineered muscle tissue was formed using human skeletal muscle cells with and without human endothelial cells (HUVECs, Lonza®) to create engineered tissues with desired properties for future in vivo applications. Cell phenotypes within the scaffold were evaluated by immunohistochemistry and Western blot. The results showed that the method of growth factor presentation during 3D culture impacted human skeletal myoblast phenotype and maturation through shifts in desmin, myogenin, pax7, and alpha-actinin expression in human skeletal muscle and CD-31 (PECAM-1), von Willebrand factor, and VE-cadherin expression in human umbilical vein endothelial cells. Therefore, the rational design of 3D scaffolds for implantation in vivo based on in vitro analyses should improve engineered tissue integration, and the patient’s muscle function following biomaterial resorption and degradation. Preliminary results confirmed biomaterial formulation impacts engineered primary rodent skeletal muscle pre-conditioning within a bioreactor system, resulting in vivo functionality following pre-conditioned scaffold implantation.