Characterization of the human smooth muscle cell secretome for regenerative medicine

Smooth muscle cells (SMC) play a central role in maintaining the structural and functional integrity of muscle tissue. Little is known about the early in vitro events that guide the assembly of 'bioartificial tissue' (constructs) and recapitulate the key aspects of smooth muscle differentiation and development before surgical implantation. Biomimetic approaches have been proposed that enable the identification of in vitro processes which allow standardized manufacturing, thus improving both product quality and the consistency of patient outcomes. One essential element of this approach is the description of the SMC secretome, that is, the soluble and deposited factors produced within the three-dimensional (3D) extracellular matrix (ECM) microenvironment. In this study, we utilized autologous SMC from multiple tissue types that were expanded ex vivo and generated with a rigorous focus on operational phenotype and genetic stability. The objective of this study was to characterize the spatiotemporal dynamics of the first week of organoid maturation using a well-defined in vitro-like, 3D-engineered scale model of our validated manufacturing process. Functional proteomics was used to identify the topological properties of the networks of interacting proteins that were derived from the SMC secretome, revealing overlapping central nodes related to SMC differentiation and proliferation, actin cytoskeleton regulation, and balanced ECM accumulation. The critical functions defined by the Ingenuity Pathway Analysis included cell signaling, cellular movement and proliferation, and cellular and organismal development. The results confirm the phenotypic and functional similarity of the SMC generated by our platform technology at the molecular level. Furthermore, these data validate the biomimetic approaches that have been established to maintain manufacturing consistency.