Developmental pathways pervade stem cell responses to evolving extracellular matrices of 3D bioprinted microenvironments

Developmental studies and 3D in vitro model systems show the production and engagement of extracellular matrix (ECM) often precedes stem cell differentiation.   Yet unclear is how the ECM triggers signaling events in sequence to accommodate multistep processes characteristic of differentiation. Here we employ transcriptome profiling and advanced imaging to delineate the specificity of ECM engagement to particular differentiation pathways and to determine whether specificity in this context is a function of long term ECM remodeling.  To this end, human mesenchymal stem cells (hMSCs) were cultured in 3D bioprinted matrices created from ECM proteins and associated controls. We found exogenous ECM provided in 3D microenvironments at early time points impacts on the composition of microenvironments at later time points and that each evolving 3D microenvironment is uniquely poised to promote stem cell differentiation. Moreover, 2D cultures undergo minimal ECM remodeling and are ill-equipped to stimulate pathways associated with development. 3D environments composed ofbovine serum albumin, collagen I, fibronectin, and laminin was bioprinted using multiphoton-excited photochemistry. Human mesenchymal stem cell transcriptome modification due to interaction with 3D environments examined via RNA seq.