3D ECM-Rich Environment Sustains the Identity of Naïve Human iPSCs

The establishment of in vitro naive human pluripotent stem cell cultures opened new perspectives for the study of early events in human development. The role of several transcription factors and signaling pathways during maintenance of human naive pluripotency has been characterized. However, little is known about the role exerted by the extracellular matrix and its three-dimensional organization. Here, using an unbiased and integrated approach combining transcriptional, proteomic and secretome analyses, we found that naive, but not primed, hiPSC colonies are characterized by a self-organized extracellular matrix (ECM)-rich microenvironment. Based on this, we developed a 3D culture system that supports robust long-term feeder-free self-renewal of naive hiPSCs, and also allows direct and timely developmental morphogenesis simply by modulating the signalling environment. Our study opens new perspectives for future applications of naive hiPSCs to study critical stages of human development in 3D starting from a single cell.