Thermoreversible cell-derived extracellular matrix only hydrogel (CEOgel): Development, characterization, and applications

Decellularized extracellular matrix (dECM) has been widely used as a biomimetic material for three-dimensional cell culture and tissue regeneration. Although tissue-derived ECM is the conventional source of dECM, cell-derived ECM (cECM) has emerged as an attractive alternative. Various cECM-based formulations such as powders, films, and preformed gels have been reported, but thermosensitive cECM hydrogels remain largely unexplored. Here, we report a novel method to produce a thermoreversible hydrogel exclusively made from cECM, termed CEOgel. Once in vitro–cultured umbilical cord mesenchymal stem cells were decellularized, cECM solubility was enhanced and its nanofibers were concentrated to generate CEOgel without additional factors. This fabrication strategy was applicable across multiple cell types and consistently yielded homogeneous gels with minimal donor- or batch-dependent variability. CEOgel exhibited sufficient mechanical stability for in vitro use and formed gels in vivo following injection, confirming its thermosensitive and biocompatible nature. It also served as a functional 3D matrix, supporting endothelial vascularization in microfluidic chips and the growth of colorectal cancer organoids. Proteomic profiling revealed that CEOgel incorporates a broad spectrum of proteins commonly expressed across human tissues. Additionally, we demonstrated that CEOgel properties can be tuned through transition-metal crosslinking and genetically engineering ECM-producing cells. Together, this study proposes a new class of thermoreversible cECM hydrogels that eliminate reliance on animal tissues or external cross-linkers while expanding the applicability of cECM materials and advancing conceptual diversity for ECM hydrogel design. Our findings highlight the potential of CEOgel as a regenerative biomaterial for tissue engineering and medical applications. • Thermoreversible CEOgel fabricated solely from decellularized cECM. • Method works across cell types and yields consistent, low-variation gels. • CEOgel contains many growth factors and universal tissue proteins. • CEOgel properties are tunable via metal crosslinking and gene engineering. • CEOgel supports vasculature and organoids and shows excellent biosafety.