Nanofiber-Supported Decellularized Matrix Membrane Creates a Biomimetic Biochemical and Biophysical Niche for Intestinal Organoid Epithelium.

Reconstructing intestinal organoid-derived epithelium on porous membranes is a promising strategy for building in vitro intestine models that combine biological complexity with experimental accessibility. A central challenge is designing membranes that reproduce the biochemical and biophysical cues of the native intestinal microenvironment. Here, we report a nanofiber-supported decellularized extracellular matrix (NaDE) membrane engineered to emulate these key features. Specifically, a colon-specific NaDE (C-NaDE) membrane is fabricated by integrating porcine colon-derived decellularized extracellular matrix (CdECM) hydrogel with an ultra-thin nanofiber scaffold. The resulting membrane preserves a collagen-rich extracellular matrix (ECM) composition and exhibits basement membrane-level stiffness (∼230 kPa), thereby providing a supportive niche for intestinal epithelial reconstruction. Colon organoid-derived epithelium cultured on the C-NaDE membrane shows enhanced expression of genes associated with stem cell maintenance and epithelial proliferation. Notably, the C-NaDE membrane also increases markers related to secretory epithelial lineages that are typically underrepresented in conventional synthetic stiff membrane systems, including enteroendocrine, tuft, and Paneth cell populations. These findings demonstrate that integrating organ-specific matrix composition with tissue-relevant mechanics can improve membrane-based intestinal epithelial models. The C-NaDE membrane, therefore, provides a physiologically enhanced platform for studying intestinal biology, disease mechanisms, barrier function, and drug responses.