Agarose-Based Core-Shell Scaffolds Facilitate Mammary Epithelial Tubulogenesis

Three-dimensional mammary epithelial cultures are important for modeling tissue organization and developing cell-based milk technologies, yet conventional Matrigel systems provide limited control over geometry, confinement, and long-range tissue remodeling. Here, we developed a Matrigel-agarose core-shell bio-casting platform that separates biochemical and structural cues by confining MCF10A cells or MCF10A organoids-laden Matrigel within a hollow agarose shell. This defined architecture supported the formation of elongated mammary epithelial mini-tissues and induced dynamic morphological remodeling, including longitudinal compaction, lateral tissue reorganization, and budding-like protrusions. Compared to MCF10A cultured in Matrigel without the agarose shell, agarose core-shell constructs showed enhanced epithelial and lumen-like organization, as indicated by increased Ep-CAM and MUC1 expression and enlarged lumen-like regions. Strikingly, core-shell filaments developed a transparent tube-like phenotype with visible internal structures and retained epithelial marker expression. Gene-expression analysis showed increased expression of selected mammary- and lipid-associated markers, including Perilipin-2, fatty acid synthase, and casein, suggesting early mammary-associated molecular expression. Together, these results established Matrigel-agarose core-shell bio-casting as a modular culture strategy for guiding mammary epithelial tubulogenesis in vitro.