Homeostatic Branch Angle Oscillation Influences Mammary Branching Morphogenesis: Regulation by MMP14-Mediated ECM Remodeling

Branching morphogenesis shapes the epithelial tissue architecture as proliferative branches navigate the extracellular matrix (ECM) microenvironment. In the mammary gland, pubertal terminal end buds (TEBs) drive stochastic ductal invasion to build a tree-like network. Although stem cell programs and hormonal regulation have been extensively characterized, the geometric principles underlying branch adaptation to ECM mechanics during morphogenesis remain incompletely defined. Here, using in vivo whole-mount imaging and ex vivo 3D organoid live imaging, we show that the mammary bifurcation angle occupies a characteristic range and, in organoids, that it oscillates dynamically during branch morphogenesis. We show that branch angles maintain a confined dynamic range with regular oscillations during both bud bifurcation and branch elongation, thereby supporting efficient branching morphogenesis. These angle dynamics are independent of major mammary-specific developmental growth and hormonal signaling, pointing to non-endocrine microenvironmental cues as potential regulators. Integrative analysis of branched epithelial RNA-seq and in vivo TEB microarray data further reveals the remodeling of cell adhesion pathways and epithelial–ECM interactions during network formation, with matrix metalloproteinase 14 (MMP14) specifically enriched in branched epithelia as a promising candidate mediator. Pharmacological inhibition of MMP14 disrupted branch angle oscillations and impaired organoid branching growth. Taken together, our findings, derived from murine tissue and organoid models, identify the homeostatic branch angle as a previously unrecognized biophysical signature of mammary branching that is maintained, in organoids, through MMP14-mediated ECM remodeling.