Volumetric analysis of the terminal ductal lobular unit architecture and cell phenotypes in the human breast

The major lactiferous ducts of the human breast branch out and ultimately end at terminal ductal lobular units (TDLUs). These glandular structures are the source of milk upon lactation, and also the most common origin of breast cancer. Despite their functional and clinical importance, the three dimensional (3D) architecture of TDLUs has remained undetermined, largely due to their absence in rodent animal models. By utilizing recent technological advances in optical tissue clearing, 3D reconstruction histology and microscopy, we imaged the 3D structure of healthy human breast tissue to determine whether general branching patterns or cell type specific niches exist in the TDLU. Our data demonstrate that highly branched TDLUs, also exhibiting elevated proliferation, are uncommon in the breast tissue regardless of donor age, parity or hormonal contraception. Overall, TDLUs have a consistent shape and their branch parameters are largely comparable between different TDLUs and individuals irrespective of donor age or parity. Simulation of TDLU branching morphogenesis in 3D by mathematical modelling suggests that evolutionarily conserved mechanisms regulate mammary gland branching in humans and mice, despite their anatomical differences. The data also demonstrate a new level of organization within the TDLU structure identifying a main subtree that dominates in bifurcation events and exhibits a more duct-like keratin expression pattern. In all, our data provide the first structural insights into 3D human breast anatomy and branching, and exemplify the power of volumetric imaging in deeper understanding of human breast biology. ### Competing Interest Statement The authors have declared no competing interest.