Sustained postconfluent culture of human mammary epithelial cells enriches for luminal and c-Kit+ subtypes.
Publication Year:
2023
PubMed ID:
36653787
Public Summary:
Luminal cells are thought to be the cells of origin for several breast cancer subtypes and as a result are the subject of intense interest among breast cancer researchers. In the past it has been difficult to study these cells because they represent a rare population among cells grown in a lab dish. To tackle this issue, Todhunter et al. tested a new way of growing these cells called "postconfluent culture." Instead of frequently splitting dense cultures into new dishes, the authors let them grow for 2–5 weeks without any disturbance save regular media changes. This technique greatly increased the proportion of luminal cells in culture and created an environment more like the human breast. As such, postconfluent culture is a valuable tool for breast cancer researchers wanting to understand and intervene in early stages of breast cancer initiation.
Scientific Abstract:
BACKGROUND: A challenge in human mammary epithelial cell (HMEC) culture is sustaining the representation of competing luminal, myoepithelial, and progenitor lineages over time. As cells replicate in culture, myoepithelial cells come to dominate the composition of the culture with serial passaging. This drift in composition presents a challenge for studying luminal and progenitor cells, which are prospective cells of origin for most breast cancer subtypes. METHODS: We demonstrate the use of postconfluent culture on HMECs. Postconfluent culture entails culturing HMECs for 2-5 weeks without passaging but maintaining frequent feedings in low-stress M87A culture medium. In contrast, standard HMEC culture entails enzymatic subculturing every 3-5 days to maintain subconfluent density. RESULTS: When compared to standard HMEC culture, postconfluent culture yields increased proportions of luminal cells and c-Kit+ progenitor cells. Postconfluent cultures develop a distinct multilayered morphology with individual cells showing decreased physical deformability as compared to cells in standard culture. Gene expression analysis of postconfluent cells shows increased expression of lineage-specific markers and extracellular matrix components. CONCLUSIONS: Postconfluent culture is a novel, useful strategy for altering the lineage composition of HMECs, by increasing the proportional representation of luminal and progenitor cells. We speculate that postconfluent culture creates a microenvironment with cellular composition closer to the physiological state and eases the isolation of scarce cell subtypes. As such, postconfluent culture is a valuable tool for researchers using HMECs for breast cancer research.