Generation and functional characterization of murine mammary organoids

Abstract

Summary 3D cultures of mammary epithelial cells purified from murine models provide a unique resource to study genetically defined breast cancer and response to targeted therapies. Here, we describe step-by-step experimental procedures for the successful establishment of murine mammary organoid lines isolated from mammary glands or mammary tumors driven by mutations in components of the PI3K pathway. These detailed protocols also include procedures to perform assays that can be adopted to screen response to drug treatments and to inform better therapies. For details on potential applications and use of this protocol, please refer to Yip et al. (2020) .

Effects of 5-fluorouracil, thymoquinone and mammary stem cells’ exosomes on in vitro cultured breast cancer cells

“…The bovine mammary stem cells (Bov-MaSCs) were isolated and characterized from the udder of lactating dairy cows, this was performed following our previously published protocol ( Abdelfattah-Hassan et al ., 2016 ). The establishment of mammary organoids using isolated Bov-MaSCs via culturing the isolated cells on ultra-low attachment plates (Corning, USA) as described previously with slight modifications ( Yip and Papa, 2021 ).…”

In vitro breast cancer models for studying mechanisms of resistance to endocrine therapy

“…A recent publication detailed the methodology for developing murine mammary organoids from both normal and tumor tissue [ 72 ]. This advance will encourage investigators to establish more elaborate genetically-engineered mouse models (GEMMs) that mirror the combinations of gene mutations observed in human tumors.…”

Methods for imaging intracellular calcium signals in the mouse mammary epithelium in 2- and 3-dimensions

“…These two methods provide scope for imaging different stages of the mature mammary gland. Of note, many published mammary dissection protocols describe methods that ensure optimal dissection of the abdominal (4 th ) mammary glands, but not the inguinal (5 th ) glands [33–35]. In our experience, the pair of inguinal mammary glands provide valuable tissue for both ex vivo imaging and primary cell isolation due to the high ratio of mammary epithelial cells in comparison to surrounding stroma.…”

“…Whilst outside the scope of this chapter, following isolation, primary epithelial cells can additionally be utilised for the development of mammary organoids, which our group has shown are also suitable for Ca 2+ imaging studies. For further information on this, we recommend previous descriptions of this process [18, 35].…”