Developing and utilising a mouse mammary organoid model

The mammary gland is a complex organ, relying upon multiple cell types and signalling pathways to orchestrate its predominantly postnatal development. To obtain a detailed understanding of such development, and also the mechanisms whose deregulation lead to mammary tumourigenesis, a physiologically relevant, quantifiable and qualitative model system is required. In vitro, currently available mammary culture systems have allowed studies into prospective stem cell populations, cell-cell interactions, differentiation, proliferation, paracrine networks and hormonal responses, but are limited in the degree to which they can truly recapitulate in vivo mammary biology. Three dimensional organoid culture systems from many tissues have recently been shown to concurrently allow sustained stem-cell maintenance, proliferation and functional differentiation; however, no such recapitulative in vitro model currently exists for the mammary gland. This thesis therefore details the development of a novel murine mammary organoid culture system. Culture conditions described, including R-Spondin1 and Neuregulin1, enable the development and expansion of mammary epithelial organoids for up to 2.5 months in culture. Organoids possess distinct basal and luminal compartments - functional steroid receptors in the latter allowing hormonal responses - and regenerative cell populations enabling mammary gland reconstitution in vivo. Additional conditions are also described promoting the growth of abnormal, tumour-like organoids. Utilisation of the organoid model allowed the study of key signalling pathways associated with mammary development; most notably revealing the critical importance of the Wnt signalling pathway in regulating normal development, its role in mammary tumourigenesis and demonstrating its potential as a target in anti-tumour therapy. Cell-cell signalling, individual cell populations and behaviours, and tumour organoid growth were studied, the latter offering a platform in which to study potential therapeutic compounds. Evidence provided furthers the understanding of mammary biology and supports that the organoid model comprises a suitable physiologically relevant tool for use in mammary research and drug discovery