Production of kidney organoids arranged around single ureteric bud trees, and containing endogenous blood vessels, solely from embryonic stem cells.

There is intense worldwide effort in generating kidney organoids from pluripotent stem cells, for research, for disease modelling and, perhaps, for making transplantable organs. Organoids generated from pluripotent stem cells (PSC) possess accurate micro-anatomy, but they lack higher-organization. This is a problem, especially for transplantation, as such organoids will not be able to perform their physiological functions. In this study, we develop a method for generating murine kidney organoids with improved higher-order structure, through stages using chimaeras of ex-fetu and PSC-derived cells to a system that works entirely from embryonic stem cells. These organoids have nephrons organised around a single ureteric bud tree and also make vessels, with the endothelial network approaching podocytes.

Long-term live imaging and multiscale analysis identify heterogeneity and core principles of epithelial organoid morphogenesis.

BACKGROUND: Organoids are morphologically heterogeneous three-dimensional cell culture systems and serve as an ideal model for understanding the principles of collective cell behaviour in mammalian organs during development, homeostasis, regeneration, and pathogenesis. To investigate the underlying cell organisation principles of organoids, we imaged hundreds of pancreas and cholangiocarcinoma organoids in parallel using light sheet and bright-field microscopy for up to 7 days. RESULTS: We quantified organoid behaviour at single-cell (microscale), individual-organoid (mesoscale), and entire-culture (macroscale) levels. At single-cell resolution, we monitored formation, monolayer polarisation, and degeneration and identified diverse behaviours, including lumen expansion and decline (size oscillation), migration, rotation, and multi-organoid fusion. Detailed individual organoid quantifications lead to a mechanical 3D agent-based model. A derived scaling law and simulations support the hypotheses that size oscillations depend on organoid properties and cell division dynamics, which is confirmed by bright-field microscopy analysis of entire cultures. CONCLUSION: Our multiscale analysis provides a systematic picture of the diversity of cell organisation in organoids by identifying and quantifying the core regulatory principles of organoid morphogenesis.

Sfrp3 modulates stromal-epithelial crosstalk during mammary gland development by regulating Wnt levels.

Mammary stroma is essential for epithelial morphogenesis and development. Indeed, postnatal mammary gland (MG) development is controlled locally by the repetitive and bi-directional cross-talk between the epithelial and the stromal compartment. However, the signalling pathways involved in stromal-epithelial communication are not entirely understood. Here, we identify Sfrp3 as a mediator of the stromal-epithelial communication that is required for normal mouse MG development. Using Drosophila wing imaginal disc, we demonstrate that Sfrp3 functions as an extracellular transporter of Wnts that facilitates their diffusion, and thus, their levels in the boundaries of different compartments. Indeed, loss of Sfrp3 in mice leads to an increase of ductal invasion and branching mirroring an early pregnancy state. Finally, we observe that loss of Sfrp3 predisposes for invasive breast cancer. Altogether, our study shows that Sfrp3 controls MG morphogenesis by modulating the stromal-epithelial cross-talk during pubertal development.

Macrophages restrict the nephrogenic field and promote endothelial connections during kidney development.

The origins and functions of kidney macrophages in the adult have been explored, but their roles during development remain largely unknown. Here we characterise macrophage arrival, localisation, heterogeneity, and functions during kidney organogenesis. Using genetic approaches to ablate macrophages, we identify a role for macrophages in nephron progenitor cell clearance as mouse kidney development begins. Throughout renal organogenesis, most kidney macrophages are perivascular and express F4/80 and CD206. These macrophages are enriched for mRNAs linked to developmental processes, such as blood vessel morphogenesis. Using antibody-mediated macrophage-depletion, we show macrophages support vascular anastomoses in cultured kidney explants. We also characterise a subpopulation of galectin-3+ (Gal3+) myeloid cells within the developing kidney. Our findings may stimulate research into macrophage-based therapies for renal developmental abnormalities and have implications for the generation of bioengineered kidney tissues.