In vitro differentiation of mouse pluripotent stem cells into corticosteroid-producing adrenocortical cells

Directed differentiation of pluripotent stem cells into specialized cell types represents an invaluable tool for a wide range of applications. Here, we have exploited single-cell transcriptomic data to develop a stepwise in vitro differentiation system from mouse embryonic stem cells into adrenocortical cells. We show that during development, the adrenal primordium is embedded in an extracellular matrix containing tenascin and fibronectin. Culturing cells on fibronectin during differentiation increased the expression of the steroidogenic marker NR5A1. Furthermore, 3D cultures in the presence of protein kinase A (PKA)-pathway activators led to the formation of aggregates composed of different cell types expressing adrenal progenitor or steroidogenic markers, including the adrenocortical-specific enzyme CYP21A1. Importantly, in-vitro-differentiated cells responded to adrenocorticotropic hormone (ACTH) and angiotensin II with the production of glucocorticoids and mineralocorticoids, respectively, thus confirming the specificity of differentiation toward the adrenal lineage. [Display omitted] •Mouse adrenal progenitors differentiate in a tenascin/fibronectin-rich ECM•Stepwise differentiation of embryonic stem cells yields NR5A1+ adrenocortical cells•3D aggregates produce glucocorticoids in response to ACTH•Treatment with angiotensin II induces aldosterone production Cellular systems that allow studying adrenocortical biology are sparse. Here, Neirijnck and colleagues have leveraged single-cell transcriptomic data to develop a protocol for the directed differentiation of mouse ES cells into cellular aggregates that express key markers of the adrenal cortex and respond to ACTH and angiotensin II by secreting glucocorticoids and mineralocorticoids, respectively.