Niche-inspired tubuloid differentiation enables modeling of the distal nephron in health and disease

Tubuloid culture allows long-term expansion of primary human renal epithelium. However, the progenitor state in expanding tubuloids comes at expense of differentiation, which limits application. To improve differentiation, we developed niche-inspired culture conditions. A single cell atlas revealed that differentiation predominantly generated ascending limb of Henle and principal cells. Using differentiated tubuloids, we subsequently modeled lithium nephropathy in vitro, and performed single-cell RNA sequencing to look at the transcriptional effects caused by lithium. We found that lithium suppresses electrolyte and solute transporters in multiple cell types and induces a progenitor state in kidney tubuloids. Overall design: Kidney tubuloids were grown for 3 days post-dissociation in expansion medium (EM) and then treated for 7 days with EM or differentiation medium (FAF). Samples were thereafter harvested for sequencing. For sequencing of lithium samples, tubuloids were grown to a confluent 2D monolayers in EM and then treated for 7 days with FAF + 10 mM sodium (Na) or lithium (Li).