Directed differentiation to nephron progenitor cells [RNA Seq 1]

Metabolic pathways driving differentiation into nephron progenitor cells (NPCs) and renal organoids from the pluripotency stage remain poorly understood. In this study, we systematically performed comprehensive metabolite and transcriptome profiling of human pluripotent stem cells (hPSCs) during differentiation into NPCs and further into multicellular organoids. We found activation of distinct metabolic pathways during early-stage progression from hPSCs to NPCs; however, later-stage differentiation from NPCs to organoids, and the intervening developmental stages between them, largely shared similar metabolic profiles. Among the pathways changing in early differentiation, the alanine-aspartate-glutamate and glutamine pathways were found to be significantly altered by both an enrichment and by pathway impact analysis. Moreover, hPSCs survived glutamine deprivation during in vitro differentiation, and NPCs were successfully generated both in the absence and presence of glutamine and glutamate. Surprisingly, glutamine deprivation resulted in enhanced maturation, by accelerating PAX8 expression, and enriching for podocytes as detected through single cell RNA-Seq analysis. Taken together, these findings highlight a critical regulatory role of glutamine metabolism in the derivation of nephron progenitor cells and renal organoids and identify a controlling metabolic pathway of early renal differentiation. Overall design: Global gene profiling on days 0, 1, 3, 4, 6, 7 and 9 during directed differentiation to nephron progenitor cells