Direct Isolation and Characterization of Human Nephron Progenitors.

: Mature nephrons originate from a small population of uninduced nephrogenic progenitor cells (NPs) within the cap mesenchyme. These cells are characterized by the coexpression of SIX2 and CITED1. Many studies on mouse models as well as on human pluripotent stem cells have advanced our knowledge of NPs, but very little is known about this population in humans, since it is exhausted before birth and strategies for its direct isolation are still limited. Here we report an efficient protocol for direct isolation of human NPs without genetic manipulation or stepwise induction procedures. With the use of RNA-labeling probes, we isolated SIX2(+)CITED1(+) cells from human fetal kidney for the first time. We confirmed their nephrogenic state by gene profiling and evaluated their nephrogenic capabilities in giving rise to mature renal cells. We also evaluated the ability to culture these cells without complete loss of SIX2 and CITED1 expression over time. In addition to defining the gene profile of human NPs, this in vitro system facilitates studies of human renal development and provides a novel tool for renal regeneration and bioengineering purposes.

SIGNIFICANCE: The use of RNA-labeling probes allowed for the first time the definition of an efficient protocol for direct isolation of human nephron progenitors coexpressing SIX2 and CITED1, the master genes regulating renal development. These SIX2/CITED1-positive cells were derived from fetal kidneys without the use of any reprogramming strategy or laborious stepwise induction protocols. Their nephrogenic state was confirmed by gene profiling, and their nephrogenic specification and culture conditions were evaluated. This first "snapshot" of the transcriptional network of human nephron progenitors opens new avenues in understanding human kidney development and nephron specification and supports the study's ultimate goal of understanding possible mechanisms for kidney regeneration.