Generation of Distal Renal Segments Involves a Unique Population of Aqp2+ Progenitor Cells

Significance Statement Renal progenitor cells could become critical for regenerative medicine and reveal mechanisms of pathology. Understanding their role in development may help kidney organoid generation. The identity of the progenitors of the distal renal segments, however, remains mysterious. We identify a unique subset of Aqp2(+) cells through Aqp2Cre- and Aqp2(ECE/+)-based lineage tracing in mice as the progenitors. Unlike regular principal cells and intercalated cells, the progenitors coexpress Aqp2 and V-ATPase subunits B1 and B2, and they are clonogenic, multipotent, and self-renew, generating DCT2, CNT1, CNT2, and CD cells. We demonstrate (1) high fidelity of both models; (2) the common origin and molecular identity of DCT2, CNT1, CNT2, and CD segments; and (3) a unique population of Aqp2(+) cells functioning as progenitors. Background Progenitor cells have clonogenicity, self-renewal, and multipotential capacity, and they can generate multiple types of cells during development. Evidence demonstrating the existence of such progenitor cells for renal distal segments is lacking. Methods To identify Aqp2(+) progenitor (AP) cells, we performed in vivo lineage tracing using both constitutive (Aqp2Cre RFP/+) and Tamoxifen-inducible (Aqp2(ECE/+) RFP/+, Aqp2(ECE/+) Brainbow/+, and Aqp2(ECE/+) Brainbow/Brainbow) mouse models. Aqp2Cre RFP/+ mice were analyzed from E14.5 to adult stage. The inducible models were induced at P1 and examined at P3 and P42, respectively. Multiple segment- or cell-specific markers were used for high-resolution immunofluorescence confocal microscopy analyses to identify the cell types derived from Aqp2(+) cells. Results Both Aqp2Cre and Aqp2(ECE/+) faithfully indicate the activation of the endogenous Aqp2 promoter for lineage tracing. A subset of Aqp2(+) cells behaves as potential AP. Aqp2Cre-based lineage tracing revealed that embryonic APs generate five types of cells, which form the late distal convoluted tubule (DCT2), connecting tubule segments 1 and 2 (CNT1 and CNT2, respectively), and collecting ducts (CDs). The ?- and ?-intercalated cells were apparently derived from embryonic AP in a stepwise manner. Aqp2(ECE/+)-based lineage tracing identified cells coexpressing Aqp2 and V-ATPase subunits B1 and B2 as the potential AP. Neonate APs generate daughter cells either inheriting their property (self-renewal) or evolving into various DCT2, CNT, or CD cells (multipotentiality), forming single cell-derived multiple-cell clones (clonogenicity) during development. Conclusion Our study demonstrates that unique Aqp2(+) B1B2(+) cells are the potential APs to generate DCT2, CNT, CNT2, and CD segments.

Automated summary

A unique subset of Aqp2(+) cells has been identified as potential progenitor cells for distal renal segments, demonstrating self-renewal, multipotentiality, and clonogenicity abilities to generate various cell types.