A Comprehensive Molecular Portrait of Human Urine-derived Renal Progenitor Cells

Abstract

Human urine is a non-invasive source of stem cells with regeneration potential. Here, we investigated the cellular and molecular identities, and the gene regulation driving self-renewal and differentiation of these cells in vitro. These cells express pluripotency-associated markers enabling easy reprogramming. Based on the expression of renal associated genes, proteins and functionality, we refer to these cells as urine derived renal progenitor cells-UdRPCs. CHIR99021-induced differentiation of UdRPCs activated WNT-related genes-AXIN2, JUN and NKD1. Protein interaction network identified JUN as a putative regulator of differentiation whereas self-renewal is maintained by FGF2-driven TGFβ-SMAD2/3. Our data will enhance understanding of the molecular identities of UdRPCs, and enable the generation of renal disease models in vitro and eventually kidney-associated regenerative therapies.3 Abstract Background Human urine is now recognised as a non-invasive source of stem cells with regeneration potential. These cells are mesenchymal stem cells but their detailed molecular and cellular identities are poorly defined. Furthermore, unlike the mouse, the gene regulatory network driving self-renewal and differentiation into functional renal cells in vitro remain unresolved. MethodsWe isolated urine stem cells from 10 individuals from both genders and distinct ages, characterized them as renal progenitor cells and explored the gene regulatory network sustaining self-renewal. ResultsThese cells express pluripotency-associated proteins-TRA-1-60, TRA-1-81, SSEA4, C-KIT and CD133. Expression of pluripotency-associated proteins enabled rapid reprogramming into iPSCs using episomal-based plasmids without pathway perturbations. Transcriptome analysis revealed expression of a plethora of nephrogenesis-related genes such as SIX2, OSR1, CITED1, NPHS2, NPHS1, PAX2, SALL1, AQP2, EYA1, SLC12A1 and UMOD. As expected, the cells transport Albumin by endocytosis. Based on this, we refer to these cells as urine derived renal progenitor cells-UdRPCs. Associated GO-term analysis of UdRPCs and UdRPC-iPSCs underlined their renal identity and functionality. Upon differentiation by WNT activation using the GSK3β-inhibitor (CHIR99021), transcriptome and KEGG pathway analysis revealed upregulation of WNT-associated genes-AXIN2, JUN and NKD1. Protein interaction network identified JUN-a downstream target of the WNT pathway in association with STAT3, ATF2 and MAPK1 as a putative regulator of self-renewal and differentiation in UdRPCs. Furthermore, like pluripotent stem cells, self-renewal is maintained by FGF2-driven TGFβ-SMAD2/3 pathway.Conclusion This in vitro model and the data presented should lay the foundation for studying nephrogenesis in man.

Urinary single-cell sequencing captures kidney injury and repair processes in human acute kidney injury

“…The feasibility of urinary single-cell RNA sequencing (scRNAseq) has been proven in chronic kidney diseases such as diabetic nephropathy (DN) and focal-segmental glomerulosclerosis (FSGS) [18][19][20] and indicates that the urine has untapped potential as a source for the non-invasive study of renal epithelial cells. Moreover, urine-derived progenitor or stem cells, which were first isolated in pediatric patients 21 , have since been studied extensively 22 .…”

Cell competition for cancer treatment with Urine Progenitor Cells

“…Once we ascertained that the UPC's antitumor response depends on direct cell-to-cell contacts, we wanted to identify genes expressed in UPCs capable of triggering cell competition responses towards three main known mechanisms: apoptosis, phagocytosis, and extrusion. To this end, we evaluated the expression of genes involved in cell competition mechanisms using publicly available differential gene expression datasets, where the gene expression profiles of the UPCs were compared to MSCs and fibroblasts [20][21][22] . Interestingly, we found that UPCs overexpress three genes related to the antitumor response through cell competition.…”

Renal progenitors derived from urine for personalized diagnosis of kidney diseases

“…Specifically, they demonstrated that RPC are the only urinary cells that can undergo numerous rounds of cell division in culture, exhibiting clonogenic and amplification potential over time [36]. Transcriptomic profile analysis proved that urine-derived RPC (u-RPC) are the same as those residing in the kidney in terms of phenotypic and functional markers [36,45]. More recently, single-cell RNA sequencing (scRNAseq) analysis confirmed the presence of a population of RPC in urine samples endowed with bipotency, giving rise potentially to both podocytes and tubular cells [46].…”