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A comprehensive cellular anatomy of normal human kidney is crucial to address the cellular origins of renal disease and renal cancer. Some kidney diseases may be cell type-specific, especially renal tubular cells. To investigate the classification and transcriptomic information of the human kidney, we rapidly obtained a single-cell suspension of the kidney and conducted single-cell RNA sequencing (scRNA-seq). Here, we present the scRNA-seq data of 23,366 high-quality cells from the kidneys of three human donors. In this dataset, we show 10 clusters of normal human renal cells. Due to the high quality of single-cell transcriptomic information, proximal tubule (PT) cells were classified into three subtypes and collecting ducts cells into two subtypes. Collectively, our data provide a reliable reference for studies on renal cell biology and kidney disease.

Tubuloids have become a promising tool for modeling and regenerating kidney disease, although their ability for integration and regeneration in vivo is not well documented. Here, we established, characterized, and compared human tubuloids using two optimized protocols: one involving prior isolation of tubular cells (Crude tubuloids) and the other involving prior isolation of proximal tubular cells (F4 tubuloids). Next, healthy rat-derived tubuloids were established using this protocol. Finally, we compared two strategies for delivering GFP tubuloids to a kidney host: 1) subcapsular/intracortical injection and 2) tubuloid infusion during normothermic preservation in a rat transplantation model and a discarded human kidney. F4 tubuloids achieved a higher level of differentiation state compared to Crude tubuloids. When analyzing tubuloid delivery to the kidney, normothermic perfusion was found to be more efficient than in vivo injection. Moreover, fully developed tubules were observed in the host parenchyma at 1 week and 1 month after infusion during normothermic perfusion represent a potential strategy to enhance the translatability of kidney regenerative therapies into clinical practice to condition kidney grafts and to treat kidney diseases.

MicroRNAs, activated by the enzyme Dicer1, control post-transcriptional gene expression. Dicer1 has important roles in the epithelium during nephrogenesis, but its function in stromal cells during kidney development is unknown. To study this, we inactivated Dicer1 in renal stromal cells. This resulted in hypoplastic kidneys, abnormal differentiation of the nephron tubule and vasculature, and perinatal mortality. In mutant kidneys, genes involved in stromal cell migration and activation were suppressed as were those involved in epithelial and endothelial differentiation and maturation. Consistently, polarity of the proximal tubule was incorrect, distal tubule differentiation was diminished, and elongation of Henle’s loop attenuated resulting in lack of inner medulla and papilla in stroma-specific Dicer1 mutants. Glomerular maturation and capillary loop formation were abnormal, whereas peritubular capillaries, with enhanced branching and increased diameter, formed later. In Dicer1-null renal stromal cells, expression of factors associated with migration, proliferation, and morphogenic functions including α-smooth muscle actin, integrin-α8, -β1, and the WNT pathway transcriptional regulator LEF1 were reduced. Dicer1 mutation in stroma led to loss of expression of distinct microRNAs. Of these, miR-214, -199a-5p, and -199a-3p regulate stromal cell functions ex vivo, including WNT pathway activation, migration, and proliferation. Thus, Dicer1 activity in the renal stromal compartment regulates critical stromal cell functions that, in turn, regulate differentiation of the nephron and vasculature during nephrogenesis.

To understand renal functions and disease, it is important to define the molecular constituents of the various compartments of the kidney. Here, we used comparative transcriptomic analysis of all major organs and tissues in the human body, in combination with kidney tissue micro array based immunohistochemistry, to generate a comprehensive description of the kidney-specific transcriptome and proteome. A special emphasis was placed on the identification of genes and proteins that were elevated in specific kidney subcompartments. Our analysis identified close to 400 genes that had elevated expression in the kidney, as compared to the other analysed tissues, and these were further subdivided, depending on expression levels, into tissue enriched, group enriched or tissue enhanced. Immunohistochemistry allowed us to identify proteins with distinct localisation to the glomeruli (n = 11), proximal tubules (n = 120), distal tubules (n = 9) or collecting ducts (n = 8). Among the identified kidney elevated transcripts, we found several proteins not previously characterised or identified as elevated in kidney. This description of the kidney specific transcriptome and proteome provides a resource for basic and clinical research to facilitate studies to understand kidney biology and disease.

Current kidney organoids model development and diseases of the nephron but not the contiguous epithelial network of the kidney’s collecting duct (CD) system. Here, we report the generation of an expandable, 3D branching ureteric bud (UB) organoid culture model that can be derived from primary UB progenitors from mouse and human fetal kidneys, or generated de novo from human pluripotent stem cells. In chemically-defined culture conditions, UB organoids generate CD organoids, with differentiated principal and intercalated cells adopting spatial assemblies reflective of the adult kidney’s collecting system. Aggregating 3D-cultured nephron progenitor cells with UB organoids in vitro results in a reiterative process of branching morphogenesis and nephron induction, similar to kidney development. Applying an efficient gene editing strategy to remove RET activity, we demonstrate genetically modified UB organoids can model congenital anomalies of kidney and urinary tract. Taken together, these platforms will facilitate an enhanced understanding of development, regeneration and diseases of the mammalian collecting duct system. Here, the authors model the collecting duct system in kidneys by taking ureteric bud (UB) progenitor cells from both mouse and human primary tissues, as well as from hESC and hiPSC to generate organoids, which can model congenital anomalies of the kidney and urinary tract.

The incidence of kidney disease is rising, constituting a significant burden on the healthcare system and making identification of new therapeutic targets increasingly urgent. The heme oxygenase (HO) system performs an important function in the regulation of oxidative stress and inflammation and, via these mechanisms, is thought to play a role in the prevention of non-specific injuries following acute renal failure or resulting from chronic kidney disease. The expression of HO-1 is strongly inducible by a wide range of stimuli in the kidney, consequent to the kidney’s filtration role which means HO-1 is exposed to a wide range of endogenous and exogenous molecules, and it has been shown to be protective in a variety of nephropathological animal models. Interestingly, the positive effect of HO-1 occurs in both hemolysis- and rhabdomyolysis-dominated diseases, where the kidney is extensively exposed to heme (a major HO-1 inducer), as well as in non-heme-dependent diseases such as hypertension, diabetic nephropathy or progression to end-stage renal disease. This highlights the complexity of HO-1’s functions, which is also illustrated by the fact that, despite the abundance of preclinical data, no drug targeting HO-1 has so far been translated into clinical use. The objective of this review is to assess current knowledge relating HO-1’s role in the kidney and its potential interest as a nephroprotection agent. The potential therapeutic openings will be presented, in particular through the identification of clinical trials targeting this enzyme or its products.

The (Pro)renin receptor (P)RR/Atp6ap2 is a cell surface protein capable of binding and non-proteolytically activate prorenin. Additionally, (P)RR is associated with H(+)-ATPases and alternative functions in H(+)-ATPase regulation as well as in Wnt signalling have been reported. Kidneys express very high levels of H(+)-ATPases which are involved in multiple functions such as endocytosis, membrane protein recycling as well as urinary acidification, bicarbonate reabsorption, and salt absorption. Here, we wanted to localize the (P)RR/Atp6ap2 along the murine nephron, exmaine whether the (P)RR/Atp6ap2 is coregulated with other H(+)-ATPase subunits, and whether acute stimulation of the (P)RR/Atp6ap2 with prorenin regulates H(+)-ATPase activity in intercalated cells in freshly isolated collecting ducts. We localized (P)PR/Atp6ap2 along the murine nephron by qPCR and immunohistochemistry. (P)RR/Atp6ap2 mRNA was detected in all nephron segments with highest levels in the collecting system coinciding with H(+)-ATPases. Further experiments demonstrated expression at the brush border membrane of proximal tubules and in all types of intercalated cells colocalizing with H(+)-ATPases. In mice treated with NH4Cl, NaHCO3, KHCO3, NaCl, or the mineralocorticoid DOCA for 7 days, (P)RR/Atp6ap2 and H(+)-ATPase subunits were regulated but not co-regulated at protein and mRNA levels. Immunolocalization in kidneys from control, NH4Cl or NaHCO3 treated mice demonstrated always colocalization of PRR/Atp6ap2 with H(+)-ATPase subunits at the brush border membrane of proximal tubules, the apical pole of type A intercalated cells, and at basolateral and/or apical membranes of non-type A intercalated cells. Microperfusion of isolated cortical collecting ducts and luminal application of prorenin did not acutely stimulate H(+)-ATPase activity. However, incubation of isolated collecting ducts with prorenin non-significantly increased ERK1/2 phosphorylation. Our results suggest that the PRR/Atp6ap2 may form a complex with H(+)-ATPases in proximal tubule and intercalated cells but that prorenin has no acute effect on H(+)-ATPase activity in intercalated cells.

Key Points The mTOR pathway has a central role in the regulation of cell metabolism, growth and proliferation Pharmacological inhibition of mTOR and selective gene targeting of mTORC1 or mTORC2 in podocytes and tubular epithelial cells has helped to elucidate their role in renal cell homeostasis, including in autophagy Mechanistic insights into the roles of mTOR complexes in regulating immune cell function are helping to improve understanding of the effects of mTOR inhibitors in renal disease and transplant rejection mTOR is increasingly recognized as having a fundamental role in the development of glomerular disease and in acute kidney injury; its role in fibrotic kidney disease is less certain New generation dual mTORC1 and mTORC2 inhibitors offer potential for the treatment of renal cell carcinoma mTOR inhibitors are associated with reduced rates of skin cancer and cytomegalovirus infection in renal transplant recipients The mTOR pathway has a role in the development of renal disease, kidney transplant rejection and malignancies. Here, the authors discuss the mechanisms by which mTOR complexes drive the pathogenesis of these diseases as well as the therapeutic potential of mTOR inhibitors. The mTOR pathway has a central role in the regulation of cell metabolism, growth and proliferation. Studies involving selective gene targeting of mTOR complexes (mTORC1 and mTORC2) in renal cell populations and/or pharmacologic mTOR inhibition have revealed important roles of mTOR in podocyte homeostasis and tubular transport. Important advances have also been made in understanding the role of mTOR in renal injury, polycystic kidney disease and glomerular diseases, including diabetic nephropathy. Novel insights into the roles of mTORC1 and mTORC2 in the regulation of immune cell homeostasis and function are helping to improve understanding of the complex effects of mTOR targeting on immune responses, including those that impact both de novo renal disease and renal allograft outcomes. Extensive experience in clinical renal transplantation has resulted in successful conversion of patients from calcineurin inhibitors to mTOR inhibitors at various times post-transplantation, with excellent long-term graft function. Widespread use of this practice has, however, been limited owing to mTOR-inhibitor- related toxicities. Unique attributes of mTOR inhibitors include reduced rates of squamous cell carcinoma and cytomegalovirus infection compared to other regimens. As understanding of the mechanisms by which mTORC1 and mTORC2 drive the pathogenesis of renal disease progresses, clinical studies of mTOR pathway targeting will enable testing of evolving hypotheses.

Chronic kidney disease is a leading cause of death in the United States. Tubulointerstitial fibrosis (TIF) is considered the final common pathway leading to end-stage renal disease (ESRD). Here, we used pharmacologic, genetic, in vivo, and in vitro experiments to show that activation of the Notch pathway in tubular epithelial cells (TECs) in patients and in mouse models of TIF plays a role in TIF development. Expression of Notch in renal TECs was found to be both necessary and sufficient for TIF development. Genetic deletion of the Notch pathway in TECs reduced renal fibrosis. Consistent with this, TEC-specific expression of active Notch1 caused rapid development of TIF. Pharmacologic inhibition of Notch activation using a γ-secretase inhibitor ameliorated TIF. In summary, our experiments establish that epithelial injury and Notch signaling play key roles in fibrosis development and indicate that Notch blockade may be a therapeutic strategy to reduce fibrosis and ESRD development.

In recent years, although the development of clinical therapy for diabetic kidney disease (DKD) has made great progress, the progression of DKD still cannot be controlled. Therefore, further study of the pathogenesis of DKD and improvements in DKD treatment are crucial for prognosis. Traditional studies have shown that podocyte injury plays an important role in this process. Recently, it has been found that glomerulotubular balance and tubuloglomerular feedback (TGF) may be involved in the progression of DKD. Glomerulotubular balance is the specific gravity absorption of the glomerular ultrafiltrate by the proximal tubules, which absorbs only 65% to 70% of the ultrafiltrate. This ensures that the urine volume will not change much regardless of whether the glomerular filtration rate (GFR) increases or decreases. TGF is one of the significant mechanisms of renal blood flow and self‐regulation of GFR, but how they participate in the development of DKD in the pathological state and the specific mechanism is not clear. Injury to tubular epithelial cells (TECs) is the key link in DKD. Additionally, injury to glomerular endothelial cells (GECs) plays a key role in the early occurrence and development of DKD. However, TECs and GECs are close to each other in anatomical position and can crosstalk with each other, which may affect the development of DKD. Therefore, the purpose of this review was to summarize the current knowledge on the crosstalk between TECs and GECs in the pathogenesis of DKD and to highlight specific clinical and potential therapeutic strategies.