In a previous Cell of The Month article, we explored the biology of the kidney. We highlighted the structure and function of the glomerulus podocytes and proximal tubules; these are specialized cell types found in the nephron, which is the kidney’s key structural and functional unit. We also touched upon some of the ways in which podocytes and proximal tubular epithelial cells can contribute to kidney disease, including inherited or acquired podocyte injury that leads to reduced filtration barrier function, and injury to the proximal tubules that is increasingly believed to play a key role in the progression from acute to chronic kidney disease, as well as most cases of renal cell carcinoma.
Here, we look at how podocytes and proximal tubules contribute to kidney fibrosis, which is a key end stage in most progressive kidney diseases, and explore their importance in drug-induced nephrotoxicity assessment. After a quick recap on kidney biology, we introduce kidney fibrosis and nephrotoxicity, providing a brief overview of what is known about how podocytes and proximal tubular cells respond to injury and their potential roles in kidney fibrosis. We also highlight how these specialized cell types serve as critical models for evaluating potential kidney damage during drug development.
A quick recap on kidney structure and function
Kidneys play critical roles in toxin removal, electrolyte homeostasis, acid-base regulation, and the secretion of hormones including erythropoietin and renin, which control red blood cell production and blood pressure.
Each human kidney contains approximately 2 million nephrons; each nephron consists of a glomerulus for filtration and a tubular reabsorption compartment. Podocytes, which are specialized terminally differentiated cells within the glomerulus, form foot processes that create filtration slits at the glomerular basement membrane. This arrangement allows selective filtration, retaining blood cells and proteins while allowing small solutes to pass through. The proximal tubule, distinguished by its brush border and mitochondria-rich epithelial cells, reabsorbs most filtered substances through energy-dependent transport mechanisms powered by sodium gradients, with water following passively along concentration pathways.
This complex filtration and reabsorption system makes the kidney susceptible to nephrotoxicity, which can affect the entire kidney and be fatal in the most severe cases. Nephrotoxicity occurs because of the high metabolic activity of proximal tubular cells and the concentration of substances during filtration; both can cause cellular damage when exposed to certain chemicals or medications. Advanced cellular models of podocytes and proximal tubule cells provide an opportunity to study toxicity during early-stage drug development, enabling more human relevant nephrotoxicity assessments than previously possible.
Kidney fibrosis: The pathological endpoint of chronic kidney disease
Kidney fibrosis affects approximately 10-14% of the global population, and represents the common pathological endpoint of chronic kidney disease (CKD), which is an escalating threat to global public health (1,2 and references within).
CKD is clinically diagnosed when abnormalities in kidney structure or function persist for more than three months and impact health. Widely used diagnostic criteria include reduced estimated glomerular filtration rate or elevated urinary albumin-to-creatinine ratio, both of which suggest sub-optimal kidney function.
Kidney fibrosis develops when normal wound healing becomes dysregulated, leading to excessive accumulation of extracellular matrix (ECM) proteins including fibronectin and collagens. Its morphological characteristics include glomerulosclerosis, tubular atrophy, interstitial inflammation, and vascular rarefaction, leading to severe inflammation as a result.
The process begins with injury-triggered activation of fibroblasts and pericytes, a multi-functional cell type whose role in immune modulation is still being characterized (3). This causes those cells to increase their contractility, release inflammatory mediators, and synthesize ECM [extracellular matrix] components. When injury persists or is severe, continuous ECM accumulation leads to progressive tissue disruption, functional decline, and eventually, organ failure. Despite advances in understanding kidney fibrosis, translating this knowledge into new therapies remains challenging.
The fibrotic niche
The findings of studies carried out within the past decade support the idea that organ fibrosis starts from a fibrotic niche. This may be defined as the complex interplay between the injured parenchyma (functional tissue) and multiple non-parenchymal cell types that are located near the scarred tissue (4, and references within). A kidney spatial transcriptomic analysis published in Nature in 2021 revealed mesenchymal cells, immune cells, and specific types of kidney tubular epithelial cells as the cellular components of the fibrotic niche within the human kidney. The identification of these cell types, along with pathways that are upregulated in response to tissue injury (e.g., Notch, Wnt, Hedgehog and SOX9) and factors expressed during fibrosis, represents progress in the field, but the precise mechanisms by which these cells interact to mediate fibrosis are not fully understood (2, 4).
While kidney fibrosis involves multiple cell types in a complex interplay, podocytes and proximal tubule cells are attractive therapeutic targets because of their distinct vulnerabilities to injury, central roles in initiating and driving fibrotic cascades, and their potential as valuable cellular models for drug development using human iPSC-derived kidney cell types, both to understand disease mechanisms and develop targeted therapies.
So, how do proximal tubules and podocytes contribute to kidney fibrosis?
Proximal tubules
Single-cell (sc)RNA-seq studies during the last decade have provided new insights into the vulnerability and response of proximal tubular epithelial cells (PTCs) following injury. These cells are highly susceptible to acute damage, with studies revealing significant decreases in PTC abundance after acute kidney injury (AKI).
Distinct injury-related PTC sub-populations have also been identified; these are reported to express markers such as VCAM1, HAVCR1, PROM1, or DCDC2—all of which have been implicated in liver fibrosis or other liver disease—during the transition from AKI to chronic kidney disease in both murine and human kidneys.
Of note, PTCs that express VCAM1—which is known to regulate inflammation-associated vascular adhesion—expand after injury or with aging and are detectable in the urine of diabetic kidney disease patients. These cells lose their terminal differentiation markers while developing a pro-inflammatory phenotype and have been termed ‘failed repair PTCs’ due to their long-term persistence post-injury. Similar PTC sub-populations have been found within the thick ascending limb of the loop of Henle, often with expression of PROM1, HAVCR1, and DCDC2.
Although definitions for these PTC sub-populations vary between studies, their marker profiles are similar with a pro-fibrotic phenotype. These cells also express some mesenchymal markers and have been suggested to overlap with cells previously described as undergoing partial epithelial-mesenchymal transition, which is a crucial step in fibrosis.
Several studies have explored how pro-fibrotic PTCs contribute to kidney fibrosis through various cellular crosstalk mechanisms. Through receptor-ligand interaction analyses, researchers found that pro-fibrotic PTCs secrete signaling molecules including Edn1, Ccl2, Wnt, Notch, TNF, and TGF-β that can directly or indirectly activate mesenchymal cells by affecting circulating hemopoietic cells.
It has also been demonstrated that pro-fibrotic PTCs release the chemoattractant CXCL1 to recruit CXCR2-positive basophils, which then promote fibrosis through interactions with Th17 helper T cells. Additionally, active cell death of pro-fibrotic PTCs through pyroptosis or ferroptosis has been shown to drive immune cell activation and fibrosis. These findings in the last decade highlight the complex interplay between injured and pro-fibrotic PTCs, mesenchymal cells, and immune cells in driving kidney remodeling and fibrosis and suggests that therapies to target tubule dedifferentiation, injury, or cellular crosstalk might be promising strategies to slow kidney fibrosis progression.
Much of what we now understand about pro-fibrotic PTCs and their role in kidney fibrosis has been gleaned through single-cell genomics approaches (5 and references within). Future studies incorporating human kidney biopsies will further elucidate the mechanisms behind fibrosis and address critical questions, such as whether these mechanisms are conserved across different kidney diseases or if there are disease-specific differences in how PTCs drive fibrosis. While single-cell analyses provide valuable clues about cellular interactions and pathways, follow-up studies that include targeted disruption of identified pathways and interactions are necessary to confirm their functional significance and potential as therapeutic targets.
Podocytes
While our understanding of how proximal tubular cells in kidney fibrosis has greatly improved in the last decade, less is known about the contribution of podocytes to this process. What is known (summarised in Ref. 7) suggests that podocytes primarily influence kidney fibrosis through their injury and dysfunction in glomerular diseases.
For example, in a study of diabetic kidney disease, podocytes exhibited altered expression patterns in genes related to glucose and lipid metabolism, increased activity in apoptotic pathways, and enhanced production of matrix proteins, with progressive podocyte loss in advanced disease stages. Podocyte injury was also reported to be a key initiator of proteinuria and focal segmental glomerulosclerosis, which is a rare kidney disease in which the glomeruli are scarred and the podocytes are damaged. Podocytes are also known to engage in complex cellular crosstalk through specific receptor-ligand interactions with mesangial and endothelial cells, including CCN1-ITGAV/ITGB3/ITGB5 and SLIT3-ROBO2 signaling pathways, although the function and implications of these interactions and how they play into kidney fibrosis are not fully understood (7 and references within).
The importance of nephrotoxicity in drug discovery and development
As mentioned above, advanced cellular models of podocytes and proximal tubule cells enable more accurate nephrotoxicity assessments than before. Kidney injury is a major cause of drug failure during clinical development, making reliable preclinical nephrotoxicity screening a crucial part of drug development.
Podocytes and proximal tubule cells are both vulnerable to drug-induced injury albeit in different ways; proximal tubules express critical drug transporters (OATs, OCTs, P-glycoprotein) while podocytes contain filtration barrier proteins whose disruption causes proteinuria. Incorporating both cell types in screening enables earlier identification of nephrotoxicity, which should help to reduce drug failure in clinical trials and improve patient safety.
Tempo-iKidneyPod represents a new model for nephrotoxicity evaluation, utilizing iPSC-derived kidney cells that maintain key functional characteristics of mature podocytes. These cells enable real-time qPCR analysis of various injury biomarkers and the penetration of molecules (e.g. drugs) can be studied through live-cell imaging assays. Moreover, iKidneyPod cells can be adapted for mitochondrial toxicity assessments and ATP depletion assays, providing further insights into potential nephrotoxicity mechanisms.
As our understanding of kidney fibrosis continues to advance, these cellular models will not only serve as tools for nephrotoxicity assessment but also help to pave the way for new, targeted therapies for kidney fibrosis.
References:
- Kidney disease: a global health priority. Nat Rev Nephrol. 2024 Jul;20(7):421-423.
- Huang R, Fu P, Ma L. Kidney fibrosis: from mechanisms to therapeutic medicines. Signal Transduct Target Ther. 2023 Mar 17;8(1):129.
- Dabravolski SA, Andreeva ER, Eremin II, et al. The Role of Pericytes in Regulation of Innate and Adaptive Immunity. Biomedicines. 2023 Feb 17;11(2):600.
- Kuppe C, Ibrahim MM, Kranz J, et al. Decoding myofibroblast origins in human kidney fibrosis. Nature. 2021 Jan;589(7841):281-286.
- Yamashita N, Kramann R. Mechanisms of kidney fibrosis and routes towards therapy. Trends Endocrinol Metab. 2024 Jan;35(1):31-48.
- Lake BB, Menon R, Winfree S, et al. An atlas of healthy and injured cell states and niches in the human kidney. Nature. 2023 Jul;619(7970):585-594.
- Huang, R., Fu, P. & Ma, L. Kidney fibrosis: from mechanisms to therapeutic medicines. Sig Transduct Target Ther 2023, 8:129.
Karen O’Hanlon Cohrt is an independent Science Writer with a PhD in biotechnology from Maynooth University, Ireland (2011). After her PhD, Karen relocated to Denmark where she held postdoctoral positions in mycology and later in human cell cycle regulation, before moving to the world of drug discovery. Karen has been a full-time science writer since 2017, and has since then held numerous contract roles in science communication and editing spanning diverse topics including diagnostics, molecular biology, and gene therapy. Her broad research background provides the technical know-how to support scientists in diverse areas, and this in combination with her passion for learning helps her to keep abreast of exciting research developments as they unfold. Karen is currently based in Ireland, and you can follow her on Linkedin here.
