In terms of cellular diversity, it may not be the brain, but the kidney boasts more cell types than most other organs. Now, researchers at the Institute for Bioengineering of Catalonia, the Gwangju Institute of Science and Technology, and the University of Pennsylvania Perelman School of Medicine have used single-cell RNA sequencing to identify further subtypes of cells in the kidney, and gain new insights into the molecular mechanisms of chronic kidney disease (CKD).
The team demonstrated major changes in cellular diversity in mouse models of kidney fibrosis, in which proximal tubule (PT) cells in particular showed altered differentiation and were highly vulnerable to dysfunction.
Notably, the nuclear receptors estrogen-related receptor alpha (ESRRA) and peroxisome proliferator-activated receptor alpha (PPARA), were shown to maintain both cell differentiation and metabolism by regulating expression of cellular metabolism and key cell type-specific genes.
These findings provide new opportunities for the therapeutic manipulation of cell fate, PT cell differentiation and metabolism, based on their dependence on nuclear receptors, the study authors reported in the December 9, 2020, online edition of Cell Metabolism.
"This is the first study to show that nuclear receptors such as ESRRA and PPARA protect against CKD by coupling cell metabolism and differentiation," said study lead author Katalin Susztak.
Associated with the increasing global incidence of type 2 diabetes, CKD affects approximately 800 million people worldwide and can progress to end-stage renal disease (ESRD).
As such, CKD has now become the 10th leading cause of global mortality and represents a significant health and economic burden, with limited treatment options.
"Approximately 30% of diabetes patients develop kidney disease, with approximately 30 million people in the U.S. having CKD, of whom half a million have ESRD," said Susztak, a professor in the Department of Medicine at the University of Pennsylvania Perelman School of Medicine in Philadelphia.
To help mitigate this progression, "we use inhibitors of the renin angiotensin system, while the new sodium glucose co-transport inhibitors show promise in this regard, but no drugs can halt or reverse disease progression or the PT atrophy that characterizes CKD.
PT cell atrophy strongly correlates with impaired renal function, with PT cells being particularly susceptible to toxic and hypoxic injury, being the main cause of acute kidney injury (AKI), while genetic studies have highlighted the role of PT-specific genes in kidney function.
Comprehensive genome-wide kidney tissue transcriptomic analysis has been used to define the molecular hallmarks of AKI in patients and mouse models, correlating multiple transcripts and kidney fibrosis.
Moreover, genes regulating lipid metabolism, including fatty acid oxidation (FAO) and oxidative phosphorylation (OXPHOS), are robustly correlated with disease in patients and mouse CKD models.
Pharmacological or genetic approaches that enhance FAO and mitochondrial metabolism have been shown to improve renal function via not fully understood mechanisms.
"The [cholesterol lowering] PPARA agonist fenofibrate, has shown early promise in this regard in multiple studies, but is associated with multiple side effects," said Susztak.
Furthermore, mitochondrial defects can lead to leakage of mitochondrial DNA into the cytoplasm, activating cytokine release, immune cell influx and fibrosis.
Single-cell RNA sequencing (scRNA-seq) analysis is changing the understanding of complex diseases, with previous research having identified 21 distinct cell types, including three novel kidney cells, and defined cell identity genes that can classify key kidney cell types in mice and humans.
In the new study co-led by Susztak, Jihwan Park, an assistant professor in the School of Life Sciences at the Gwangju Institute of Science and Technology in Korea and Nuria Montserrat, group leader of Pluripotency for Organ Regeneration at the Barcelona Institute of Technology in Spain, used scRNA-seq to identify differences in injured kidney tissue cellular composition and cell type-specific gene expression in mouse models of kidney disease.
This analysis highlighted major changes in cellular diversity in kidney disease, which markedly impacted whole-kidney transcriptomics findings.
"We found that bulk scRNAseq was mostly a read-out for cell diversity, although it provides little information on cell type specific changes," Susztak told BioWorld Science.
The researchers therefore used cell type-specific differential expression analysis to identify PT cells as being the key vulnerable cell type in CKD, with unbiased cell trajectory analyses showing that PT cell differentiation was altered in kidney disease.
Metabolism, including FAO and OXPHOS in PT cells, showed the strongest and most reproducible association with PT cell differentiation and disease.
"Gene expression data indicated that PT cell differentiation correlates the best with metabolism, specifically lipid metabolism," noted Susztak.
Coupling of cell differentiation and metabolism was then shown to be established by PPARA and especially by ESRRA, which directly controlled metabolic and PT cell-specific gene expression in mice and patient samples, while protecting mice from kidney disease.
"We demonstrated that ESRRA directly bound to the promoters of metabolic- and kidney-specific genes and improved both kidney function and metabolism," said Susztak.
This finding has important implications for drug discovery and development, "as nuclear receptors are targetable pathways, suggesting that better PPARA and ESRRA agonists could be developed," she said.
"Furthermore, using molecular modifications we might be able to generate drugs that are more specific to PT cells."