Nicotinamide riboside activates renal metabolism and protects the kidney in a model of Alport syndrome.

Chronic kidney disease (CKD) is associated with renal metabolic disturbances, including impaired fatty acid oxidation (FAO). Nicotinamide adenine dinucleotide (NAD + ) is a small molecule that participates in hundreds of metabolism-related reactions. NAD + levels are decreased in CKD, and NAD + supplementation is protective. However, both the mechanism of how NAD + supplementation protects from CKD, as well as the cell types most responsible, are poorly understood. Using a mouse model of Alport syndrome, we show that nicotinamide riboside (NR), an NAD + precursor, stimulates renal peroxisome proliferator-activated receptor α signaling and restores FAO in the proximal tubules, thereby protecting from CKD in both sexes. Bulk RNA-sequencing shows that renal metabolic pathways are impaired in Alport mice and dramatically activated by NR in both sexes. These transcriptional changes are confirmed by orthogonal imaging techniques and biochemical assays. Single nuclei RNA-sequencing and spatial transcriptomics, both the first of their kind from Alport mice, show that NAD + supplementation restores FAO in the proximal tubules with minimal effects on the podocytes. Finally, we also report, for the first time, sex differences at the transcriptional level in this Alport model. Male Alport mice had more severe inflammation and fibrosis than female mice at the transcriptional level. In summary, the data herein identify both the protective mechanism and location of NAD + supplementation in this model of CKD.

The chromatin landscape of healthy and injured cell types in the human kidney.

There is a need to define regions of gene activation or repression that control human kidney cells in states of health, injury, and repair to understand the molecular pathogenesis of kidney disease and design therapeutic strategies. Comprehensive integration of gene expression with epigenetic features that define regulatory elements remains a significant challenge. We measure dual single nucleus RNA expression and chromatin accessibility, DNA methylation, and H3K27ac, H3K4me1, H3K4me3, and H3K27me3 histone modifications to decipher the chromatin landscape and gene regulation of the kidney in reference and adaptive injury states. We establish a spatially-anchored epigenomic atlas to define the kidney's active, silent, and regulatory accessible chromatin regions across the genome. Using this atlas, we note distinct control of adaptive injury in different epithelial cell types. A proximal tubule cell transcription factor network of ELF3, KLF6, and KLF10 regulates the transition between health and injury, while in thick ascending limb cells this transition is regulated by NR2F1. Further, combined perturbation of ELF3, KLF6, and KLF10 distinguishes two adaptive proximal tubular cell subtypes, one of which manifested a repair trajectory after knockout. This atlas will serve as a foundation to facilitate targeted cell-specific therapeutics by reprogramming gene regulatory networks.

miRNA and mRNA Signatures in Human Acute Kidney Injury Tissue.

Acute kidney injury (AKI) is an important contributor to the development of chronic kidney disease (CKD). There is a need to understand molecular mediators that drive either recovery or progression to CKD. In particular, the role of miRNA and its regulatory role in AKI is poorly understood. We performed miRNA and mRNA sequencing on biobanked human kidney tissues obtained in the routine clinical care of patients with the diagnoses of AKI and minimal change disease (MCD), in addition to nephrectomized (Ref) tissue from individuals without known kidney disease. Transcriptomic analysis of mRNA revealed that Ref tissues exhibited a similar injury signature to AKI, not identified in MCD samples. The transcriptomic signature of human AKI was enriched with genes in pathways involved in cell adhesion and epithelial-to-mesenchymal transition (e.g., CDH6, ITGB6, CDKN1A ). miRNA DE analysis revealed upregulation of miRNA associated with immune cell recruitment and inflammation (e.g., miR-146a, miR-155, miR-142, miR-122). These miRNA (i.e., miR-122, miR-146) are also associated with downregulation of mRNA such as DDR2 and IGFBP6 , respectively. These findings suggest integrated interactions between miRNAs and target mRNAs in AKI-related processes such as inflammation, immune cell activation and epithelial-to-mesenchymal transition. These data contribute several novel findings when describing the epigenetic regulation of AKI by miRNA, and also underscores the importance of utilizing an appropriate reference control tissue to understand canonical pathway alterations in AKI.

The chromatin landscape of healthy and injured cell types in the human kidney.

There is a need to define regions of gene activation or repression that control human kidney cells in states of health, injury, and repair to understand the molecular pathogenesis of kidney disease and design therapeutic strategies. However, comprehensive integration of gene expression with epigenetic features that define regulatory elements remains a significant challenge. We measured dual single nucleus RNA expression and chromatin accessibility, DNA methylation, and H3K27ac, H3K4me1, H3K4me3, and H3K27me3 histone modifications to decipher the chromatin landscape and gene regulation of the kidney in reference and adaptive injury states. We established a comprehensive and spatially-anchored epigenomic atlas to define the kidney's active, silent, and regulatory accessible chromatin regions across the genome. Using this atlas, we noted distinct control of adaptive injury in different epithelial cell types. A proximal tubule cell transcription factor network of ELF3 , KLF6 , and KLF10 regulated the transition between health and injury, while in thick ascending limb cells this transition was regulated by NR2F1 . Further, combined perturbation of ELF3 , KLF6 , and KLF10 distinguished two adaptive proximal tubular cell subtypes, one of which manifested a repair trajectory after knockout. This atlas will serve as a foundation to facilitate targeted cell-specific therapeutics by reprogramming gene regulatory networks.

Spatial transcriptomics and the kidney.

PURPOSE OF REVIEW: The application of spatial transcriptomics technologies to the interrogation of kidney tissue is a burgeoning effort. These technologies share a common purpose in mapping both the expression of individual molecules and entire transcriptomic signatures of kidney cell types and structures. Such information is often superimposed upon a histologic image. The resulting datasets are readily merged with other imaging and transcriptomic techniques to establish a spatially anchored atlas of the kidney. This review provides an overview of the various spatial transcriptomic technologies and recent studies in kidney disease. Potential applications gleaned from the interrogation of other organ systems, but relative to the kidney, are also discussed. RECENT FINDINGS: Spatial transcriptomic technologies have enabled localization of whole transcriptome mRNA expression, correlation of mRNA to histology, measurement of in situ changes in expression across time, and even subcellular localization of transcripts within the kidney. These innovations continue to aid in the development of human cellular atlases of the kidney, the reclassification of disease, and the identification of important therapeutic targets. SUMMARY: Spatial localization of gene expression will complement our current understanding of disease derived from single cell RNA sequencing, histopathology, protein immunofluorescence, and electron microscopy. Although spatial technologies continue to evolve rapidly, their importance in the localization of disease signatures is already apparent. Further efforts are required to integrate whole transcriptome and subcellular expression signatures into the individualized assessment of human kidney disease.

Skeletal muscle metabolic responses to physical activity are muscle type specific in a rat model of chronic kidney disease.

Chronic kidney disease (CKD) leads to musculoskeletal impairments that are impacted by muscle metabolism. We tested the hypothesis that 10-weeks of voluntary wheel running can improve skeletal muscle mitochondria activity and function in a rat model of CKD. Groups included (n = 12-14/group): (1) normal littermates (NL); (2) CKD, and; (3) CKD-10 weeks of voluntary wheel running (CKD-W). At 35-weeks old the following assays were performed in the soleus and extensor digitorum longus (EDL): targeted metabolomics, mitochondrial respiration, and protein expression. Amino acid-related compounds were reduced in CKD muscle and not restored by physical activity. Mitochondrial respiration in the CKD soleus was increased compared to NL, but not impacted by physical activity. The EDL respiration was not different between NL and CKD, but increased in CKD-wheel rats compared to CKD and NL groups. Our results demonstrate that the soleus may be more susceptible to CKD-induced changes of mitochondrial complex content and respiration, while in the EDL, these alterations were in response the physiological load induced by mild physical activity. Future studies should focus on therapies to improve mitochondrial function in both types of muscle to determine if such treatments can improve the ability to adapt to physical activity in CKD.