Methylseq, single-nuclei RNAseq, and discovery proteomics identify pathways associated with nephron-deficit CKD in the HSRA rat model

Low nephron numbers are associated with an increased risk of developing chronic kidney disease (CKD) and hypertension, which are significant global health problems. To investigate the impact of nephron deficiency, our laboratory developed a novel inbred rat model (HSRA rat). In this model, ∼75% of offspring are born with a single kidney (HSRA-S), compared with two-kidney littermates (HSRA-C). HSRA-S rats show impaired kidney development, resulting in ∼20% fewer nephrons. Our previous data and current findings demonstrate that nephron deficit (failure of one kidney to form and altered development in the remaining kidney) predisposes HSRA-S to CKD late in life (with increased proteinuria by 18 mo of age in HSRA-S = 51 ± 3.4 vs. HSRA-C = 8 ± 1.5 mg/24 h). To understand early molecular mechanisms contributing to the increased predisposition to CKD, Methylseq using reduced representation bisulfite sequencing, single-nuclei (sn)RNAseq, and discovery proteomics were performed in kidneys of 4-wk-old HSRA rats. Methylation analysis revealed a small number of differences, including five differentially methylated cytosines and six differentially methylated regions between groups. The snRNAseq analysis identified differentially expressed genes in most kidney cell types, with several hundred genes dysregulated depending on the analysis method (Seurat vs. DESeq2). Notably, many genes are involved in kidney development. Discovery proteomic analysis identified 366 differentially expressed proteins. A key finding was dysregulation of <i>Deptor</i>/DEPTOR and <i>Amdhd2</i>/AMDHD2 across omics layers, suggesting a potential role in compensatory mechanisms or the genetic basis of altered kidney development. Further understanding of these mechanisms may guide interventions to preserve nephron health and slow kidney disease progression.<b>NEW & NOTEWORTHY</b> The HSRA rat is a novel model of nephron deficiency and provides a unique opportunity to study the association between nephron number and chronic kidney disease (CKD). Previous work characterized the impact of age, hypertension, and diabetes on the development of CKD in HSRA animals. This study examined early changes in epigenetics, cell-type specific transcriptome, and proteomic changes in the kidney that likely predispose the model to CKD with age.