Kidney single-cell atlas reveals myeloid heterogeneity in progression and regression of kidney disease

Abstract

Short running title: Myeloid cell heterogeneity in kidney injury and repair Corresponding authors: Bryan Conway (Bryan.Conway@ed.ac.uk), Eoin O'Sullivan (Eoin.osullivan@ed.ac.uk), Tamir Chandra (Tamir.Chandra@ed.ac.uk) Laura Denby (Laura.Denby@ed.ac.uk) Word Count = 4810 Number of figures = 8 2 AbstractThe kidney has a limited capacity to repair following injury, however, the endogenous reparative pathways are not well understood. Here we employ integrated droplet-and platebased scRNA-seq in the murine reversible unilateral ureteric obstruction model to dissect the transcriptomic landscape at the single cell level during renal injury and resolution of fibrosis.We generate a comprehensive catalogue of the changes induced during injury and repair, revealing significant myeloid cell heterogeneity, which would not have been identifiable by conventional flow cytometry. We identify new markers for the myeloid populations within the kidney as well as identification of novel subsets including an Arg1 + monocyte population specific to early injury and a Mmp12 + macrophage subset exclusive to repair. Finally, using paired blood exchange to track circulating immune cells, we confirm that monocytes are recruited to the kidney early after injury and are the source of Ccr2 + macrophages that accumulate in late injury. Our data demonstrate the utility of complementary technologies to identify novel myeloid subtypes that may represent therapeutic targets to inhibit progression or promote regression of kidney disease. Methods Animal modelsAll protocols and surgical procedures were approved by the Animal Ethics Committee, University of Edinburgh. Animal experiments were conducted in accordance with the Animals Scientific Procedures Act UK 1986, under Home Office project licenses 70/8093 and 70/8867. Reversible unilateral ureteric obstruction model (R-UUO)The R-UUO model was performed as previously described (33). Briefly, 8 week old male C57BL/6JOlaHsd mice (Enviago) underwent laparotomy and the left ureter was isolated and the distal portion was ligated twice with 6/O black braided silk suture close to the bladder. In

The renal inflammatory network of nephronophthisis

“…Our findings add to the recently identified expression of CXCL17 in the kidney 37 , the last identified member of the CXC chemokine family in mammals 38,39 . CXCL17 attracts professional antigen presenting cells 40 , including macrophages, through its receptor CXCR8.…”

Identifying cell-enriched miRNAs in kidney injury and repair

“…During injury (UUO), the proportion of live PT cells significantly decreased, whereas the proportion of CD45 + immune cells, macrophages, and PDGFR-β + cells significantly increased ( Figure 1E). Upon reversal, there was significant reduction in the proportion of macrophages but a persistence of CD45 + cells, which likely reflects T cell presence during the repair phase as has been observed before (22,23). An increase in ECs with injury was noted; this may be a consequence of increased expression of CD31 upon endothelial activation and/or increased liberation of ECs from injured tissue.…”

“…Deconvolution of whole-kidney tissue sRNA-Seq data. The R-UUO model exhibits increased inflammatory cell numbers and a loss of healthy proximal tubules with injury, with partial recovery in R-UUO ( Figure 1, B and E), as we have previously described (22). We set out to explore whether the expression changes in whole-tissue sRNA-Seq of our highly enriched miRNAs mirrored the changes in the proportions of renal cells determined by flow cytometry.…”

“…To gain insight into the potential biological pathways influenced by the dynamic expression of miR-18a-5p and miR-194-5p, we determined the validated targets (Supplemental Table 1. We then cross-referenced the list of validated targets with expression data from single-cell RNA-sequencing (scRNA-sequencing) performed in the same model (22) and looked for targets for which the gene expression was inversely correlated with miRNA expression. This analysis revealed that for macrophage-derived miR-18a-5p several targets were involved in the cell cycle, e.g., Ccnd1 (Supplemental Figure 11), and TGF-β signaling, e.g., Tgfbr1.…”

Single-cell transcriptomics: a novel precision medicine technique in nephrology

“…These analyses identify key factors involved in diabetic kidney disease progression and thereby help in implementing new treatment approaches[80].Using the reversible unilateral ureteric obstruction model (R-UUO), changes observed in renal injury and repair were noted at the single-cell level[81]. The scRNA-seq analysis performed and revealed detailed characteristics of myeloid cell heterogeneity in damaged and recovering kidneys by identifying new monocyte and macrophage subsets that have not been observed in the kidney[81]. These data could identify potential therapeutictargets that can inhibit the progression of kidney disease and aid in the development of therapeutics for kidney diseases.…”