| Authors |
Wu H, Kirita Y, Donnelly EL, Humphreys BD
|
| Submitted By |
Benjamin Humphreys on 1/23/2019 |
| Status |
Published |
| Journal |
Journal of the American Society of Nephrology : JASN |
| Year |
2019 |
| Date Published |
1/1/2019 |
| Volume : Pages |
30 : 23 - 32 |
| PubMed Reference |
|
| Abstract |
A challenge for single-cell genomic studies in kidney and other solid tissues is
generating a high-quality single-cell suspension that contains rare or
difficult-to-dissociate cell types and is free of both RNA degradation and
artifactual transcriptional stress responses., We compared single-cell RNA
sequencing (scRNA-seq) using the DropSeq platform with single-nucleus RNA
sequencing (snRNA-seq) using sNuc-DropSeq, DroNc-seq, and 10X Chromium platforms
on adult mouse kidney. We validated snRNA-seq on fibrotic kidney from mice 14
days after unilateral ureteral obstruction (UUO) surgery., A total of 11,391
transcriptomes were generated in the comparison phase. We identified ten
clusters in the scRNA-seq dataset, but glomerular cell types were absent, and
one cluster consisted primarily of artifactual dissociation-induced stress
response genes. By contrast, snRNA-seq from all three platforms captured a
diversity of kidney cell types that were not represented in the scRNA-seq
dataset, including glomerular podocytes, mesangial cells, and endothelial cells.
No stress response genes were detected. Our snRNA-seq protocol yielded 20-fold
more podocytes compared with published scRNA-seq datasets (2.4% versus 0.12%,
respectively). Unexpectedly, single-cell and single-nucleus platforms had
equivalent gene detection sensitivity. For validation, analysis of frozen day 14
UUO kidney revealed rare juxtaglomerular cells, novel activated proximal tubule
and fibroblast cell states, and previously unidentified tubulointerstitial
signaling pathways., snRNA-seq achieves comparable gene detection to scRNA-seq
in adult kidney, and it also has substantial advantages, including reduced
dissociation bias, compatibility with frozen samples, elimination of
dissociation-induced transcriptional stress responses, and successful
performance on inflamed fibrotic kidney.
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