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Abstract: PO0887

Single-Cell Transcriptional and Chromatin Accessibility Profiling Redefines Cellular Heterogeneity in the Adult Human Proximal Tubules

Session Information

Category: Development, Stem Cells, and Regenerative Medicine

  • 500 Development, Stem Cells, and Regenerative Medicine

Authors

  • Muto, Yoshiharu, Washington University in Saint Louis School of Medicine, Saint Louis, Missouri, United States
  • Wilson, Parker C., Washington University in Saint Louis School of Medicine, Saint Louis, Missouri, United States
  • Waikar, Sushrut S., Boston Medical Center, Boston, Massachusetts, United States
  • Humphreys, Benjamin D., Washington University in Saint Louis School of Medicine, Saint Louis, Missouri, United States
Background

Single nucleus RNA sequencing (snRNA-seq) has improved our understanding of cell-specific genes and pathways, however, relatively less is known about how chromatin accessibility contributes to cell identity. We hypothesized that an integrated analysis by snRNA and ATAC sequencing (snATAC-seq) would enhance our ability to detect unique cell types and states in the kidney, uncovering previously unrecognized cellular heterogeneity.

Methods

We performed snRNA-seq and snATAC-seq on 5 healthy adult kidney samples (3M and 2F, mean age = 55.8y, mean sCr = 1.07 mg/dl). Nuclear preparations were processed using 10x Genomics 5’ v2 (snRNA) and Single Cell ATAC (snATAC) Chromium kits, sequenced and counted with Cell Ranger. Seurat was used to integrate snRNA and snATAC datasets with label transfer. Chromatin interactions were predicted with Cicero and pseudotemporal ordering was performed with Monocle.

Results

We analyzed a total of 52,097 nuclei by snRNA-seq (n=19,985) and snATAC-seq (n=32,112), identifying 214,890 accessible chromatin regions that confer kidney cell type identity. This multi-modal analysis highlighted a unique subpopulation of proximal tubule (PT) cells characterized by increased chromatin accessibility in VCAM1 and a pro-inflammatory gene expression signature. Immunofluorescence studies showed that these cells are present in a scattered distribution in the kidney cortex. Transcription factor motif analysis implicated NFkB signaling in the transition between healthy PT and the VCAM1-positive subpopulation. We identified candidate regulatory regions that are predicted to interact with the VCAM1 promoter via cis-coaccessibility networks. Inter-species snRNA-seq data integration suggests this subpopulation exists in the post-IRI mouse kidney and bulk RNA-seq deconvolution implicates a potential role in CKD and kidney aging.

Conclusion

Our multi-omics approach improves the ability to detect unique cell states within the kidney and reveals a previously unrecognized subpopulation of proximal tubule cells with a pro-inflammatory signature.

Funding

  • NIDDK Support