Abstract: FR-PO1006
Single-Cell Analysis Identifies the Interaction of Altered Renal Tubules With Basophils Orchestrating Kidney Fibrosis
Session Information
- CKD: Pathobiology - I
November 04, 2022 | Location: Exhibit Hall, Orange County Convention Center‚ West Building
Abstract Time: 10:00 AM - 12:00 PM
Category: CKD (Non-Dialysis)
- 2203 CKD (Non-Dialysis): Mechanisms
Authors
- Doke, Tomohito, University of Pennsylvania, Philadelphia, Pennsylvania, United States
- Abedini, Amin, University of Pennsylvania, Philadelphia, Pennsylvania, United States
- Aldridge, Daniel, University of Pennsylvania, Philadelphia, Pennsylvania, United States
- Park, Jihwan, University of Pennsylvania, Philadelphia, Pennsylvania, United States
- Balzer, Michael S., University of Pennsylvania, Philadelphia, Pennsylvania, United States
- Shrestha, Rojesh, University of Pennsylvania, Philadelphia, Pennsylvania, United States
- Coppock, Gaia M., University of Pennsylvania, Philadelphia, Pennsylvania, United States
- Hunter, Christopher A., University of Pennsylvania, Philadelphia, Pennsylvania, United States
- Susztak, Katalin, University of Pennsylvania, Philadelphia, Pennsylvania, United States
Background
Inflammation is an important component of fibrosis; however, immune processes that orchestrate kidney fibrosis are not well understood. Here we apply single-cell sequencing to a mouse model of kidney fibrosis. We identify a subset of kidney tubule cells with a profibrotic-inflammatory phenotype characterized by the expression of cytokines and chemokines associated with immune cell recruitment.
Methods
Single-cell libraries were generated from the control and unilateral ureteral obstruction (UUO) model of kidney fibrosis and patient samples using the 10x system and sequenced 77,393 high-quality mice and 13,380 human cells after quality control. Clustering and data processing and cell trajectory analysis followed established pipelines. Bulk RNA sequencing and in silico deconvolution was used to define cell fractions. Flow sorting (FACS) with established antibodies and in situ hybridization (ISH) were used for validation. Two complementary methods were used to delete basophils, using the Mcpt8Cre-DTR mice and MAR-1 antibody. Flow sorted basophils were analyzed in vitro.
Results
Unbiased clustering identified 28 cell populations in renal fibrosis. Sub-clustering of UUO proximal tubule cells (PT) cells identified 8 PT cell types including profibrotic PT cells that expressed a proinflammatory gene signature (Il34, Tnfrsf12a, CD74, Pdgfb, Cxcl1, Cxcl10, and Cxcl16). Receptor-ligand interaction analysis and experimental validation indicate that CXCL1 secreted by profibrotic tubules recruits CXCR2+ basophils. Basophils were identified based on the expression of Mcpt8, Fcer1a, and Cd200r3, and their presence were validated by FACS and ISH. In mice, these basophils are an important source of IL-6 and recruitment of Th17 cells. Genetic deletion or antibody-based depletion of basophils as well as IL-6 receptor blockade results in reduced renal fibrosis. Human kidney single-cell, bulk gene expression, and immunostaining validated the functional role of basophils in patients with kidney fibrosis.
Conclusion
Collectively, we identify profibrotic proximal tubules activating basophils and Th17 cells as important contributors to the development of renal fibrosis and suggest that targeting these cells might be a useful clinical strategy.
Funding
- NIDDK Support