Kidney Week

Abstract: FR-PO088

Kidney-Resident Macrophages Exist in Unique Subsets and Demonstrate Subset-Specific Responses to AKI

Session Information

• AKI: Mechanisms - Inflammation/Sepsis/Remote Injury
  November 08, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Acute Kidney Injury

• 103 AKI: Mechanisms

Authors

• Lever, Jeremie M., University of Alabama at Birmingham, Birmingham, Alabama, United States

Jeremie M. Lever,

• Mrug, Michal, University of Alabama at Birmingham, Birmingham, Alabama, United States

Michal Mrug,

• Yoder, Bradley K., University of Alabama at Birmingham, Birmingham, Alabama, United States

Bradley K. Yoder,

• Agarwal, Anupam, University of Alabama at Birmingham, Birmingham, Alabama, United States

Anupam Agarwal,

• George, James F., University of Alabama at Birmingham, Birmingham, Alabama, United States

James F. George,

Background

Myeloid cell-mediated inflammation plays a key role in AKI. Kidney resident macrophages (KRMs) are embryonically derived, self-renewing, and are a distinct lineage from infiltrative macrophages. The means by which these cells differentiate and facilitate these events is unknown. Our objectives were to define KRM subsets using single cell RNA-sequencing (scRNAseq), identify subsets that are responsive to AKI, and determine target genes for intervention.

Methods

C57BL/6J mice were subjected to 20 min bilateral ischemia-reperfusion AKI (n=3) under ketamine/xylazine anesthesia. At 6 d post-injury, KRMs (F4/80^Hi/CD11b^Int) were isolated by FACS and subjected to scRNAseq using the Chromium 10X genomics platform. KRMs from untreated mice were used for controls. 6885 and 4253 cells were sequenced, for AKI and control, respectively.

Results

We used Seurat V3.0 to cluster cells with UMAP (uniform manifold approximation and projection) to reveal four major and two minor subsets. Cluster 0 constituted the majority of cells (57%). After IRI, cluster 0 decreased to 14% and cluster 1 increased from 0.2% to 36%. There were minimal changes to proportions of clusters 2 to 5. Expression of MHCII related transcripts (H2-Aa, H2-Ab1, and Cd74) in cluster 1 were decreased by 2 to 8-fold compared with cluster 0 (p<10^-100). Transcripts increased in cluster 1 included Ctsd, F13a1, Fabp5, Fos, Cxcl2 (p<10^-20). Pseudotemporal ordering produced by the Monocle algorithm (V2.3.6), which constructs a “trajectory” of cellular progress through differentiation, predicted that cluster 0 transitions to cluster 1 and that there are 2 potential decision points during that transition. Gene ontology analysis suggests cluster 1 is enriched for transcripts involved in an immune defensive response (e.g. innate immune response p=2.9x10^-12, leukocyte differentiation p=1.6x10^-11, inflammatory response p=2.0x10^-11).

Conclusion

Six transcriptionally unique KRM subsets exist and demonstrate subset-specific responses to AKI. The largest subset (cluster 0) demonstrated injury-responsive changes defined by downregulation of transcripts associated with MHCII that are consistent with a defensive posture involving innate immunity. KRM heterogeneity at the single cell level provides an opportunity to leverage KRM biology for interventions for AKI or the AKI to CKD transition.

Funding

• NIDDK Support