Background/Purpose:

Poor renal prognosis in lupus nephritis (LN) is associated with an abundance of renal macrophages and dendritic cells (DCs) but the role of these cells is not well understood. Because mouse models provide a system for studying the immune response in LN, we characterized the transcriptomes of myeloid cells during pre- and post-nephritic disease in Sle.Yaa1 mice that have an extra copy of TLR7 and NZB/W mice in which an activated adaptive immune system drives disease.

Methods: We sorted PBMCs and dissociated renal cells from pre- and post-nephritic Sle.Yaa1 and NZB/W mice (3-4 per group) using (i) CD45 beads/antibodies to isolate all immune cells or (ii) CD11b/CD11c to enrich for myeloid cells. We performed single cell transcriptomic profiling using 10x genomics and analyzed droplets that contained >500 genes and UMIs after doublet removal. We performed coarse and fine clustering and differential expression (DE) (Seurat v3.0); gene set enrichment; trajectory (analysis Monocle v2.0 and Velcyto); and integration (Harmony). We identified residential vs. infiltrating cells by co-clustering intrarenal with blood data and by using published residential and infiltration gene signatures. We further identified macrophage and DCs subsets by scoring clusters using gene signatures generated from bulk RNASeq of intrarenal myeloid cells sorted by CD11b/CD11c and F4/80 expression.

Results: Several myeloid clusters were identified using data from >5000 cells from each mouse strain. Several novel subclusters of macrophages and DCs were identified in nephritic mice. Macrophages from pre- and post-nephritic mice clustered separately. Trajectory analysis of these cells indicated the acquisition of activation and inflammatory genes in nephritic mice including IL1b, AP1 (Fos and Jun), chemokines, cytoskeleton remodeling genes and polarization towards an alternative phenotype. Loss of Mmp13 and increased Mmp12 expression further suggests acquisition of a profibrotic phenotype. Fcrl5, Fcrg4 patrolling monocytes were found only in nephritic Sle.Yaa1 mice, confirming their expansion in TLR7 overexpressing models. Putative cluster functions were based on the top DE genes and gene set enrichment: macrophages had characteristics of resident cells and were enriched for phagocytosis, lysosomal function and antigen processing/presentation whereas DCs were enriched for transendothelial migration, chemokine signaling and cell adhesion genes, all characteristics of infiltrating cells.  pDCs were enriched in genes in the glycolysis pathway whereas CD103 DCs were highly enriched for genes involved in oxidative phosphorylation.

Conclusion: Resident macrophages are the major myeloid subpopulation in pre-nephritic kidneys in both models and acquire an activated and profibrotic phenotype during nephritis. Post-nephritic kidneys from both models contained novel subclusters of expanded residential and infiltrating macrophages and DCs. Only Sle.Yaa1 kidneys contained patrolling monocytes. These analyses generate new hypotheses about the development of tissue damage in lupus kidney disease and can be used to identify molecular features shared with myeloid cells from human lupus kidneys for targeted follow-up studies.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/the-identification-of-shared-and-unique-myeloid-cell-states-in-pre-and-post-nephritic-lupus-mouse-models-sle-yaa1-and-nzbw/