1844: Disease-Associated Macrophages Express an Injury-Associated Gene Program and Localize to Distinct Compartments in Proliferative and Mixed Histologic Classes of Lupus Nephritis

Background/Purpose: In collaboration with the AMP-RA/SLE network, we identified disease-associated macrophages (D-Macs) in kidney biopsies from 155 patients with active lupus nephritis (LN) and 30 controls, using single-cell RNA sequencing of ~25,000 myeloid cells (Fig. 1A, B). D-Macs comprised several macrophage subsets and expressed an injury-associated gene program, including TREM2, CD63, CTSD, APOE, SPP1, GPNMB, CD9, and FABP5 (Fig. 1C), which are conserved across monocytes and macrophages in other injured tissues. D-Macs showed positive correlation with the ISN/RPS activity index and were more prevalent in proliferative and mixed histological classes (III/IV±V) compared to the pure membranous class (V) or healthy individuals (not shown). In this follow-up study, we explored the origins and significance of D-Macs in proliferative/mixed LN.

Methods: We analyzed kidney biopsies from LN patients and controls using single-cell RNA sequencing to identify distinct cell populations. Spatial transcriptomics was performed with a custom gene panel to map myeloid subtypes in FFPE kidney samples. We also examined transcriptomic responses to putative nephritic glomerular factors plus M-CSF in healthy CD14 monocytes using flow cytometry and RNA sequencing.

Results: Bioinformatic analysis suggested that D-Macs likely differentiate in situ from infiltrating monocytes or resident macrophages, indicating a process of "convergent differentiation" where blood and tissue myeloid cells adopt similar transcriptional profiles in response to factors from diseased kidneys (Fig. 1D-G). To gain further insight into D-Mac differentiation and function, we mapped each D-Mac subset alongside other myeloid populations across renal compartments and in relation to lesions in FFPE sections from 6 LN patients with proliferative or mixed histologic classes (mean ISN/RPS activity = 13.5; S.D = 5.8) and 2 controls. Certain D-Mac subsets were statistically enriched in glomeruli with proliferative lesions (Glomerular D-Macs), while others were mainly found in the tubulointerstitium (Tubulointerstitial D-Macs) (Fig. 2), suggesting that distinct D-Mac subsets may differentiate from different precursors in a compartment-specific manner.

To determine whether factors from nephritic glomeruli drive D-Mac differentiation from infiltrating CD14+ or CD16+ monocytes, we screened 40 inflammatory mediators identified through single-cell data, spatial transcriptomics, histology, and literature review. Results showed that several glomerular factors—such as immune complexes with resiquimod, cell debris, and TGF-β—promoted the induction of D-Mac-like states, including an injury-associated gene program similar to that observed in patients, from both CD14+ (Fig. 3) and CD16+ (not shown) monocyte precursors.

Conclusion: Our findings highlight key glomerular mediators in proliferative lupus nephritis that may promote D-Mac differentiation from infiltrating monocytes. They support the idea that distinct monocyte states respond similarly to nephritic factors, converging on a shared cellular phenotype. Ongoing studies are focused on elucidating the functional roles of these cells.