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Abstract
Background Lupus nephritis (LN) is a common and severe complication of systemic lupus erythematosus requiring renal biopsy to guide treatment decisions. Despite standard-of-care therapy, a third of patients with LN class III, IV, or V show a progressive decline in kidney function. Identifying distinct inflammatory processes associated with LN using non-invasive tools may improve treatment targeting.
Method We applied mass cytometry using four 48-marker panels to characterize peripheral blood mononuclear cells from 140 patients with active, biopsy-proven proliferative (class III or IV +/- V, n=98) or membranous (class V, n=42) nephritis and 40 healthy controls in the Accelerated Medicine Partnership RA/SLE Network Phase II study. Renal response was determined at 1 year (n=107), and longitudinal samples were collected (n=49).
Results By applying covarying neighborhood analysis, we observed a marked enrichment of cell neighborhoods that expressed type I interferon (IFN-I)-induced proteins, including MX1 (figure 1A) and Siglec1, reflecting a simple cytometric detection of an IFN-I signature. Specific immunophenotypic alterations, including activated and proliferating cells (Ki67hi) across immune cell types, such as plasmablasts (B8) and Tph (T11), were associated with IFN-I signaling (figure 1B). Amongst cells strongly associated with IFN-I, we identified a naive-like (CD45RA+CCR7+) CD4 T cell cluster (T12) expressing low levels of TCF1, suggesting early activation (figure 1C). We confirmed that IFN-I had an additive effect with TCR stimulation in decreasing TCF1 (TCF7 gene) expression in naive T cells using a recently published bulk RNAseq dataset1 (figure 2). Longitudinally, the cell subsets strongly associated with IFN-I, including T12, did not change in abundance over time independently from the renal outcome (longitudinal mixed effect model in complete responders p = 0.37, non-responder p = 0.62). As IFN-I signaling and the frequency of cell subsets varied across individuals, we stratified all samples using an unsupervised analysis based on 55 immune cell subsets. We identified 3 LN patient subgroups: the first group (G0) included all controls and 20% baseline LN patients; the two others (G1, G2) included only LN patients (46 patients each). G2 was associated with an increased proportion of GZMB+ T cells which were not correlated with IFN-I score (figure 1C). G1 displayed the highest cytometric IFN- I scores and was driven mainly by cells enriched with IFN-I-induced proteins, including T12 (figure 3A). Additionally, IFN-I cytometric score was strongly correlated with an IFN-I score in kidney leukocytes from paired samples (rho = 0.73, p < 0.001). Finally, after adjusting for a history of previous biopsy and immunosuppression at baseline, group G1 was significantly associated with non-response at 1 year (figure 3B).
Conclusion IFN-I signaling in LN patients is associated with persistent enrichment of altered and activated immune cells, despite immunosuppression. Patients with increased IFN-I signaling in the peripheral blood have increased IFN-I signaling in the kidney and represent a group with poor response to standard-of-care.
Reference
Sumida T, et al. Type I interferon transcriptional network regulates expression of coinhibitory receptors in human T cells, Nature Immunology. 2022;23(4):632–42.
Identification of cells enriched in type I interferon (IFN-I) signaling in lupus nephritis (LN). A. UMAP representing covarying neighborhoods results comparing LN patients against controls; colored cells are in neighborhoods associated positively (red) or negatively (blue) with LN after passing FDR 5%. The highlighted cells and names are the results of the cell clustering. B. UMAP of marker expression. C. Correlation plot of all cell clusters identifying amongst T cells, B cells, Myeloid (M) cells, and NK cells and the type I interferon score (rectangles). The rows and columns are organized using hierarchical clustering. The lower triangle represents all Spearman’s rho coefficients, and the upper triangle shows only the correlations passing FDR 5%.
Decreased expression of TCF7 gene (= TCF1 protein) in naïve T cells after TCR stimulation (Th0) with or without interferon b (IFNb) over time. Data generated from Sumida et al.
Identification of 3 subgroups of samples based on blood immunophenotyping. A. PCA colored by subgroups defined by k-means (left) and disease condition (right). B. Results of covarying neighborhood analysis comparing lupus nephritis patients in one subgroup against the others at baseline. The colors represent the frequencies of cells passing FDR 5% for each cell cluster after adjusting for race, previous biopsy, and immunosuppression use. Cells in red are enriched in the subgroup mentioned, and in blue are decreased. C. Baseline likelihood of complete response (CR) versus non-response (NR) to treatment at 1 year.
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