296 Activated stress response genes and perturbation of regulatory pathways in anti-nuclear antibody positive individuals and SLE patients vary by cell type and race in single-cell transcriptomic analyses

Abstract

Background A universal hallmark of systemic lupus erythematosus (SLE) is the presence of antinuclear antibodies (ANAs). While their cognate antigens and ANA levels can associate with SLE subphenotypes, many individuals develop ANAs years prior to autoimmune disease manifestations or never develop any clinical symptoms. At present, the mechanisms governing the transition from autoantibody production to disease onset remain unknown. Methods To ascertain differences in immune cell populations and their transcriptional state, we performed scRNA-seq on peripheral blood mononuclear cells from 30 patients, including African and European American individuals with lupus-associated autoantibodies (n=10), SLE patients (n=10), and ANA- healthy controls (n=10). Transcriptomes were analyzed using canonical correlation analysis, the results of which were used by a smart local moving algorithm to identify cell population clusters and by Uniform Manifold Approximation and Projection for visualization. Differential expression of genes was assessed using Model-based Analysis of Single Cell Transcriptomics. Results Distinct cell populations could be identified for each disease classification. EA ANA +individuals showed a decrease in the number of naive B cells relative to SLE and healthy individuals; in contrast, AA ANA +individuals had higher numbers of centrocytes. Additionally, AA SLE patients demonstrated an enrichment of plasma cells relative to both ANA +and healthy individuals while FCRL5, a marker of dysfunctional B cells, was increased only in SLE memory B cells. Overall, AA ANA +cell populations showed an upregulation in a number of heat shock genes in ANA+ (HSPA6, HSPA1A, DNAJB1). Multipotent progenitor (MPP) cells from SLE patients of both ethnicities were marked by upregulation of interferon stimulated genes; ANA +MPPs, however, differed in that cells from AA individuals had an upregulation of housekeeping genes (GAPDH, PGK1) while anti-inflammatory NFKB1A was increased in EA MPPs. Comparison of a manually curated list of regulatory factors demonstrated dysregulation of several genes, including upregulation of TGFB across all AA ANA +cell clusters and an increase in CD46 in EA ANA +clusters. Similarly, gene set enrichment analysis of differentially regulated genes indicated activation of stress response pathways in AA and EA ANA +individuals. Conclusions These data indicate that substantial transcriptional and cell population differences exist among immune cells from ANA+, SLE, and healthy individuals, including an activated stress response and enhanced immune regulation in the cells from ANA +individuals. Our results suggest that cells from ANA +individuals may be regulating reactions to stimulation, and exhaustion of that response may allow for transition to SLE. Funding Source(s): NIH U54GM104938, U01AI101934, U19AI082714, and P30AR053483.