Trained innate immunity in response to nuclear antigens in systemic lupus erythematosus

Abstract Background and Aims Systemic lupus erythematosus (SLE) is an autoimmune disease directed against nuclear antigens, including those derived from apoptotic microparticles (MPs) and neutrophil extracellular traps (NETs). Innate immune cells display an hyperreactive phenotype in patients with SLE, with increased expression of immunostimulatory surface markers and increased production of proinflammatory cytokines. We hypothesize that the hyperreactive phenotype of innate immune cells in SLE is caused by the induction of trained immunity. Trained immunity is a de facto innate immune memory elicited by an initial stimulus that induces metabolic and epigenetic changes and results in a more vigorous inflammatory response to subsequent stimuli. Here we aim to investigate whether nuclear autoantigens derived from MPs and NETs can induce trained immunity in SLE patients. Method To investigate the capability of MPs and NETs to induce trained immunity we stimulated healthy PBMCs with isolated NETs and MPs or with plasma from SLE patients for 24 hours, washed and rested the cells for five days. Cells were restimulated with lipopolysaccharide (LPS) and Pam3CSK4. To test the activation status of innate immune cells, PBMCs were isolated from patients with SLE and healthy controls and stimulated the cells with different TLR agonists. Cytokine production was measured using ELISA. Immune cell subsets in SLE patients were analyzed by flow cytometry. We performed RNA sequencing and Chromatin immunoprecipitation (ChIP) sequencing for histone 3 lysine 4 trimethylation (H3K4me3) on monocytes from SLE patients and healthy controls. Results We found that in vitro both MPs and NETs, as well as plasma from SLE patients, can induce trained immunity. Initial stimulation with MPs, NETs or SLE plasma resulted in increased production of Tumor necrosis factor (TNF) and Interleukin (IL) 6 upon restimulation with different TLR agonists. Assessment of circulating immune cells showed higher percentages of monocytes in SLE patients compared to healthy controls, and we found that circulating monocytes from SLE patients produce increased levels of pro-inflammatory cytokines (IL-6, IL-1ß, TNF) after stimulation with Toll-like receptor agonists, indicating trained immunity. This is accompanied by increased expression of metabolism and inflammation-related genes, underscoring the hyperreactive phenotype typical in trained innate immune cells. Epigenetic analysis of monocytes revealed major changes in H3K4me3, an epigenetic mark associated with trained immunity. Conclusion Our findings provide new insight into the pathogenesis of SLE by showing that trained immunity can be elicited by SLE-related antigens present in MPs and NETs, and demonstrating that that circulating monocytes from SLE patients have a trained immunity phenotype. Trained immunity yields a possible biomarker for the risk of SLE flares and offers a new potential target for developing therapeutic strategies.