Dnase1L3 deficiency leads to 100% penetrance of pediatric onset Systemic lupus erythematous (SLE) while reduce Dnase1L3 levels is seen in adult-onset. Dnase1L3 is an endonuclease primarily secreted by myeloid cells like macrophages into the serum, where it degrades apoptotic bodies and chromatin, which reduces auto antibody formation and prevents SLE in both humans and mice. Importantly, reduction in the Dnase1L3 protein levels occurs both in adult-onset SLE in humans, and in polygenic murine models. This suggests that different levels of Dnase1L3 contribute to SLE progression. Also, decreasing Dnase1L3 levels leads to progressively worse SLE. However, the amount of serum Dnase1L3 needed, or whether the impact of Dnase1L3 is analog or digital remains unknown. In order to reduce Dnase1L3 protein levels, we generated Dnase1L3fl/fl x LysM-cre+/− conditional knockout (cKO) mice on B6 background. We determined the contribution of macrophage Dnase1L3 by measuring the Dnase1L3 protein levels and enzymatic activity by Dnase activity assay. We found that cKO mice showed ~2-log reduction in the serum Dnase activity. These cKO mice represent a cell-specific model to study the impact of Dnase1L3 protein levels on SLE pathogenesis. Together these findings suggest that macrophage specific Dnase1L3 may be a key player in lupus, however there could be other nucleases involved which we plan to inhibit using certain specific inhibitors. Overall, our findings suggest that macrophage specific Dnase1L3 is critical to prevent autoantibody formation in protecting lupus onset

A hallmark symptom of SLE is elevated autoantibodies, especially anti-dsDNA antibodies in multiple organs and tissues. Deficiency of Dnase1L3, a serum endonuclease, allows the accumulation of self-DNA leading to autoantibody generation. The source of Dnase1L3 is primarily expressed by macrophages and dendritic cells. Complete Dnase1L3 deficiency is known to promote the SLE phenotype, however, the dosage required to prevent disease onset is unknown. We hypothesize that macrophages provide sufficient Dnase1L3 expression to regulate autoantibody levels in the host’s system. We generated conditional, macrophage specific Dnase1L3 knockout mice to assess disease progression. We collected serum samples from 6–50 weeks of age and measured total IgG and anti-dsDNA levels by ELISA to develop a longitudinal analysis of autoantibody elevation. At sacrifice, we observed pathology and measured IgG deposition in the kidney and heart by immunofluorescence. We observed that the elimination of Dnase1L3 in macrophages increased anti-dsDNA antibodies in the conditional knockout mice. In addition, we observed minor abnormalities in the glomeruli size for the conditional knockout mice as compared to the wild type mice. Our results indicate that macrophages are capable of providing protection against the development of SLE.

One leading cause of death in women is autoimmunity. One prominent autoimmune disease is Systemic Lupus Erythematosus (lupus). During lupus, inflammation is driven by autoantibodies that target cell-free DNA. Cell-free DNA is degraded by two serum endonucleases, Dnase1 and Dnase1L3. The loss of Dnase1L3 causes lupus in humans and mice, but the amount of Dnase1L3 necessary to prevent lupus onset is unknown. Since DnaselL3 is secreted by macrophages and dendritic cells, we hypothesized that loss of macrophage-derived DnaselL3 is sufficient to cause lupus-like phenotypes in mice. We generated conditional knockout (cKO) mice lacking DnaselL3 expression in macrophages. Sera was collected weekly from these mice until 50 weeks of age. To measure autoantibody induction, total IgG, total IgM, anti-dsDNA and anti-nuclear antibody levels were analyzed at 4, 20, 30 and 50 weeks. Sandwich ELISA was used to quantitate antibody levels in control and cKO mice. Homogeneous and peripheral anti-nuclear antibodies were detected by immunofluorescence, consistent with anti-dsDNA antibodies. Total IgG, anti-ds DNA antibody levels increased in cKO mice. We conclude that SLE-like phenotypes arise in mice due to the loss of macrophage Dnase1L3. This suggest that macrophage derived DnaselL3 is critical to controlling lupus. Plains Bridges to the Baccalaureate Program (PBB) (NIH 2R25GM83730-B)