IMIQUIMOD-INDUCED ONSET OF DISEASE IN LUPUS-PRONE NZB/W F1 MICE

PV012 / #474 Poster Topic: AS02 - Animal Models Background/Purpose Systemic lupus erythematosus (SLE) is a heterogeneous disease and onset of the disease in lupus-prone NZB/W F1 mice are typically between weeks 20 to 25. Kidney failure in female mice normally develops within 9 months. Here, we wanted to study if we could accelerate disease onset and possibly induce a more homogenous disease by topical treatment of imiquimod (IMQ), a toll-like receptor (TLR) 7 agonist. Activation of TLRs, especially TLR7 and TLR9 by nucleic acids, have been linked to one of many mechanisms contributing to the development and aggravation of autoimmune diseases such as SLE. Methods We applied IMQ to the ears of 5 weeks old NZB/W F1 mice 3 times per week for 5 weeks with endpoints set at experimental weeks 6 and 10. We used age-matched C57BL/6J and NZB/W F1 as control mice. Disease progression was closely monitored by weekly measurements of proteins in urine, serum levels of anti-dsDNA antibodies, and hematological analysis using IDEXX ProCyte Dx. At endpoint, spleen, lymph nodes and kidney were collected for flow cytometric analysis on lymphocyte activation, macrophage, and dendritic cell populations. IgG deposition in glomeruli and tubules was analyzed by immune electron microscopy. Classification of kidney damage was performed according to the 2003 ISN/RPS criteria on zinc-fixated paraffin embedded kidney sections stained with Periodic acid–Schiff staining. Cellular composition and tissue changes were assessed using immunohistochemistry and immunofluorescence, and mRNA gene expression of genes related to disease progression was analyzed using qPCR. Results In this study, none of the mice developed full-blown proteinuria. At experimental endpoints, at week 6 and 10 after treatment start, IMQ treated mice had elevated production of autoantibodies against dsDNA, RNA, and cardiolipin compared to control mice. Hematological analysis revealed hemolysis, thrombocytopenia, neutrophilia, and lymphocytopenia. Immune complex deposition in the kidneys was observed at both endpoints. Increased infiltration of activated lymphocytes into the kidneys was observed by both histological and flow cytometric analyses. An increase in CD8+ tissue residential memory cells (CD69+CD44+), monocytes and CD11c+ monocyte derived dendritic cells, CX3CR1+ M2 macrophages, and CX3CR1+ dendritic cells was observed. In contrast, a decrease in plasmacytoid dendritic cells and XCR1+ conventional dendritic cells were seen. In addition, we detected a rise in regulatory immune mechanisms such as an increase of FoxP3+CD4+ Tregs in kidneys, spleen and lymph nodes. Conclusions IMQ treatment in young NZB/W F1 mice induced breakage of tolerance against nuclear antigens, but failed to induce nephritis in these mice. The highest disease burden was in general seen after 5 to 6 weeks following treatment initiation. However, anti-dsDNA antibodies production increased after 4 to 5 weeks and 8 to 10 weeks post-treatment onset. The age-matched controls did not manifest disease within the experimental time frame.

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