Background/Purpose: The role of complement in Systemic Lupus Erythematosus (SLE) is complex as complement deficiencies in early components of the classical pathway cause lupus with high penetrance, yet terminal complement activation at the level of the glomeruli causes kidney inflammation and damage in SLE. Aberrant accumulation or processing of dying cells that expose nucleosomes is thought to stimulate the production of lupus autoantibodies. To determine the relative contributions of excess dying cells, complement and autoantibodies, we created novel genetic models of lupus that contained one or more of these abnormalities, all on the C57BL/6 background.

Methods: Bicongenic mice were created by crossing mice deficient in the apoptotic cell (AC) opsonin, MFG-E8 (that accumulates AC in the germinal centers of the spleen), to sle1 mice (that produce autoantibodies to chromatin). These mice were then backcrossed to either C1q or C3 KO mice to create triple mutants. Anti-chromatin and anti-ds-DNA autoantibodies were measured by ELISA assay (n=20-30 per group). Autoantibodies specificities were determined on pooled sera (n=15-20 per strain) with the Autoantigen Microarray Super Panel (UT Southwestern Microarray Core). AC accumulation in the spleen and kidneys was detected by TUNEL staining (Roche). Kidney damage was evaluated by blood urea nitrogen (Amplite, AAT Bioquest), immuno-fluorescence for complement C3 (MP Biomedicals) and IgG2c (Jackson Immunoresearch), PAS staining and electron microscopy (E/M) (University of Washington Histology and Specialized Pathology Services).

Results:  sle1.MFG-E8 double mutant mice had significantly higher anti-DNA autoantibodies compared to sle1 mice alone (P<0.001). When sle1.MFG-E8 mice were crossed to C1q or C3 KO to create triple mutant mice, anti-chromatin and anti-DNA autoantibodies increased further (triple mutants compared to double mutants; p<0.05). In addition, when these strains were compared on an autoantigen array, the triple mutants demonstrated a significant broadening in antigenic specificity to include aggrecan, collagens, alpha-actinin, and others. Both triple mutant mouse strains showed AC accumulation in the kidneys and developed severe glomerulonephritis as determined by PAS staining. E/M analysis confirmed leukocyte infiltration and electron dense deposits near podocytes with podocyte destruction, as well as extensive tubular damage. Immunofluorescence analysis C1q triple mutants had abundant C3 deposition in the glomeruli, likely via activation of the alternative pathway, F4/80+ cell infiltrates in the interstitial and periglomerular space, and very prominent IgG2c deposits.

Conclusion: These studies reveal a stepwise contribution of all three elements studied here to lupus-like disease, and indicate that defective clearance of dying cells coupled with autoantibody production and impaired early complement function leads to a severe glomerulonephritis. Whereas it is likely that C3 activation contributed to nephritis in triple mutants lacking C1q, triple mutants lacking C3 also showed broadening of the autoantibody repertoire and developed intense nephritis suggesting that IgG FcgR engagement is sufficient to result in severe kidney damage.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/dissecting-the-role-of-single-complement-deficiencies-in-a-novel-model-for-apoptotic-cell-induced-lupus/