Complement in ANCA-associated glomerulonephritis.

BACKGROUND: Anti-neutrophil cytoplasmic antibodies (ANCA) are found in pauci-immune necrotizing crescentic glomerulonephritis. In the past, the role of complement in ANCA-associated vasculitis (AAV) was assumed to be minimal. More recently, however, it was found that blocking the complement cascade in a mouse model of AAV reduces glomerular damage. Immune complex deposits have been found in biopsies from AAV patients. In this study, we questioned whether immune complex formation or deposition may result in complement activation in ANCA-associated glomerulonephritis. METHODS: ANCA-positive patients from the Limburg Renal Registry were included between 1979 and 2011. Renal histology was documented together with immunoglobulin and complement immunofluorescence. In addition, C3d, properdin, C4d and mannose-binding lectin (MBL) were stained. Electron microscopy was performed. Circulating immune complexes were determined in a subset of patients, as well as C3 allotypes. RESULTS: C3c was found in 78 of 187 renal biopsies (41.7%) divided over 32.3% of proteinase-3 (PR3)-AAV patients and 52.3% of myeloperoxidase (MPO)-AAV patients (P = 0.006), whereas C3d was found positive in 51.1% of PR3-AAV patients and 70.4% of MPO-AAV patients (P = 0.105). C4d was found positive in 70.8%, properdin in 38.7% and MBL in 30.4% of patients. Whereas C4d and MBL positivity was similar between the AGN groups, properdin was more common in biopsies classified as crescentic compared with biopsies classified as focal or mixed. Renal biopsies positive for C3d and/or properdin showed more cellular crescents and less normal glomeruli compared with biopsies negative for C3d and/or properdin (P < 0.05). In 3 out of 43 renal biopsies analysed by electron microscopy, small electron dense deposits were found. In 14 of 46 patients analysed, circulating immune complexes were detectable. No association between histological findings and C3 allotypes was found. CONCLUSIONS: In the majority of AAV patients, no immune complex deposits were found in their renal biopsies. C3d, C4d and C5b-9 staining, however, was found to be positive in a majority of analysed renal biopsies. Importantly, C3d and properdin staining was associated with cellular crescents. We hypothesize that local immune complexes are quickly degraded in AAV and therefore not visible by electron microscopy. Our findings are compatible with the hypothesis that complement activation in AAV occurs predominantly via alternative pathway activation.

Evaluation of anti-PLA2R1 as measured by a novel ELISA in patients with idiopathic membranous nephropathy: a cohort study.

OBJECTIVES: Autoantibodies against the M-type phospholipase A2 receptor 1 (anti-PLA2R1) have been demonstrated to be very specific for idiopathic membranous nephropathy (MN). We studied a novel enzyme-linked immunosorbent assay (ELISA) and compared results with results obtained using an indirect immunofluorescence (IIF) and a Western blotting test (WB). METHODS: One-hundred nine patients with idiopathic MN were recruited between November 1979 and March 2011. The control cohort comprised serum samples from patients with secondary MN (n = 16) and nephrotic controls (n = 17). The presence of anti-PLA2R1 in serum samples obtained at the time of renal biopsy was determined using ELISA, IIF, and WB. RESULTS: With similar specificity (≥ 97%), sensitivity varied from 68% (IIF) to 72% (ELISA, WB). Remarkably, patients who were seronegative for anti-PLA2R1 more often entered spontaneous remission (P = .038), whereas seropositive patients were more frequently treated with immunosuppressive agents (P < .001). CONCLUSIONS: ELISA performs excellently in differentiating idiopathic from secondary MN. Furthermore, ELISA shared high agreement with WB and IIF.

Dendritic cells in renal biopsies of patients with ANCA-associated vasculitis.

BACKGROUND: Dendritic cells (DCs) maintain immune tolerance and are able to initiate immune responses. Their involvement in ANCA-associated vasculitis (AAV) is unknown. In this study, the participation of DC subsets is investigated in renal biopsies of AAV patients. METHOD: A total of 25 patients with biopsy-proven AAV and five healthy controls (HC) with normal renal histology were included. Renal biopsies were stained for mature (CD208), immature (CD209), plasmacytoid (CD303) and Langerhans (CD1a) DC subsets. Furthermore, T-cells were stained using a T-cell marker (CD3). The interstitial cellular infiltrate was graded semi-quantitatively from 0+ (= absence of cells) to 3+ (= numerous cells). Within the glomeruli, an absolute count was performed for positive cells. RESULTS: CD208+ and CD209+ cells were found within patients' glomeruli but not in HC (1 +/- 0.3 versus 0.08 +/- 0.1 cells/glom; 2 +/- 0.3 versus 0.1 +/- 0.07 cells/glom). An average of 0.3 +/- 0.1 cell/glom expressed CD3 in patients while few cells were found in HC (0.1 +/- 0.7 cell/glom). Focal interstitial cellular infiltrates were observed in patients' biopsies but not in HC. Interstitial infiltration with CD3+ and CD209+ cells was assessed at an average of 1+, but some glomeruli and tubuli were surrounded by CD3+ and CD209+ cells forming clusters. Serial sections revealed that CD209+ cells were present in CD3+ rich areas. CONCLUSION: Both mature and immature glomerular DCs are found in renal biopsies of patients with AAV. Immature DCs cluster with T-cells in interstitial infiltrates in these biopsies. Since DCs form aggregates in T-cell areas, we hypothesize that these cells interact with each other and are involved in lymphoid neogenesis.