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.

Connective tissue growth factor and the cicatrization of cellular crescents in ANCA-associated glomerulonephritis.

BACKGROUND: Outcome in patients with anti-neutrophil cytoplasmic antibodies (ANCA)-associated glomerulonephritis (AGN) is difficult to predict. Scoring of renal biopsies has significant but limited predictive value. We investigated whether analysis of plasma and urine levels, and immunostaining of biopsies for the pro-fibrotic peptide connective tissue growth factor (CTGF), might improve prediction of renal outcome. METHODS: ANCA-positive patients were included. Renal biopsies were classified according to the AGN classification. Biopsies were stained for CTGF protein. CTGF was measured by ELISA at the time of renal biopsy in plasma and urine, and during follow-up in plasma. RESULTS: Eighty-two patients were included. CTGF staining was positive in crescentic lesions. Plasma CTGF at the time of renal biopsy was 2.4 ± 1.7 pmol/mL when compared with 0.5 ± 0.0 pmol/mL in healthy controls (P < 0.01). Plasma CTGF was associated with cellular crescents, but not when corrected for renal function. Plasma CTGF at baseline was associated with fibrous crescents in the follow-up biopsy, also after correction for renal function. Plasma CTGF at baseline predicted renal survival more accurately than the AGN classification. CONCLUSION: In AGN patients, CTGF was overexpressed in crescentic glomeruli. Baseline plasma CTGF predicted the percentage of fibrous crescents in later biopsies, and renal survival, suggesting that CTGF is involved in the cicatrization, as opposed to resolution of cellular crescents in AGN.

Estimating renal survival using the ANCA-associated GN classification.

A histopathological classification system for ANCA-associated vasculitis was recently published, but whether this system predicts renal outcome requires validation. Here, we analyzed data from 164 consecutive patients with biopsy-proven renal involvement of ANCA-associated vasculitis. The ANCA-associated GN (AGN) classification categorizes patients as having focal, mixed, crescentic, or sclerotic GN. Five-year renal survival rates by categories of the AGN classification scheme were 91% for focal, 69% for mixed, and 64% for crescentic (log-rank P<0.0001). Only one patient was classified as sclerotic. Furthermore, the percentage of normal glomeruli found on biopsy estimated renal survival with the same precision as did the AGN classification scheme. Patients classified as crescentic or mixed, however, had worse survival when the percentage of normal glomeruli was <25%. In conclusion, the AGN classification for renal biopsy specimens is a practical and informative scheme with which to categorize patients with ANCA-associated vasculitis, but adding the percentage of normal glomeruli to the system seems to improve its predictive value.

Improved outcome in anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis: a 30-year follow-up study.

BACKGROUND: Anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis has a poor prognosis. In the current study, we assessed whether prognosis in these patients improved over the last three decades. METHODS: In a large inception cohort, all consecutive patients with ANCA-associated glomerulonephritis were included between January 1979 and December 2009. Inclusion criteria were the presence of ANCA and the availability of a kidney biopsy. To assess renal and patient survival, patients were divided in three groups through time: 1979-89, 1990-2000 and 2001-09. RESULTS: A total of 181 patients were included. One-, 5- and 10-year survival was 77, 66 and 49%, respectively. Survival within the time groups was significantly different, yielding a hazard ratio for death of 2.9 for 1990-2000 and 3.9 for 1979-89 compared with 2001-09 (P < 0.001). Serum creatinine and active lesions as found in the kidney biopsy significantly decreased through the three decades. CONCLUSIONS: Both patient and renal survival in patients with ANCA-associated renal vasculitis have improved over the last three decades. We postulate that both earlier diagnosis and better therapeutic management of patients are responsible for this effect.

Immune complexes in acute adult-onset Henoch-Schonlein nephritis.

BACKGROUND: Adult-onset Henoch-Schönlein purpura nephritis (HSPN) and primary IgA (IgAN) nephropathy have been considered indistinguishable immunohistopathologically and are often considered as two extremes of one disease entity. We postulate that adult-onset Henoch-Schönlein can be distinguished histologically from primary IgAN and that both diseases differ in their immunopathological mechanisms. METHODS: Twenty consecutive patients with adult-onset HSPN were studied. Serum was analysed for circulating IgA immune complexes; renal biopsies were analysed by light and electron microscopy (EM). As disease controls, 40 IgAN patients were studied. RESULTS: Intracapillary leukocyte margination was seen in 15 of the 20 patients and cellular crescent formation in all renal biopsies of the HSPN patients. IgAN biopsies showed a few small crescents without intracapillary leukocytes. In 16 HSPN patients, EM was performed and in 10, no dense deposits were found. In all biopsies of IgAN patients, typical 'humps' were found. In 6 of 9 analysed HSPN patients, intermediate to large circulating immune complexes were found, whereas in 4 of 28 analysed patients with primary IgAN small circulating immune complexes were found. CONCLUSIONS: We consider adult-onset HSPN distinguishable in histology and ultrastructure from primary IgAN. We believe adult-onset Henoch-Schönlein to be a circulating immune complex disease.

Coexistence of anti-glomerular basement membrane antibodies and myeloperoxidase-ANCAs in crescentic glomerulonephritis.

BACKGROUND: In a substantial proportion of patients with crescentic glomerulonephritis (CGN), both anti-glomerular basement membrane (GBM) antibodies and antineutrophil cytoplasmic antibodies (ANCAs) with specificity for myeloperoxidase (MPO-ANCA) are detected. In the present study, we questioned whether histological and clinical features of patients with both ANCA and anti-GBM antibodies differ from those of patients with either ANCA or anti-GBM alone. METHODS: We reviewed the Limburg renal biopsy registry (1978 to 2003; n = 1,373) for cases of CGN. The presence of linear fluorescence on renal biopsy and the presence of ANCA and/or anti-GBM antibodies were measured. Subsequently, we assessed patient characteristics and follow-up and compared histological findings among the different groups. RESULTS: We identified 46 MPO-ANCA-positive, 10 double-positive, and 13 anti-GBM-positive patients. Mean ages were 63, 64, and 52 years (P = 0.04), and serum creatinine levels were 5.0, 10.3, and 9.6 mg/dL (445, 910, and 850 micromol/L), respectively (P = 0.01). Granulomatous periglomerular inflammation was found in either MPO-ANCA- or double-positive patients, but not in anti-GBM-positive patients with CGN without MPO-ANCAs. Patient survival among the 3 groups was different, although not statistically significant (log rank P = 0.17, with 75%, 79%, and 100% alive at 1 year, respectively). Renal survival analysis showed significant differences among the 3 groups (P = 0.04, with 65%, 10%, and 15% off dialysis therapy at 1 year, respectively). CONCLUSION: In patients with both anti-GBM antibodies and MPO-ANCAs, histological findings differ from those of patients with anti-GBM antibodies only. However, renal survival in these patients is not better than that in anti-GBM-positive patients and is worse compared with patients with MPO-ANCAs only.

Signs and symptoms of thin basement membrane nephropathy: a prospective regional study on primary glomerular disease-The Limburg Renal Registry.

BACKGROUND: To chart the epidemiology of primary glomerular disease by means of a prospective regional study in the southern part of The Netherlands. METHODS: Experienced renal technicians collected renal biopsies, blood, and 24-hour urine samples at the bed site in each of the participating hospitals. The material was processed and analyzed at the University Hospital Maastricht. Analysis included light microscopy, immunohistochemistry, and electron microscopy of the biopsies as well as serologic and chemical analysis. RESULTS: Primary IgA nephropathy (IgAN), membranous glomerulopathy, antineutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis and thin basement membrane nephropathy (TBMN) are the most common primary glomerular diseases in this order of sequence. Our data show the clinical and histologic phenotype of TBMN to be diverse: the vast majority of TBMN has chronic microscopic hematuria, frequently associated with hypertension in late middle age; about 15% of TBMN has in addition substantial proteinuria which is associated in the majority of cases with the lesions of focal segmental glomerulosclerosis (FSGS). In 5% of TBMN a nephrotic syndrome is observed, occasionally associated with FSGS tip lesions. CONCLUSION: These results support the notion that TBMN is a disease of genetic heterogeneity; it is not a benign renal condition in a substantial number of patients, particularly those in late middle age.

Susceptibility to cyclosporin A-induced autoimmunity: strain differences in relation to autoregulatory T cells.

Cyclosporin A-induced autoimmunity (CsA-AI), also called autoimmune syngeneic graft-vs-host disease, is a thymus dependent, T cell mediated rodent animal model of disease and is considered to be an experimental model for human scleroderma. Since adoptive transfer of CsA-AI by effector T cells can be prevented by autoregulatory T cells, there may also be a role for dominant tolerance in the resistance of certain rat strains to develop clinical manifest CsA-AI. LEW rats have been reported to be susceptible, whereas BN rats are resistant to CsA-AI. In the present study we first demonstrate that PVG, but not DA rats, are susceptible to CsA-AI and that disease characteristics in PVG rats are comparable to LEW rats in terms of pathogenesis and T cell kinetics, although of more rapid onset and greater severity. Next, we examined whether the relative presence of autoregulatory T-helper cells, i.e. CD25+ and/or CD45RClow CD4 T cells, is increased in resistant BN and DA rats. The results obtained reveal that the genetically determined CD45RChigh/CD45RClow ratio, but not the percentage CD25+ cells, within the CD4 T cell compartment of naïve rats is correlated with resistance to CsA-AI in these rat strains. We conclude that the relative presence of autoregulatory T cells with a CD45RClow T-helper cell phenotype may be a critical determinant in susceptibility to CsA-AI.

The CD8 T cell compartment plays a dominant role in the deficiency of Brown-Norway rats to mount a proper type 1 immune response.

Differential cytokine production by T cells plays an important role in regulating the nature of an immune response. In the rat, Brown-Norway (BN) and Lewis (LEW) strains differ markedly in their susceptibility to develop either type 1 or type 2-mediated autoimmune manifestations. BN rats are susceptible to type 2-dependent systemic autoimmunity, while LEW rats are resistant. Conversely, type 1-mediated, organ-specific autoimmune disease can be easily induced in LEW, but not in BN, rats. The mechanisms involved in the differential development of type 1 and type 2 immune responses by these two strains are still unknown. In the present study we analyzed the contributions of APC, CD4 and CD8 T cells, and MHC molecules in the difference between LEW and BN rats to develop a type 1 immune response. First, we show that the defect of BN T cells to produce type 1 cytokines in vitro does not require the presence of APC and, by using an APC-independent stimulation assay, we have localized the defect within the T cell compartment. Both CD4 and CD8 T cells are involved in the defect of BN rats to develop a type 1 immune response with a major contribution of the CD8 T cell compartment. This defect is associated with an increase in the type 2 cytokine IL-4 in both BN T cell populations, but neutralization of this cytokine does not restore this defect. Finally, by using MHC congenic rats, we show that the MHC haplotype is not involved in the defect of BN T cells to mount a proper type 1 cytokine response.