Impaired deoxyribonuclease activity in monoglandular and polyglandular autoimmunity.

OBJECTIVE: The enzyme desoxyribonuclease (DNase) degrades DNA during early apoptosis. Impaired DNase activity might increase susceptibility to autoimmune diseases. This study examined for the first time DNase activity in endocrine autoimmunity. METHODS: Included were 112 patients with monoglandular (MGA) or polyglandular autoimmunity (PGA), their 93 healthy relatives, and 41 healthy controls. Serum DNase activity was quantified with a solid phase enzyme immunometric assay comprising degradation of the specific immobilized DNase substrate, formation of enzyme conjugate complexes using horseradish peroxidase conjugate solution, and enzymatic colour reaction. RESULTS: The Bland-Altman plot of the interassay differences suggested good reproducibility (n=96). Compared to healthy controls (median 9.8, range 5.2-16.7 ng/ml), DNase activity was markedly lowered in patients with endocrine autoimmunity (5.8, 2.6-26.2 ng/ml; p<0.0001). Corresponding values in the following MGA, PGA, and relatives groups were 4.8 (2.8-19.0) ng/ml, 7.9 (2.6-26.2) ng/ml, and 8.4 (1.5-19.0) ng/ml, respectively. When MGA patients were splitted up by disease, patients with type 1 diabetes had the lowest DNase activity (3.6, 3.2-3.9 ng/ml) which positively correlated with HbA1c in females (r=0.486, p=0.041). Pathological reduction of DNase activity (below 5 ng/ml) was noted in 54%, 31%, 24%, and 0% of MGA, PGA, relatives, and controls, respectively. Anti-ds-DNA and anti-nucleosome antibodies were negative in the patients with MGA and PGA. CONCLUSIONS: These findings indicate the potential relevance of DNase activity in patients with monoglandular and polyglandular autoimmunity and their clinically healthy relatives. The impaired DNase activity might reduce removal of circulating self- or pathogen-derived DNA thereby favoring autoimmune mechanisms by Toll-like receptor 9 co-activation.

A novel high sensitivity ELISA for detection of antineutrophil cytoplasm antibodies against proteinase-3.

OBJECTIVE: Conventional direct enzyme-linked immunosorbent assays (ELISA) for the detection of anti-neutrophil cytoplasm antibodies (ANCA) often lack sensitivity because epitopes of the target antigen are hidden by binding to the ELISA plate. This study was designed to evaluate a novel ELISA method for detection of ANCA against proteinase-3 (PR3) for the diagnosis of Wegener's granulomatosis (WG) using PR3 presented in its native form. METHODS: Sera from four subgroups of patients with a diagnosis of WG (n=86), 80 healthy controls and 450 disease controls were tested for the presence of C-ANCA/PR3-ANCA by anchor ELISA, direct ELISA, capture ELISA, indirect immunofluorescence (IFT) and immunoblotting. RESULTS: In prospectively analysed consecutive patients, anchor ELISA showed the highest sensitivity for a diagnosis of WG of 96.0% (95% CI: 79.6-99.3), followed by IFT 92.0% (73.9-98.8), capture ELISA 72.0 (50.6-87.9) and direct ELISA 60.0 (38.7-78.8). Specificity was high for all methods and ranged from 98.5 (97.0-99.4) to 95.5% (97.9-99.8). Receiver operating characteristics curve analysis revealed that the overall diagnostic performance of the anchor ELISA was significantly superior compared to the direct ELISA and the capture ELISA in patients with generalized WG, and also compared to IFT and immunoblotting in patients with localised WG. CONCLUSION: Anchor ELISA is a novel highly sensitive and specific method for the detection of PR3-ANCA in patients with WG, which may replace the need for a combined analysis with IFT and ELISA in the future.