Increased levels of BLyS and sVCAM-1 in anti-neutrophil cytoplasmatic antibody (ANCA)-associated vasculitides (AAV).

OBJECTIVES: Anti-neutrophil antibodies (ANCA)-associated vasculitides (AAV) comprise different forms of small vessel vasculitis characterised by B-cell driven autoimmune processes and endothelial cell activation. Aim of this study was to correlate markers of B- and endothelial cell activation with clinical manifestations of disease in AAV. METHODS: Consecutive serum samples of patients fulfilling the Chapel Hill Consensus Conference (CHCC) and American College of Rheumatology (ACR) criteria for AAV and healthy donors were used for the determination of ANCA, B-lymphocyte stimulator (BLyS), soluble vascular cell adhesion molecule-1 (sVCAM-1) and soluble E-selectin (sE-selectin) levels using enzyme-linked immunosorbent assay (ELISA). Subset and follow-up analyses were performed in cytoplasmatic ANCA (C-ANCA) or perinuclear ANCA (P-ANCA) positive patients with respect to change in ANCA-titres during the course of disease. RESULTS: Levels of sVCAM-1 were elevated in all patient groups with vasculitis compared to healthy controls. In contrast, significantly increased levels of BLyS were only observed in patients with Wegener's granulomatosis (WG), but not in patients with microscopic polyangiitis (mPAN)/Churg-Strauss-syndrome (CSS). Remarkably, there were no differences in the levels of sE-selectin between the vasculitis groups and healthy controls. In follow-up analysis, a significant correlation was shown for sE-Selectin and P-ANCA titres as well as sVCAM-1 levels. Furthermore, a strong correlation was detected for sVCAM-1 and creatinine levels. Interestingly, sE-selectin levels and C-ANCA titres were negatively correlated. CONCLUSIONS: Enhanced levels of sVCAM-1 represent a marker for endothelial cell activation in AAV. The observed correlation between sVCAM-1 and creatinine levels might indicate the influence of the vasculitic process on renal function. Signalling pathways for B-cells provided by BLyS could play a significant role in the pathogenesis of WG.

Effects of dietary energy intake during gestation and lactation on milk yield and composition of first, second and fourth parity sows.

In order to determine the effects of a varied level of dietary energy intake during pregnancy and lactation on milk yield and composition, first, second and fourth parity sows (Large White x German Landrace) were provided with energy at a level of either: (i) 100% of ME requirement (MEreq) during pregnancy and lactation, (ii) 120% MEreq during pregnancy and 80% during lactation, and (iii) 80% MEreq during pregnancy and 120% during lactation. In spite of equal target levels feed analysis revealed that gestating first parity sows with 120/80 treatment combination and lactating sows of 80/120 treatment combination received 25, and 11-17% more digestible N than in the respective 100/100 treatment combination. Irrespective of this 120/80 sows responded with the highest milk DM, fat, and energy contents, and the lowest lactose concentrations whereas protein levels where not affected, irrespective of parity (p < 0.05). Milk yield of sows in 1st and 4th lactation was 85 and 106% of that in 2nd lactation, respectively. Average milk composition was 18.1% DM, 4.9% protein, 6.8% fat, 5.6% lactose, and 0.8% ash. Milk composition changes ceased at day 7 of lactation with a reduction of milk GE and protein, and an increase of lactose content. Concentrations of threonine, arginine, valine, leucine, tyrosine, phenylalanine, cystine, and tryptophan, as well as stearic, oleic, and linoleic acid were higher in colostrum than in milk at later lactation stages. In contrast, laurine, myristic, palmitic, and palmitoleic acids were lower concentrated in colostrum. In conclusion, these results illustrate the importance of body reserve mobilization for milk production in sows and indicate that low energy supply during gestation cannot be compensated by higher energy supply during lactation.

Mutation and citrullination modifies vimentin to a novel autoantigen for rheumatoid arthritis.

OBJECTIVE: Modification of antigens represents a trigger for the generation of autoantibodies. In the pathogenesis of rheumatoid arthritis (RA), citrullination of proteins has been shown to be a critical process, and the determination of antibodies against citrullinated antigens has been a diagnostic milestone. We undertook this study to determine whether antibodies to mutated and citrullinated vimentin (MCV) could serve as a diagnostic and prognostic marker for RA. METHODS: We identified novel isoforms of human MCV in the synovial fluid of RA patients. The significance of these disease-related modifications was investigated by the analysis of autoantibody reactivities. In a group of 1,151 RA patients, the diagnostic significance and the prognostic value of an anti-MCV enzyme-linked immunosorbent assay (ELISA) were compared with that of an anti-cyclic citrullinated peptide (anti-CCP) ELISA. RESULTS: In RA, sensitivities of 82% and 72% were calculated for the anti-MCV and anti-CCP assays, respectively. The specificity of both assays was comparable (98% and 96%, respectively). In followup analyses of 16 RA patients with moderate disease activity (mean Disease Activity Score in 28 joints [DAS28] of 2.72) and 26 RA patients with active disease (mean DAS28 of 5.07), disease stratification of RA was possible using the anti-MCV assay (P = 0.0084). A significant correlation of anti-MCV antibodies with the DAS28 was documented (r = 0.5334, P = 0.0003), in 42 RA patients (n = 427 antibody determinations at different time points). CONCLUSION: Antigenic properties of vimentin were determined by mutation and citrullination. Anti-MCV antibodies are a novel diagnostic marker for RA. Furthermore, they may allow monitoring and-if confirmed in even larger series of patients-stratification of disease.

Tissue-specific up-regulation of the proteasome subunit beta5i (LMP7) in Sjögren's syndrome.

OBJECTIVE: Sjögren's syndrome (SS) is characterized by autoimmune infiltration and focal accumulation of lymphocytes in the exocrine glands, with a predominance of CD4-positive T cells. Since these histologic findings are nonspecific, determination of clinical and serologic abnormalities contribute to the diagnosis. The aim of this study was to identify a novel, disease-specific, immunologically relevant marker for SS. METHODS: To analyze disease-related and tissue-specific expression of candidate markers, we examined biopsied minor salivary glands and peripheral blood mononuclear cells from patients with primary and secondary SS (n = 26) as well as from patients with sicca symptoms without autoimmune sialadenitis (n = 15). Expression of the Th1/Th2-related chemokines CCL3 (macrophage inflammatory protein 1alpha) and CCL2 (monocyte chemoattractant protein 1), CXCL7 (neutrophil-activating peptide 2 [NAP-2]), interleukin-1beta, inducible costimulator, and the proteasome subunits alpha3 (C9) and beta5i (LMP7) was analyzed at the messenger RNA (mRNA) level using real-time polymerase chain reaction techniques. Immunohistochemical analysis was used to identify the beta5i (LMP7)-expressing cell populations in minor salivary glands. RESULTS: The expression profiles revealed a significant up-regulation of beta5i (LMP7) exclusively in the salivary glands of SS patients. Immunohistochemistry confirmed expression of the immunoproteasome subunit beta5i (LMP7) within the acinar and ductal epithelial cells. No significant difference in the distinct histologic focus scores was evident for the expression of the markers investigated. In the peripheral blood compartment, the expression of CXCL7 was up-regulated both in primary and in secondary SS. CONCLUSION: Tissue-specific up-regulation of beta5i (LMP7) mRNA was shown to be characteristic of SS, indicating a disease-specific modulation of the proteasome system. Expression of beta5i (LMP7) represents an independent parameter that can be used in addition to the focus score to distinguish SS in biopsied labial salivary glands.

Circulating proteasomes are markers of cell damage and immunologic activity in autoimmune diseases.

OBJECTIVE: The 20S proteasome plays a leading immunologic role in the cytosolic generation of MHC class I restricted antigens, and it represents an abundant antigen in several autoimmune diseases. To investigate the effects of autoimmune inflammatory and perioperative traumatic cellular damage, we determined qualitative and quantitative properties of released proteasomes (circulating proteasomes, cProteasomes) from serum samples of patients with a variety of autoimmune diseases. METHODS: cProteasomes were analyzed from serum samples of 314 patients with several systemic and organ-specific autoimmune diseases and 85 healthy controls. The concentrations of cProteasomes were determined by sandwich ELISA using a monoclonal and a polyclonal proteasome-specific antibody. Followup analyses were performed in patients with systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) as well as in patients with myasthenia gravis undergoing thoracoscopic thymectomy. RESULTS: Strongly increased levels of cProteasomes (> 1000 ng/ml) were detected in samples obtained from patients with autoimmune myositis, SLE, primary Sjögren's syndrome, RA, and autoimmune hepatitis. Significant differences were observed in the mean values of cProteasomes comparing systemic with organ-specific autoimmune diseases. Followup analyses revealed a close correlation of cProteasome with the autoimmune process as well as cellular damage. Moreover, cProteasomes were isolated in intact and native as well as in degraded or dissociated forms from the serum samples. The immuno-subunit LMP7 was found to be incorporated in the circulating protease complex. CONCLUSION: Levels of cProteasomes are markedly elevated in patients with systemic autoimmune diseases, apparently correlating with disease activity. The cProteasomes represent novel sensitive markers of the autoimmune inflammatory processes and/or reflect the magnitude of cellular damage.