Identification of target antigens of antiendothelial cell antibodies in healthy individuals: A proteomic approach.
ABSTRACT In order to identify target antigens of anti-endothelial cell (anti-EC) antibodies (AECA) in healthy individuals, we have used a proteomic approach combining 2-DE and immunoblotting. Whole cell protein extracts obtained from human umbilical vein EC (HUVEC) cultures were used as a source of antigens. Serum IgG from 12 healthy blood donors were tested at a concentration of 200 microg/mL. Targeted spots were identified by MS. The HUVEC proteome was composed of 884 protein spots. Among these, 61 +/- 25.8 (mean +/- SD) spots were recognized by serum IgG from healthy individuals, with marked differences from one individual to another. Among these spots, 11 were recognized by serum IgG from all healthy individuals tested. These spots corresponded to six different proteins with several spots corresponding to different isoforms of the same protein. Target antigens were: cytoskeletal proteins (beta-actin, alpha-tubulin, and vimentin); glycolytic enzymes (glucose-3-phosphate-deshydrogenase and alpha-enolase); and prolyl-4-hydroxylase beta subunit, a member of the disulfide isomerase family. This study shows that the repertoire of IgG AECA is heterogeneous among healthy individuals. IgG from all of the healthy individuals tested recognized a restricted set of highly conserved ubiquitous proteins playing key roles in cell biology and maintenance of homeostasis.
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ABSTRACT: Susac syndrome is a rare disease that is characterised by the clinical triad of encephalopathy, branch retinal artery occlusion, and sensorineural hearing loss. It was first described as a distinctive syndrome by Susac in 1979. There have been 304 reported individual patients with Susac syndrome. Etiophatogenesis is not clear, although it is now thought that it is an immune-mediated endotheliopathy that affects the microvasculature of the brain, retina, and inner ear. Antiendothelial Cell Antibodies (AECAs) play an important role in mediating the endothelial cell injury with consequent deposition of thrombotic material in the lumen of the small vessel. In biopsies of the brain, microinfarcts with atrophy of the white and grey matter could be detected. These microinfarcts are caused by a microangiopathic process with arteriolar wall proliferation, lymphocytic infiltration and basal lamina thickening. At clinical onset, the most common manifestation was Central Nervous System symptoms, followed by visual symptoms and hearing disturbances. Diagnosis is based on Magnetic Resonance Imaging (MRI), retinal fluorescein angiography, and audiometry; these are considered crucial tests to enable diagnosis. Antiendothelial cell antibodies (AECAs) are also of diagnostic relevance. Based on the hypothesis of being an autoimmune disease, treatment has to be immunosuppressive. In addition, anticoagulation measures, antiplatelet agents and antivasospastic agents should be considered. The majority of patients did not initially present with the complete triad of symptoms. An appropriate approach would be to perform a search for absent components of the triad if the clinical presentation is suggestive of Susac syndrome. Improved understanding of the presentation of Susac syndrome, will prevent misdiagnosis and ensure that patients receive the best possible care.Autoimmunity reviews 04/2014; · 6.37 Impact Factor
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ABSTRACT: Hepatoportal sclerosis accompanied by dense elastic fibre deposition is generally regarded as the primary lesion in the development of idiopathic portal hypertension (IPH). This study was performed to clarify the mechanism of elastic fibre deposition in the peripheral portal tracts of IPH liver in relation to serum anti-endothelial cell antibodies (AECA). In-vitro experiments were performed using human dermal microvascular endothelial cells (HMVEC) and patients' sera. The presence of serum AECA was assayed by a cell-based enzyme-linked immunosorbent assay (ELISA) using HMVEC. Immunohistochemical analysis of elastin was performed using liver tissue sections of IPH patients. IPH sera contained one or more AECA that could bind to the vascular endothelial cells of the peripheral portal tracts of the liver. When the value of AECA greater than the mean ± 2 standard deviations of healthy controls was regarded as positive, the positive detection rate of either immunoglobulin (Ig)G, IgA or IgM AECA in IPH sera was 30% (10 of 33 cases). IPH sera induced the expression of elastin in HMVEC, which appeared to be associated with the presence of AECA. Apoptosis was also induced in HMVEC by the stimulation with IPH sera. In vivo, elastin expression was observed in the endothelial cells of the peripheral portal tracts of IPH livers in a proportion of cases. The disease pathogenesis of IPH seems to be heterogeneous, and this study elucidated a possible contribution of the induction of elastin expression in the portal vessels to hepatoportal sclerosis of IPH, which might be linked to serum AECA as a causative factor.Clinical & Experimental Immunology 03/2012; 167(3):532-42. · 3.41 Impact Factor
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ABSTRACT: Profiling the autoantibody repertoire with proteome-wide antigen collections is emerging as a powerful tool for the identification of biomarkers for autoimmune diseases. Here, a systematic and undirected approach was taken to screen for profiles of IgG in human plasma from 90 individuals with multiple sclerosis (MS) related diagnoses. Reactivity pattern of 11,520 antigens (corresponding to ~38% of all human protein encoding genes) were generated on planar protein microarrays built within the Human Protein Atlas. IgG reactivity was observed towards more than 2,000 antigens, among which 64% were recognized only in single individuals. Reactivity distributions among MS subgroups were used to select 384 antigens, which were then re-evaluated on planar microarrays, corroborated with suspension bead arrays in a larger cohort (n=376), and confirmed for specificity in inhibition assays. Among the heterogeneous pattern within and across MS subtypes, differences in recognition frequencies were found for 51 antigens, which were enriched for proteins of transcriptional regulation. In conclusion, a strategy on complementary high-throughput protein array platforms facilitated the discovery and verification of disease-associated autoimmunity signatures that are proposed as additional antigens for larger scaled validation across MS biobanks.Molecular & Cellular Proteomics 06/2013; · 7.25 Impact Factor