Influence of ethnic background on clinical and serologic features in patients with systemic sclerosis and anti-DNA topoisomerase I antibody.
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OBJECTIVE: To investigate the effect of ethnicity on clinical and serologic expression in patients with systemic sclerosis (SSc) and anti-DNA topoisomerase I (anti-topo I) antibody. METHODS: Clinical and serologic features, as well as HLA class II allele frequencies, were compared among 47 North American white, 15 North American black, 43 Japanese, and 12 Choctaw Native American SSc patients with anti-topo I antibody. RESULTS: The frequency of progressive pulmonary interstitial fibrosis was lower, and cumulative survival rates were better in white compared with black and Japanese patients. Sera of white and black patients frequently recognized the portion adjacent to the carboxyl terminus of topo I, sera of Japanese patients preferentially recognized the portion adjacent to the amino terminus of topo I, and sera of Choctaw patients recognized both portions of topo I. Anti-RNA polymerase II and anti-SSA/Ro antibodies were present together with anti-topo I antibody more frequently in sera of Japanese patients than in sera of white patients. The HLA-DRB1 alleles associated with anti-topo I antibody differed; i.e., DRB1*1101-*1104 in whites and blacks, DRB1*1502 in Japanese, and DRB1*1602 in Choctaws. Multivariate analysis showed that ethnic background was an independent determinant affecting development of severe lung disease as well as survival. CONCLUSION: Clinical and serologic features in SSc patients were strongly influenced by ethnic background. The variability of disease expression in the 4 ethnic groups suggests that multiple factors linked to ethnicity, including genetic and environmental factors, modulate clinical manifestations, disease course, and autoantibody status in SSc. (+info)
Immune responses to cartilage link protein and the G1 domain of proteoglycan aggrecan in patients with osteoarthritis.
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OBJECTIVE: To determine whether patients with osteoarthritis (OA) express cellular immunity to cartilage link protein (LP) and the G1 globular domain of proteoglycan (PG) aggrecan, and whether immunity to the G1 domain is influenced by the removal of keratan sulfate (KS). METHODS: LP and the G1 globular domain of PG were isolated from human and/or bovine cartilage and used in proliferation assays with peripheral blood lymphocytes (PBL) from 42 patients with OA and 40 healthy control subjects. RESULTS: Patients with OA expressed a higher prevalence of cellular immunity to human cartilage LP (42.4%) compared with the control group (13.3%). The prevalence of immune reactivity to bovine LP in patients with OA was lower (35.7%) compared with the immunity to human LP, but remained similar in the control group (13.8%). PBL from patients with OA exhibited low reactivity to the native G1 domain of bovine PG. However, removal of KS chains from the G1 globular domain resulted in increased cellular immune responses to the G1 domain in OA patients (45.8%) compared with the control group (7.7%). CONCLUSION: These results indicate the presence of immunity to cartilage-derived LP and the G1 globular domain of PG aggrecan in patients with OA and the inhibitory effect of KS chains on the G1 domain on the expression of this immunity in OA patients. This immune reactivity is commonly observed in patients with inflammatory joint disease and can experimentally induce arthritis. Thus, it may be involved in the pathogenesis of OA. (+info)
Changes in joint cartilage aggrecan after knee injury and in osteoarthritis.
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OBJECTIVE: To determine the concentrations of aggrecan fragments in synovial fluid from patients with knee joint injury, osteoarthritis (OA), or acute pyrophosphate arthritis (PPA; pseudogout), and to test their relative reactivity with the 846 epitope, a putative marker of cartilage aggrecan synthesis. METHODS: Samples of knee joint fluid from 385 patients and 9 healthy-knee volunteers were obtained in a cross-sectional study. Study groups were acute PPA/ pseudogout (n = 60), anterior cruciate ligament (ACL) rupture (n = 159), meniscus lesion (n = 129), and primary knee OA (n = 37). The 846 epitope on aggrecan was assayed by competitive solution-phase radioimmunoassay. Aggrecan fragments were assayed by enzyme-linked immunosorbent assay using a monoclonal antibody (1-F21). Cartilage oligomeric matrix protein (COMP), C-propeptide of type II collagen (CPII), bone sialoprotein, matrix metalloproteinases 1 and 3, and tissue inhibitor of metalloproteinases 1 were previously quantified by immunoassays. RESULTS: Reactivity of the 846 epitope was increased in all study groups compared with the reference group, and was highest in patients with primary OA. The median levels (in microg fetal aggrecan equivalents/ml) of the epitope were 0.28 (range 0.24-0.47) in the reference group, 0.48 (range 0.26-1.32) in PPA/pseudogout, 0.61 (range 0.12-2.87) in ACL rupture, 0.53 (range 0.22-3.02) in meniscus lesion, and 0.68 (range 0.31-4.31) in primary OA. The 846 epitope reactivity per microg aggrecan fragments in the joint fluid was higher in late-stage OA than in early-stage OA. Epitope 846 reactivity correlated positively with several markers of matrix turnover, particularly with COMP (r(s) = 0.421) and CPII (r(s) = 0.307). CONCLUSION: The observed differences in 846 epitope reactivity in synovial fluid, and its concentration in relation to aggrecan and other markers of matrix turnover, were consistent with marked ongoing changes in aggrecan turnover after joint injury and in the development of OA. OA is thus a disease characterized by dynamic changes in tissue macromolecule turnover, which is reflected by measurable changes in aggrecan epitopes in the synovial fluid. (+info)
CFTR expression does not influence glycosylation of an epitope-tagged MUC1 mucin in colon carcinoma cell lines.
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The cause of the mucus clearance problems associated with cystic fibrosis remains poorly understood though it has been suggested that mucin hypersecretion, dehydration of mucins, and biochemical abnormalities in the glycosylation of mucins may be responsible. Since the biochemical and biophysical properties of a mucin are dependent on O-glycosylation, our aim was to evaluate the O-glycosylation of a single mucin gene product in matched pairs of cells that differed with respect to CFTR expression. An epitope-tagged MUC1 mucin cDNA (MUC1F) was used to detect variation in mucin glycosylation in stably transfected colon carcinoma cell lines HT29 and Caco2. The glycosylation of MUC1F mucin was evaluated in matched pairs of Caco2 cell lines that either express wild-type CFTR or have spontaneously lost CFTR expression. The general glycosylation pattern of MUC1F was evaluated by determining its reactivity with a series of monoclonal antibodies against known blood group and tumor-associated carbohydrate antigens. Metabolic labeling experiments were used to estimate the gross levels of glycosylation and sulfation of MUC1F mucin in these matched pairs of cell lines. Expression of CFTR in this experimental system did not affect the gross levels of glycosylation or sulfation of the MUC1F mucin nor the types of carbohydrates structures attached to the MUC1F protein. (+info)
Antifactor VIII antibody inhibiting allogeneic but not autologous factor VIII in patients with mild hemophilia A.
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Two unrelated patients with the same Arg2150His mutation in the factor VIII (FVIII) C1 domain, a residual FVIII activity of 0.09 IU/mL, and inhibitor titres of 300 and 6 Bethesda Units, respectively, were studied. Further analysis of patient LE, with the highest inhibitor titer, showed that (1) plasma or polyclonal IgG antibodies prepared from LE plasma inhibited the activity of allogeneic (wild-type) but not of self FVIII; (2) the presence of von Willebrand factor (vWF) increased by over 10-fold the inhibitory activity on wild-type FVIII; (3) the kinetics of FVIII inhibition followed a type II pattern, but in contrast to previously described type II inhibitors, LE IgG was potentiated by the presence of vWF instead of being in competition with it; (4) polyclonal LE IgG recognized the FVIII light chain in enzyme-linked immunosorbent assay and the recombinant A3-C1 domains in an immunoprecipitation assay, indicating that at least part of LE antibodies reacted with the FVIII domain encompassing the mutation site; and (5) LE IgG inhibited FVIII activity by decreasing the rate of FVIIIa release from vWF, but LE IgG recognized an epitope distinct from ESH8, a murine monoclonal antibody exhibiting the same property. We conclude that the present inhibitors are unique in that they clearly distinguish wild-type from self, mutated FVIII. The inhibition of wild-type FVIII by LE antibody is enhanced by vWF and is associated with an antibody-dependent reduced rate of FVIIIa release from vWF. (+info)
Conformational aspects of HIV-1 integrase inhibition by a peptide derived from the enzyme central domain and by antibodies raised against this peptide.
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Monospecific antibodies were raised against a synthetic peptide K159 (SQGVVESMNKELKKIIGQVRDQAEHLKTA) reproducing the segment 147-175 of HIV-1 integrase (IN). Synthesis of substituted and truncated analogs of K159 led us to identify the functional epitope reacting with antibodies within the C-terminal portion 163-175 of K159. Conformational studies combining secondary structure predictions, CD and NMR spectroscopy together with ELISA assays, showed that the greater is the propensity of the epitope for helix formation the higher is the recognition by anti-K159. Both the antibodies and the antigenic peptide K159 exhibited inhibitory activities against IN. In contrast, neither P159, a Pro-containing analog of K159 that presents a kink around proline but with intact epitope conformation, nor the truncated analogs encompassing the epitope, were inhibitors of IN. While the activity of antibodies is restricted to recognition of the sole epitope portion, that of the antigenic K159 likely requires interactions of the peptide with the whole 147-175 segment in the protein [Sourgen F., Maroun, R.G., Frere, V., Bouziane, A., Auclair, C., Troalen, F. & Fermandjian, S. (1996) Eur. J. Biochem. 240, 765-773]. Actually, of all tested peptides only K159 was found to fulfill condition of minimal number of helical heptads to achieve the formation of a stable coiled-coil structure with the IN 147-175 segment. The binding of antibodies and of the antigenic peptide to this segment of IN hampers the binding of IN to its DNA substrates in filter-binding assays. This appears to be the main effect leading to inhibition of integration. Quantitative analysis of filter-binding assay curves indicates that two antibody molecules react with IN implying that the enzyme is dimeric within these experimental conditions. Together, present data provide an insight into the structure-function relationship for the 147-175 peptide domain of the enzyme. They also strongly suggest that the functional enzyme is dimeric. Results could help to assess models for binding of peptide fragments to IN and to develop stronger inhibitors. Moreover, K159 antibodies when expressed in vivo might exhibit useful inhibitory properties. (+info)
Distribution of B-cell epitopes on the pseudorabies virus glycoprotein B.
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In order to map antigenically important regions of glycoprotein B (gB) of pseudorabies virus (PrV), a panel of recombinant fragments of gB expressed in E. coli and truncated fragments of gB generated by cleavage of purified native gB with trypsin and cyanogen bromide was analysed by using 26 monoclonal antibodies directed against gB. Three continuous epitopes were localized in the vicinity of the N terminus of gB, between amino acids (aa) 59 and 126. One continuous epitope mapped between residues 214 and 279. The residues involved in the assembly of eight discontinuous epitopes were located between aa 540 and 734. The constituents of two discontinuous epitopes were harboured in a segment encompassing aa 540-646. The clustering of continuous epitopes at the extreme N terminus of PrV gB and the locations of residues involved in the assembly of discontinuous epitopes of PrV gB are in good agreement with data on epitope locations in gB homologues from other herpesviruses. (+info)
Immunogenicity of variable regions of hepatitis C virus proteins: selection and modification of peptide epitopes to assess hepatitis C virus genotypes by ELISA.
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The immunogenicity of variable regions of hepatitis C virus (HCV) proteins was studied by ELISA by using 543 synthetic peptides from 120 variable regions and 90 sera from HCV-infected patients. Some regions from certain genotypes were less immunogenic, or even non-immunogenic, compared with their equivalents in other genotypes. However, the mean recognition of all peptides from genotypes 1a, 1b and 3 by sera infected with genotypes 1a, 1b and 3, respectively, showed no significant differences, suggesting a similar overall immunogenicity of variable regions from these genotypes. Proteins NS4a, NS4b and NS5a were found to be the most immunogenic. Recognition of individual peptides by the sera of infected patients showed that the humoral response against HCV is patient-dependent. The work shows that 15-mer peptides may encompass several B-cell epitopes. These epitopes may lie in slightly different positions in different genotypes. Thirty-one percent of the 543 peptides were recognized by some of the 35 healthy donors. This may be a reflection of the large number of antigens to which they had been exposed, but it may also reflect a strategy of HCV to respond to immune pressure. After selection and modification, a set of 40 peptides was used to assess genotypes 1a, 1b, 1, 2 and 3 in the sera of HCV-infected patients, with sensitivities of 34.1, 48.5, 68.8, 58.3 and 48.9% and specificities of 100, 99.1, 97.1, 99.5 and 99%, respectively. The overall sensitivity and specificity for the assessment of genotypes 1, 2 and 3 were 64 and 98%, respectively. (+info)