MCTD Patients Research Articles

Characterization of an HLA-DR4-restricted T cell clone recognizing a protein moiety of small nuclear ribonucleoproteins (UsnRNP).

In sera of patients with mixed connective tissue disease (MCTD, Sharp Syndrome) high titres of IgG autoantibodies to U1snRNP-specific proteins are found. The isolated occurrence of these autoantibodies is highly associated with the HLA-DR4 haplotype. snRNP-specific T cells are supposed to be involved in this autoantibody production. To address this question we cultured mononuclear cells from MCTD patients and healthy donors with a highly purified UsnRNP preparation from HeLa cells using bulk or limiting dilution cultures. Secondary responses to snRNP were detected only rarely with T cell lines from two patients and two controls, and turned out to be unstable during further expansion. One T cell line derived from a healthy individual retained its snRNP reactivity upon limiting dilution cloning and could be characterized in detail. The CD4+ T cell clone recognized native snRNP particles presented by monocytes in an HLA-DR4 (B1*0401)-restricted manner. Separation of the protein and RNA moieties of snRNP particles revealed that the T cell clone responded specifically to the protein fraction, but not to RNA and diverse control antigens. Sequencing of the T cell receptor alpha and beta chain cDNAs revealed that the clone used the V alpha 14.2 and V beta 14 elements. Upon antigen-specific and mitogenic stimulation the T cell clone showed a Th1-specific cytokine pattern, and did not provide helper activity for in vitro immunoglobulin production. This study demonstrate the presence of self-reactive snRNP-specific T cells in a healthy donor. The T cell clone may not represent a helper T cell for the formation of U1snRNP-specific autoantibodies.

CD4+ T Cells Target Epitopes Residing within the RNA-Binding Domain of the U1-70-kDa Small Nuclear Ribonucleoprotein Autoantigen and Have Restricted TCR Diversity in an HLA-DR4-Transgenic Murine Model of Mixed Connective Tissue Disease

Mixed connective tissue disease (MCTD) is a systemic autoimmune disease with significant morbidity and premature mortality of unknown pathogenesis. In the present study, we characterized U1-70-kDa small nuclear ribonucleoprotein (70-kDa) autoantigen-specific T cells in a new murine model of MCTD. These studies defined 70-kDa-reactive T cell Ag fine specificities and TCR gene usage in this model. Similar to patients with MCTD, CD4(+) T cells can be readily identified from 70-kDa/U1-RNA-immunized HLA-DR4-transgenic mice. Using both freshly isolated CD4(+) T cells from spleen and lung, and T cell lines, we found that the majority of these T cells were directed against antigenic peptides residing within the RNA-binding domain of 70 kDa. We also found that TCR-beta (TRB) V usage was highly restricted among 70-kDa-reactive T cells, which selectively used TRBV subgroups 1, 2, 6, 8.1, 8.2, and 8.3, and that the TRB CDR3 had conserved sequence motifs which were shared across different TRBV subgroups. Finally, we found that the TRBV and CDR3 regions used by both murine and human 70-kDa-specific CD4(+) T cells were homologous. Thus, T cell recognition of the 70-kDa autoantigen by HLA-DR4-transgenic mice is focused on a limited number of T cell epitopes residing primarily within the RBD of the molecule, using a restricted number of TRBV and CDR3 motifs that are homologous to T cells isolated from MCTD patients.

MCTD – Mixed Connective Tissue Disease

Mixed connective tissue disease is a disease entity characterized by overlapping symptoms of lupus erythematosus (LE), systemic sclerosis (SSc), polymyositis/dermatomyositis (PM/DM) and rheumatoid arthritis (RA). Diagnostic criteria include high titers of antibodies against U1RNP as well as the presence of at least 3 of 5 of the following clinical features: edema of hands, synovitis, myositis, Raynaud phenomenon and acroscierosis. In terms of the pathogenesis, genetic as well as infectious (viral) factors appear to play a role. The acceptance of MCTD as a distinct disease entity is controversial. Terms such as "undifferentiated connective tissue disease" or "overlapping syndromes" are not helpful. One-quarter of MCTD patients transform into LE, while one-third progress to SSc. Therapeutic recommendations are glucocorticoids in combination with immunosuppressive agents and endothelin receptor antagonists. Double blind studies are not available. The prognosis is relatively good. Causes of death include pulmonary hypertension, infections and both pulmonary and cardiac failure.

Synthesis of a Proline‐Rich [60]Fullerene Peptide with Potential Biological Activity.

Abstract A proline-rich [60]fullerene peptide was synthesized by use of (i) a 1,3-dipolar cycloaddition of an N-substituted glycine derivative to [60]fullerene, (ii) esterification of the isolated alcohol with the C-terminal amino acid of the desired peptide sequence, and finally (iii) coupling of the remaining hexapeptide to give the final product 8 as a TFA salt, with oxidized methionine. Product 8 was found to be biologically active against sera from MCTD and SLE patients (ELISA experiment).

Synthesis of a proline-rich [60]fullerene peptide with potential biological activity

A proline-rich [60]fullerene peptide was synthesized by use of (i) a 1,3-dipolar cycloaddition of an N-substituted glycine derivative to [60]fullerene, (ii) esterification of the isolated alcohol with the C-terminal amino acid of the desired peptide sequence, and finally (iii) coupling of the remaining hexapeptide to give the final product 8 as a TFA salt, with oxidized methionine. Product 8 was found to be biologically active against sera from MCTD and SLE patients (ELISA experiment).

Anti‐U1RNP antibody and aseptic meningitis in connective tissue diseases

Objective: To investigate the relationship between aseptic meningitis and anti‐U1RNP antibody in patients diagnosed with CTD.Methods: Fourteen patients with aseptic meningitis were selected from among patients with CTDs who had visited our hospital. We analyzed their medical records to clarify the clinical and immunological features of aseptic meningitis.Results: A total of 14 patients with aseptic meningitis were subsequently diagnosed as having either SLE (seven cases), MCTD (four), UCTD (one), overlap syndrome (one), or Sjögren's syndrome (one). Eight of the 14 patients had received NSAIDs, such as sulindac, naproxen, or loxoprofen, before the onset of aseptic meningitis. CRP levels were increased (mean±SD: 7.1±7.1mg/dL) and CRP levels (10.4±7.7) in the drug‐induced group were significantly increased (p<0.01). The anti‐U1RNP antibody was found in 13 of the 14 patients. There were no significant differences in cerebrospinal fluid findings between the drug‐induced group and the non‐drug‐induced group.Conclusions: SLE or MCTD patients with aseptic meningitis tend to have anti‐U1RNP antibody.

Pulmonary hypertension in connective tissue disease. Clinical analysis of sixty patients in multi-institutional study

Clinical features and prognosis of sixty patients with connective tissue disease accompanied by pulmonary hypertension (PH) (26 MCTD, 20 SLE, and 14 PSS) reported retrospectively by multi-institutions were compared. Though the obtained data were incomplete and lacking in uniformity, no significant difference in the clinical features among the three diseases were observed except high incidence of pulmonary fibrosis and low % VC in PSS and PH patients. Statistically significant difference, however, was observed between live and dead patients of three diseases gathered in post sternal pain, pulmonary diastolic murmur, right ventricular hypertrophy on ECG and mean pressure of pulmonary artery. Higher incidence of anti-nRNP antibody was observed in SLE with PH and PSS with PH patients than with the general population. A quicker occurrence of PH and shorter survival time were observed in MCTD patients with PH than in SLE and PSS patients with PH.