SUMMARY In sera of patients with mixed connective tissue disease (MCTD) high titres of IgG autoantibodies to U1snRNP‐specific proteins (70kD, A, C) are found, suggesting an antigen‐driven and T cell‐dependent process. In order to establish U1snRNP‐specific T cell lines we cultured under various culture conditions mononuclear cells from MCTD patients and healthy donors with a highly purified UsnRNP preparation from HeLa cells. Nine T cell lines were established by limiting dilution cloning from two MCTD patients and five T cell lines from a healthy individual. All T cell lines expressed the TCRαβ/CD3 complex. Surprisingly, most of the T cell lines exhibited the CD8 phenotype. Irrespective of this phenotype, all T cell lines showed a proliferative response to an N‐terminal pan (aa 51–195) of recombinant U1‐specific 70‐kD protein. One CD8+ T cell clone exhibited cytotoxic activity against an autologous B cell line pulsed with snRNP or recombinant fragments (aa 51–195 and aa 51–88). Interestingly, two T cell lines proliferated in response to four recombinant polypeptides representing different parts of the U1snRNP 70‐kD protein. Since regions of sequence homology are distributed over the 70‐kD molecule, it is suggested that conserved motifs may be recognized by the T cell lines.

SUMMARYIn sera of patients with mixed connective tissue disease (MCTD, Sharp Syndrome) high titres of IgG autoantibodies to UlsnRNP-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 a and ,B chain cDNAs revealed that the clone used the Val4.2 and VP 14 elements. Upon antigen-specific and mitogenic stimulation the T cell clone showed a Thl-specific cytokine pattern, and did not provide helper activity for in vitro immunoglobulin production. This study demonstrates 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 UlsnRNP-specific autoantibodies.

No abstract