IDENTIFICATION OF T-CELL EPITOPES ON THE RHESUS POLYPEPTIDES IN AUTOIMMUNE HEMOLYTIC-ANEMIA

We have shown previously that the Rhesus (Rh) polypeptides are the com monest targets for pathogenic anti-red blood cell (RBC) autoantibodies in patients with autoimmune hemolytic anemia (AIHA). The aim of the c urrent work was to determine whether activated T cells from such patie nts also mount recall responses to epitopes on these proteins. Two pan els of overlapping 15-mer peptides, corresponding to the sequences of the 30-kD ph proteins associated with expression of the D and Cc/Ee bl ood group antigens, were synthesized and screened for the ability to s timulate the in vitro proliferation of mononuclear cells from the peri pheral blood or spleen of nine AIHA cases. Culture conditions were cho sen that favor recall proliferation by previously activated T cells, r ather than primary responses. In seven of the patients, including all four cases with autoantibody to the ph proteins, two or more peptides elicited proliferation, but cells from eight of nine patients with oth er anemias and seven of nine healthy donors failed to respond to the p anels. Multiple peptides were also stimulatory in two positive control donors who had been alloimmunized with ph D-positive RBCs. Six differ ent profiles of peptides elicited responses in the AIHA patients, and this variation may reflect the different HLA types in the group. Stimu latory peptides were identified throughout domains shared between, or specific to, each of the related 30-kD Rh proteins, but T cells that r esponded to nonconserved regions did not crossreact with the alternati ve sequences. Anti-major histocompatibility complex class tl antibodie s blocked the responses and depletion experiments confirmed that the p roliferating mononuclear cells were T cells. Notably, splenic T cells that proliferated against multiple Rh peptides also responded to intac t RBCs. We propose that pathogenic autoantibody production in many cas es of AIHA is driven by the activation of T-helper cells specific for previously cryptic epitopes on the Rh proteins. (C) 1997 by The Americ an Society of Hematology.