Oligoclonal Expansion of Vδ1+ γδT Cells in Cyclosporine-Responsive Acquired Pure Red Cell Aplasia.

Abstract

Background: Acquired pure red cell aplasia (PRCA) is an autoimmune disease in which autoaggressive T lymphocytes selectively suppress erythropoiesis, sometimes associated with clonal T cell disorders. However, little is known about the structure of the T-cell receptors (TCR) used by T lymphocytes involved in the development of acquired PRCA. Previously, a patient with granular lymphocyte-proliferative disorder with an expansion of Vδ1+ γδ T cells has been reported to be associated with cyclosporine-resistant acquired PRCA (Handgretinger et al. NEJM 1999; 340:278–284). We hypothesized that autoreactive γδ T lymphocytes might also be involved in the pathogenesis of cyclosporine-responsive PRCA, and thus examined whether the oligoclonal expansion would be detected in cyclosporine-responsive acquired PRCA patients.Methods: Five PRCA patients who achieved remission after oral cyclosporine therapy were included in this study, and informed consent was obtained before drawing blood samples. Blood mononuclear cells were stained with monoclonal antibodies specific for Vδ1, Vδ2 or Vδ3, and were analyzed by flow cytometry. Clonality of the Vδ1+ T cell subset was determined by complementarity-determining region 3 (CDR3) size spectratyping analysis of TCR Vδ1-Cδ PCR products. Junctional sequence was determined by DNA sequencing with an ABI377 automated DNA sequencer.Results: All five patients showed an increase of the Vδ1+ T cell subset in circulating blood (Table 1), although the major subset of human γδ T cells in healthy individuals is the Vδ2 (data not shown). Oligoclonal expansion was detected by CDR3 size spectratyping analysis and were confirmed by sequencing of the TCRVδ1-Cδ PCR products in all five patients (Table 2). No apparent consensus sequence among patients was observed in the CDR3 region of clonally expanded Vδ1+ T cells.Conclusions: Oligclonally expanded Vδ1+ T cells may be responsible for the pathogenesis of cyclosporine-responsive acquired PRCA. Biological function of these clonally expanding Vδ1+ T cells, natural ligands of the TCRs and requirements of antigen-presenting molecules remain to be elucidated.Table 1. γδT cell subsets in cyclosporine-responsive acquired PRCA patientsUPN01UPN02UPN03UPN04UPN05Pan- γδ9.6516.5318.9522.896.89δ 1 V6.6218.3713.3440.082.33δ 2 V0.660.201.120.150.72δ 3 V0.400.184.950.114.06Table 2. Oligoclonal expansions of Vδ1+ T cells and the CDR3 sequencesVδ 1NDNJδColony frequencyUPN01FCALGPLGFLPRWGIRGWDTRQMFF5/17FCALGGTSSYWGHYTDKLIFGKG5/17FCALGELGQGGYPPVRTAQLFFGKG4/17UPN02FCALGERAPHWGIRRITDKLIFGKG9/23FCALGEKRTYSPPLTAQLFFGKG6/23UPN03FCALGELSLPHSLVGPGGSHTDKLIFGKG5/23FCALGYGPRVGYPWGRIIFGKG4/23FCALGRQPSYGWWGISFPPYTDKLIFGKG3/23FCALGGAYVIWGKTDKLIFGKG3/23UPN04FCALGEREYWGMYTDKLIFGKG6/16FCALGKRGFPWPYWGIRATDKLIFGKG3/23UPN05FCALGHSGLFGVQYTDKLIFGKG5/24FCALGDPSYRRWGIRLYTDKLIFGKG3/24T-cell clones appearing more than twice in each sample are presented.