title: T-cell responses associated with neonatal alloimmune thrombocytopenia: isolation of HPA-1a-specific, HLA-DRB3*0101-restricted CD4+ T cells.
abstract: T-cell responses have been implicated in the development of HPA-1a-induced neonatal alloimmune thrombocytopenia (NAIT). However, HPA-1a-specific T cells have neither been isolated nor characterized. Here, we aimed to determine whether HPA-1a-specific T cells could be isolated from HPA-1a-immunized women. In the present study, peripheral blood mononuclear cells (PBMCs) from an HPA-1a-alloimmunized woman were cultured for weeks in the presence of HPA-1a peptide, labeled with CFSE, and assayed for antigen-specific proliferation. Individual proliferating cells were isolated by fluorescence-activated cell sorting and expanded in culture. Antigen specificity and HLA restriction were determined by cytokine secretion (enzyme-linked immunospot [ELISPOT]) and proliferation assays. Several CD3(+)CD4(+) T-cell clones were isolated that proliferated and secreted cytokines in response to HPA-1a peptide. Two of these clones have been established in long-term culture in our laboratory. Both of these recognize synthetic as well as naturally processed HPA-1a antigen, and the recognition is restricted by the MHC molecule HLA-DRB3*0101 that is strongly associated with NAIT. These HPA-1a-specific T-cell clones represent unambiguous evidence for the association of T-cell responses with NAIT, and they will serve as unique tools to elucidate the cellular immune response that may result in NAIT.
authors.affiliation: Department of Immunology and Transfusion Medicine, University Hospital of North Norway, Tromsø.
authors.forename: Maria Therese
authors.initials: MT
authors.lastname: Ahlen
authors.forename: Anne
authors.initials: A
authors.lastname: Husebekk
authors.forename: Mette Kjaer
authors.initials: MK
authors.lastname: Killie
authors.forename: Bjørn
authors.initials: B
authors.lastname: Skogen
authors.forename: Tor B
authors.initials: TB
authors.lastname: Stuge
mesh_terms: D000328:Adult; D016824:Antigens, Human Platelet / Q000276:immunology*; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D049109:Cell Proliferation; D002478:Cells, Cultured; D016207:Cytokines / Q000276:immunology; D005260:Female; D006684:HLA-DR Antigens / Q000276:immunology*; D059826:HLA-DRB3 Chains; D006801:Humans; D007231:Infant, Newborn; D010455:Peptides / Q000276:immunology*; D054098:Thrombocytopenia, Neonatal Alloimmune / Q000276:immunology*
chemical_list: D016824:Antigens, Human Platelet; D016207:Cytokines; D006684:HLA-DR Antigens; D059826:HLA-DRB3 Chains; D010455:Peptides