T cell responses to human platelet antigen–1a involve a unique form of indirect allorecognition
Maria Therese Ahlen, … , Bjørn Skogen, Tor Brynjar Stuge
Maria Therese Ahlen, … , Bjørn Skogen, Tor Brynjar Stuge
Published September 8, 2016
Citation Information: JCI Insight. 2016;1(14):e86558. https://doi.org/10.1172/jci.insight.86558.
View: Text | PDF
Research Article Immunology

T cell responses to human platelet antigen–1a involve a unique form of indirect allorecognition

Abstract

Fetal and neonatal alloimmune thrombocytopenia (FNAIT) is a pregnancy-related condition caused by maternal antibodies binding an alloantigen on fetal platelets. In most cases the alloantigen is formed by a single amino acid, integrin β3 Leu33, referred to as human platelet antigen–1a (HPA-1a). Production of anti–HPA-1a antibodies likely depends on CD4+ T cells that recognize the same alloantigen in complex with the HLA-DRA/DRB3*01:01 molecule. While this complex is well characterized, T cell recognition of it is not. Here, to examine the nature of antigen recognition by HPA-1a–specific T cells, we assayed native and synthetic variants of the integrin β3 peptide antigen for binding to DRA/DRB3*01:01-positive antigen-presenting cells and for T cell activation. We found that HPA-1a–specific T cells recognize non-allogeneic integrin β3 residues anchored to DRA/DRB3*01:01 by the allogeneic Leu33, which itself is not directly recognized by these T cells. Furthermore, these T cell responses are diverse, with different T cells depending on different residues for recognition. This represents a unique form of indirect allorecognition in which a non-allogeneic peptide sequence becomes immunogenic by stable anchoring to MHC by an allogeneic residue.

Authors

Maria Therese Ahlen, Anne Husebekk, Ida Løken Killie, Bjørn Skogen, Tor Brynjar Stuge

×

Figure 2

HPA-1a–specific T cells do not exclusively recognize the Leu33 residue in the HPA-1a peptide.

Options: View larger image (or click on image) Download as PowerPoint
HPA-1a–specific T cells do not exclusively recognize the Leu33 residue i...
(A) CFSE-labeled HPA-1a–specific T cell clones were stimulated with DRA/DRB3*01:01-positive APCs (D4BL4) pulsed with a panel of designed peptides in different concentrations, without MLE. T cell proliferation was determined after 7 days, illustrated by 1 of 4 experiments (D7T4). (B) T cell activation was also measured by intracellular cytokine staining (IFN-γ) by flow cytometry. T cells were gated by light scatter and CFSE staining. One representative of 3 experiments with 10 μM peptide pulsing of D4BL4 cells, and (C) the effect of peptide pulsing concentrations on T cell activation. (D) Binding of peptides to D4BL4; same cell line used for T cell activation. Cells were pulsed with biotinylated peptides (10 μM) with or without AdEtOH, stained with Streptavidin–R-PE, and analyzed in flow cytometry. The discrimination of peptide-binding efficiency is enhanced when using AdEtOH as an MLE. Substitutions of Leu33 to Arg/Glu reduced binding, while efficient binding was seen with substitutions with small, aliphatic residues Leu33 to Val/Ile. Comparative binding experiments were conducted twice with 2 replicates; representative histograms are shown. Twelve-mer peptides were used in all experiments. HPA, human platelet antigen; MLE, MHC-loading enhancer; AdEtOH, adamantane ethanol.