Promiscuity of Peptides Presented in HLA-DP Molecules from Different Immunogenicity Groups Is Associated With T-Cell Cross-Reactivity.

In the context of HLA-DP-mismatched allogeneic stem cell transplantation, mismatched HLA-DP alleles can provoke profound allo-HLA-DP-specific immune responses from the donor T-cell repertoire leading to graft-versus-leukemia effect and/or graft-versus-host disease in the patient. The magnitude of allo-HLA-DP-specific immune responses has been shown to depend on the specific HLA-DP disparity between donor and patient and the immunogenicity of the mismatched HLA-DP allele(s). HLA-DP peptidome clustering (DPC) was developed to classify the HLA-DP molecules based on similarities and differences in their peptide-binding motifs. To investigate a possible categorization of HLA-DP molecules based on overlap of presented peptides, we identified and compared the peptidomes of the thirteen most frequently expressed HLA-DP molecules. Our categorization based on shared peptides was in line with the DPC classification. We found that the HLA-DP molecules within the previously defined groups DPC-1 or DPC-3 shared the largest numbers of presented peptides. However, the HLA-DP molecules in DPC-2 segregated into two subgroups based on the overlap in presented peptides. Besides overlap in presented peptides within the DPC groups, a substantial number of peptides was also found to be shared between HLA-DP molecules from different DPC groups, especially for groups DPC-1 and -2. The functional relevance of these findings was illustrated by demonstration of cross-reactivity of allo-HLA-DP-reactive T-cell clones not only against HLA-DP molecules within one DPC group, but also across different DPC groups. The promiscuity of peptides presented in various HLA-DP molecules and the cross-reactivity against different HLA-DP molecules demonstrate that these molecules cannot be strictly categorized in immunogenicity groups.

Emerging Concepts in Immune Thrombotic Thrombocytopenic Purpura.

Immune thrombotic thrombocytopenic purpura (iTTP) is an autoimmune disorder of which the etiology is not fully understood. Autoantibodies targeting ADAMTS13 in iTTP patients have extensively been studied, the immunological mechanisms leading to the breach of tolerance remain to be uncovered. This review addresses the current knowledge on genetic factors associated with the development of iTTP and the interplay between the patient's immune system and environmental factors in the induction of autoimmunity against ADAMTS13. HLA-DRB1*11 has been identified as a risk factor for iTTP in the Caucasian population. Interestingly, HLA-DRB1*08:03 was recently identified as a risk factor in the Japanese population. Combined in vitro and in silico MHC class II peptide presentation approaches suggest that an ADAMTS13-derived peptide may bind to both HLA-DRB1*11 and HLA-DRB1*08:03 through different anchor-residues. It is apparent that iTTP is associated with the presence of infectious microorganisms, viruses being the most widely associated with development of iTTP. Infections may potentially lead to loss of tolerance resulting in the shift from immune homeostasis to autoimmunity. In the model we propose in this review, infections disrupt the epithelial barriers in the gut or lung, promoting exposure of antigen presenting cells in the mucosa-associated lymphoid tissue to the microorganisms. This may result in breach of tolerance through the presentation of microorganism-derived peptides that are homologous to ADAMTS13 on risk alleles for iTTP.

Long-term in vitro persistence of magnetic properties after magnetic bead-based cell separation of T cells.

Magnetic-activated cell sorting (MACS) using magnetic nanoparticles coated with specific antibodies is commonly used in immunology research. For in vitro isolation purposes, it is important to know to what extent the magnetic properties remain present in the isolated cell populations and whether it has consequences for sequential isolations. We hypothesized that only upon cell division, cells will lose their magnetic properties via dilution of the particles in/on their daughter cells. We analysed residual magnetic properties of cells that divided vs cells that did not divide after magnetic bead-based cell separation. As a model, we isolated T cells using beads targeting the non-modulating surface molecule CD45RO. Cells were labelled with the cell division tracking dye PKH and cultured under different conditions to induce variable degrees of cell division. We demonstrate that T cells that underwent no, or only minimal, cell divisions after MACS retained magnetic properties for up to at least 2 weeks of in vitro culture. The presence of nanoparticles was detected on their cell surface and intracellularly using Labeling Check reagent. These results have important consequences for procedures requiring repetitive isolation rounds after in vitro culture.

Priming of Allo-HLA-DP-Specific Reactivity from the Naïve T Cell Compartment Is Not Exclusively Mediated by Professional Antigen-Presenting Cells.

Allogeneic (allo) stem cell transplantation is applied to patients suffering from hematologic malignancies to replace the diseased hematopoietic system with cells derived from a donor stem cell graft. The majority of 10/10-matched unrelated donors are HLA-DP-mismatched, and this may result in varying degrees of the graft-versus-leukemia (GVL) effect with or without the occurrence of graft-versus-host disease (GVHD). Allo-HLA-reactive T cells are commonly present in the donor T cell repertoire, and thus a very profound alloreactive immune response can be provoked in the HLA-DP-mismatched setting. The magnitude and the diversity of the allo-HLA-DP-specific immune response likely dictates the balance between the occurrence of GVL and/or GVHD after transplantation. To understand the nature of the allo-HLA-DP-specific immune response provoked under different stimulatory conditions, immune responses were induced from both the naïve and memory T cell compartments using either HLA-DP-mismatched professional antigen-presenting cells (APCs) (monocyte-derived dendritic cells [allo-DCs]) or HLA-DP-mismatched nonprofessional APCs (skin-derived fibroblasts [allo-fibroblasts]) as stimulator cells. In this study, we observed that allo-HLA-DP-reactive T cells could be provoked from both the naïve and memory compartments by both types of APCs. However, the magnitude of the allo-HLA-DP-specific immune response was greater when stimulation was performed with allo-DCs. Moreover, we found that the frequency of allo-HLA-DP-reactive T cells was greater in the naïve T cell compartment compared with the memory T cell compartment, but we observed a comparable lineage specificity of these allo-HLA-DP-specific reactivities. Overall, the data from this study illustrate that the presence of professional APCs of recipient origin will mostly dictate the magnitude of the allo-HLA-DP-specific immune response derived from both the naïve and memory T cell compartments, but does not exclusively mediate the induction of these immune responses.

The allogeneic HLA-DP-restricted T-cell repertoire provoked by allogeneic dendritic cells contains T cells that show restricted recognition of hematopoietic cells including primary malignant cells.

Stem cell grafts from 10/10 HLA-matched unrelated donors are often mismatched for HLA-DP. In some patients, donor T-cell responses targeting the mismatched HLA-DP allele(s) have been found to induce a specific graft-versus-leukemia effect without coinciding graft-versus-host disease, whereas in other cases significant graft-versus-host disease occurred. Cell-lineage-specific recognition patterns within the allogeneic HLA-DP-specific donor T-cell repertoire could explain the differential clinical effects mediated by donor T cells after HLA-DP-mismatched allogeneic stem cell transplantation. To unravel the composition of the HLA-DP T-cell repertoire, donor T-cell responses were provoked by in vitro stimulation with allogeneic HLA-DP-mismatched monocyte-derived dendritic cells. A strategy including depletion of reactivity against autologous dendritic cells allowed efficient identification and enrichment of allo-reactive T cells upon stimulation with HLA-DP-mismatched dendritic cells. In this study we elucidated that the allogeneic HLA-DP-restricted T-cell repertoire contained T cells with differential cell-lineage-specific recognition profiles. As expected, some of the allogeneic HLA-DP-restricted T cells showed broad recognition of a variety of hematopoietic and non-hematopoietic cell types expressing the targeted mismatched HLA-DP allele. However, a significant proportion of the allogeneic HLA-DP-restricted T cells showed restricted recognition of hematopoietic cells, including primary malignant cells, or even restricted recognition of only myeloid cells, including dendritic cells and primary acute myeloid leukemia samples, but not of other hematopoietic and non-hematopoietic cell types. These data demonstrate that the allogeneic HLA-DP-specific T-cell repertoire contains T cells that show restricted recognition of hematopoietic cells, which may contribute to the specific graft-versus-leukemia effect without coinciding graft-versus-host disease.