Project description:A high-throughput mass spectrometry analysis was used to identify more than 16,000 cell peptides bound to several HLA-DR and -DP class II molecules isolated from large amounts of two human cell lines (HOM-2 and JY).
Project description:Accurate prediction of antigen presentation by Human Leukocyte Antigen (HLA) class II molecules is crucial for rational development of immunotherapies and vaccines targeting CD4 T cell activation. So far, most prediction methods for HLA class II antigen presentation have focused on HLA-DR due to limited availability of immunopeptidomics data for HLA-DQ and HLA-DP, while not taking into account alternative peptide binding modes. Here, we present an update to the NetMHCIIpan prediction method which closes the performance gap between all three HLA class II loci. We accomplish this by first integrating large immunopeptidomics datasets describing the HLA class II specificity space across loci using a refined machine learning framework that accommodates inverted peptide binders. Next, we apply targeted immunopeptidomics assays to generate novel data that covers additional HLA-DP specificities. The final method, NetMHCIIpan-4.3, achieves high accuracy and molecular coverage across all HLA class II allotypes.
Project description:HLA-DR-lacking HSPCs [HLA-DR(-) HSPCs] were detected in aplastic anemia (AA) patients with HLA-DR15. HLA-DR(-) HSPCs may evade the attack by CD4+ T-cells recognizing the autoantigen presented by HLA-DR15. The goal of this study is to clarify the immune escape mechanisms from antigen-specific T-cells by comparing the trranscriptome profile of HLA-DR(+) HSPCs and HLA-DR(-) HSPCs.
Project description:Myeloid-derived suppressor cells (MDSC) is a heterogeneous population of cells that can negatively regulate T-cell function. As opposed to murine MDSC, which are characterized as Gr-1+CD11b+ cells, human MDSC are not so clearly defined due to lack of specific markers. Our lab has previously identified a new subset of MDSC as CD14+HLA-DR-neg/low cells from PBMC. CD14+HLA-DR-neg/low MDSC not only suppress proliferation and IFN-gamma secretion of autologous T cells, but also induce CD25+Foxp3+ regulatory T cells that are suppressive in vitro, whereas the counterpart CD14+HLA-DR-high monocytes don’t have the effect. In this study, we compare the immune-related gene expression between CD14+HLA-DR-neg/low MDSC and CD14+HLA-DR-high monocytes to better characterize the difference between these two populations and to find new potential specific marker for human MDSC. PBMC were isolated from fresh blood healthy donor by density centrifugation. CD14+ cells were isolated by AutoMACS CD14 microbeads using a AutoMACS (Miltenyi), and then stained with CD14 and HLA-DR antibodies. MDSC and monocytes control cells were sorted as CD14+ HLA-DR-neg/low and CD14+HLA-DR-high cells respectively. The sorted two populations were immediately frozen in liquid nitrogen and shipped to the company on dry ice for RNA isolation and further microarray.
Project description:Negative control (healthy cells) of MSV000079647. HLA-DR ligands from healthy Jy cells. Using mass spectrometry analysis of complex HLA class II-bound peptide pools isolated from large amounts of non-infected cells were identified.
Project description:Myeloid-derived suppressor cells (MDSC) is a heterogeneous population of cells that can negatively regulate T-cell function. As opposed to murine MDSC, which are characterized as Gr-1+CD11b+ cells, human MDSC are not so clearly defined due to lack of specific markers. Our lab has previously identified a new subset of MDSC as CD14+HLA-DR-neg/low cells from PBMC. CD14+HLA-DR-neg/low MDSC not only suppress proliferation and IFN-gamma secretion of autologous T cells, but also induce CD25+Foxp3+ regulatory T cells that are suppressive in vitro, whereas the counterpart CD14+HLA-DR-high monocytes don’t have the effect. In this study, we compare the immune-related gene expression between CD14+HLA-DR-neg/low MDSC and CD14+HLA-DR-high monocytes to better characterize the difference between these two populations and to find new potential specific marker for human MDSC.
Project description:HLA-DRB1 alleles have been associated with several autoimmune diseases. In anti-citrullinated protein antibody positive rheumatoid arthritis (ACPA-positive RA), HLA-DRB1 shared epitope (SE) alleles are the major genetic risk factors. In order to investigate whether expression of different alleles of major histocompatibility complex (MHC) Class II genes influence functions of immune cells, we investigated transcriptomic profiles of a variety of immune cells from healthy individuals carrying different HLA-DRB1 alleles. Sequencing libraries from peripheral blood mononuclear cells, CD4+ T cells, CD8+ T cells, and CD14+ monocytes of 32 genetically pre-selected healthy female individuals were generated, sequenced and reads were aligned to the standard reference. For the MHC region, reads were mapped to available MHC reference haplotypes and AltHapAlignR was used to estimate gene expression. Using this method, HLA-DRB and HLA-DQ were found to be differentially expressed in different immune cells of healthy individuals as well as in whole blood samples of RA patients carrying HLA-DRB1 SE-positive versus SE-negative alleles. In contrast, no genes outside the MHC region were differentially expressed between individuals carrying HLA-DRB1 SE-positive and SE-negative alleles. Existing methods for HLA-DR allele-specific protein expression were evaluated but were not mature enough to provide appropriate complementary information at the protein level. Altogether, our findings suggest that immune effects associated with different allelic forms of HLA-DR and HLA-DQ may be associated not only with differences in the structure of these proteins, but also with differences in their expression levels.
Project description:Loss of human leukocyte antigen (HLA) expression on tumor cells is frequent in diffuse large B cell lymphoma (DLBCL) arising in immune-privileged sites such as the testis and central nervous system and is associated with small homozygous deletions of HLA-DQ/DR and larger hemizygous deletions of the major histocompatibility complex (MHC) region. To better understand the significance of downregulation of HLA class II expression in relation to the homozygous and hemizygous deletions, we analyzed global gene expression patterns in a series of 26 testicular diffuse large B-cell lymphomas (DLBCL) after characterization of these deletions. Low levels of HLA-DR mRNA in whole testicular DLBCL samples were associated with a strong downregulation of numerous immune-related genes specific for T-cells, macrophages, antigen presentation and processing, lymphocyte activation, chemokines and chemokine receptors and the complement system. Interestingly, hemizygous and homozygous deletions in the MHC region did not have any additional impact on global gene expression. Keywords: Gene expression