Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Single-cell characterization of immune aplastic anemia reveals convergence of adaptive and innate immunity: in vivo

ABSTRACT: Immune aplastic anemia (AA) is a life-threatening bone marrow (BM) failure disorder driven by an autoimmune T cell attack against the hematopoietic stem and progenitor cells (HSPCs). However, the autoantigen targets and the role of other immune cells are unknown. Here, by combining scRNA+TCRαβ-seq, TCRβ-seq, flow cytometry and plasma cytokine profiling of >250 patients with AA, >250 patients with other hematological disorders and >1,000 healthy controls, we identify NK cells and CD8+ TEMRA cells expressing NK receptors with AA-associated TCRB-seq motifs as the most dysregulated immune cell populations in AA BM. Functional co-culture experiments with scRNA-TCRαβ-seq readout utilizing primary HSPCs and immune cells (in total 55 conditions) provide evidence that NK cells alone cannot kill HSPCs, but sensitize them to CD8+ T cell mediated killing. Furthermore, HSPCs induce strong activation of T cell clones with CD8+ TEMRA NK-like phenotype and AA-associated TCR motifs. To conclude, our results reveal convergent evolution of innate and adaptive immune cells in AA, where NK cells support CD8+ T cell mediated autoimmunity.

INSTRUMENT(S): Illumina NovaSeq 6000

ORGANISM(S): Homo sapiens

SUBMITTER: Jani Huuhtanen

PROVIDER: E-MTAB-13485 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:T cell receptor (TCR) αβ+ CD4- CD8- double negative T cells represent a rare T cell subset implicated in the pathogenesis of several autoimmune diseases. In this study, we investigated the DNA methylation signature of double negative T cells to gain insight into the epigenetic architecture and transcriptional capacity of peripheral blood primary human double negative T cells compared to autologous CD4+ and CD8+ T cells. We identified 2,984 CG sites across the genome with unique loss of DNA methylation in double negative T cells, and showed significant reduction in mRNA expression of DNA methyltransferases DNMT1, DNMT3A, and DNMT3B. DNA methylation was increased in CD8A/CD8B and CD4 consistent with epigenetic repression of both the CD8 and CD4 genetic loci in double negative T cells. Curiously, we show consistent increase in non-CG methylation in double negative T cells, a finding suggestive of pluripotency and previously only known to occur in embryonic stem cells and developing brain tissue. Network analyses of genes hypomethylated in double negative compared to CD4+ and CD8+ T cells indicate a strong relationship between double negative T cells and functions related to cell-cell interaction, cell adhesion, and cell activation pathways. Our data also suggest a robust pro-inflammatory epigenetic signature in double negative T cells, consistent with a transcriptional permissiveness to produce key inflammatory cytokines including IFNγ, IL-17F, IL-12B, IL-5, IL-18, TNFSF11 (RANKL), and TNFSF13B (BLYS or BAFF). These findings highlight an epigenetic basis for a role of double negative T cells in autoimmunity and extend the potential pathogenic transcriptional capacity of this unique T cell subset. Peripheral blood mononuclear cells (PBMCs) were isolated from fresh blood samples of 7 unrelated healthy women (age range 25-60) (Supplementary Table 1) by Ficoll-gradient centrifugation (GE Healthcare Bio-Sciences AB, Uppsala, Sweden). Subsequently, T cells were negatively-selected using the Human Pan T Cell Isolation II kit (Miltenyi Biotec, Cambridge, MA). T cells were then analyzed and sorted into TCRαβ+ CD4+ T cells, TCRαβ+ CD8+ T cells, and TCRαβ+ DN T cell subsets by flow cytometry. DNA was isolated from CD4+, CD8+, and DN T cell subsets using the DNeasy Blood and Tissue Kit (Qiagen, Valencia, CA), then bisulfite-converted using the EZ DNA Methylation kit (Zymo Research, Irvine, CA) for DNA methylation studies.

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Single-cell characterization of immune aplastic anemia reveals convergence of adaptive and innate immunity: in vivo

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