Dataset Information

Single-cell transcriptomics in bone marrow delineates CD56^dimGranzymeK⁺ subset as intermediate stage in NK cell differentiation.

ABSTRACT: Human natural killer (NK) cells in lymphoid tissues can be categorized into three subsets: CD56^brightCD16⁺, CD56^dimCD16⁺ and CD69⁺CXCR6⁺ lymphoid tissue-resident (lt)NK cells. How the three subsets are functionally and developmentally related is currently unknown. Therefore, we performed single-cell RNA sequencing combined with oligonucleotide-conjugated antibodies against CD56, CXCR6, CD117 and CD34 on fresh bone marrow NK cells. A minor CD56^dimGzmK⁺ subset was identified that shared features with CD56^bright and CD56^dimGzmK^- NK cells based on transcriptome, phenotype (NKG2A^highCD16^lowKLRG1^highTIGIT^high) and functional analysis in bone marrow and blood, supportive for an intermediate subset. Pseudotime analysis positioned CD56^bright, CD56^dimGzmK⁺ and CD56^dimGzmK^- cells in one differentiation trajectory, while ltNK cells were developmentally separated. Integrative analysis with bone marrow cells from the Human Cell Atlas did not demonstrate a developmental connection between CD34⁺ progenitor and NK cells, suggesting absence of early NK cell stages in bone marrow. In conclusion, single-cell transcriptomics provide new insights on development and differentiation of human NK cells.

SUBMITTER: Melsen JE

PROVIDER: S-EPMC9730327 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature

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Single-cell transcriptomics in bone marrow delineates CD56dimGranzymeK+ subset as intermediate stage in NK cell differentiation.

Melsen Janine E JE van Ostaijen-Ten Dam Monique M MM Schoorl Dorenda J A DJA Schol Pieter J PJ van den Homberg Daphne A L DAL Lankester Arjan C AC Lugthart Gertjan G Schilham Marco W MW

Frontiers in immunology 20221124

Human natural killer (NK) cells in lymphoid tissues can be categorized into three subsets: CD56brightCD16+, CD56dimCD16+ and CD69+CXCR6+ lymphoid tissue-resident (lt)NK cells. How the three subsets are functionally and developmentally related is currently unknown. Therefore, we performed single-cell RNA sequencing combined with oligonucleotide-conjugated antibodies against CD56, CXCR6, CD117 and CD34 on fresh bone marrow NK cells. ...[more]

PMID: 36505452

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Project description:BackgroundHuman memory NK cells represent a heterogeneous CD56dim population that expands and persists in human cytomegalovirus (HCMV)-seropositive healthy individuals. They are characterized by the preferential, not fully overlapping, expression of NKG2C (activating receptor for HLA-E) and CD57 maturation marker, and by the lack of FcεRIγ adaptor chain. Hyperresponsiveness to Fcγ receptor IIIA (CD16) engagement represents the distinctive functional signature of memory NK cells. Although CD16 engagement was shown to acutely enhance glycolytic and oxidative pathways, its capability to induce a persisting metabolic reprogramming of human NK cells is poorly understood yet.ResultsHere, we describe the peculiar nutrient transporter expression pattern of FcεRIγ- memory NK cells, characterized by higher levels of CD98 neutral amino acid antiporter and CD71 transferrin receptor, and lower expression of GLUT1 glucose transporter, with respect to FcεRIγ+ conventional NK cells. Although CD16 engagement acutely enhances glycolytic and oxidative pathways, its capability to induce a persisting metabolic reprogramming of human NK cells is poorly understood yet. Our results firstly show that sustained CD16 engagement by contact with IgG-opsonized target cells induces the mTORC1-dependent upregulation of CD98 and CD71 nutrient receptors on CD56dim NK cells, in a transporter-specific fashion, that is finely tuned by cell-dependent (grade of functional maturation, and memory or conventional lineage) and stimulus-dependent (time length and cooperation with cytokines) factors. We also demonstrate that CD98 antiporter function is required for CD16-dependent IFN-γ production, and that enhanced CD98-mediated neutral amino acid uptake associates with heightened memory NK cell functional response.ConclusionCollectively, our work documents that CD16 engagement leads to a metabolic rewiring of human NK cells and suggests that a distinct nutrient transporter expression pattern may contribute to memory NK cell peculiar functional features.

Project description:BackgroundSystemic lupus erythematosus (SLE) is characterized by loss of immune tolerance and imbalance of immune cell subsets. Natural killer (NK) cells contribute to regulate both the innate and adaptive immune response. In this study, we aimed to detect alterations of peripheral NK cells and explore intrinsic mechanisms involving in NK cell abnormality in SLE.MethodsBlood samples from healthy controls (HCs) and patients with SLE and rheumatoid arthritis (RA) were collected. The NK count, NK subsets (CD56bright, CD56dimCD57-, and CD56dimCD57+), phenotypes, and apoptosis were evaluated with flow cytometer. Mitochondrial reactive oxygen species (mtROS) and total ROS levels were detected with MitoSOX Red and DCFH-DA staining respectively. Published data (GSE63829 and GSE23695) from Gene Expression Omnibus (GEO) was analyzed by Gene Set Enrichment Analysis (GSEA).ResultsTotal peripheral NK count was down-regulated in untreated SLE patients in comparison to that in untreated RA patients and HCs. SLE patients exhibited a selective reduction in peripheral CD56dimCD57+ NK cell proportion, which was negatively associated with disease activity and positively correlated with levels of complement(C)3 and C4. Compared with HCs, peripheral CD56dimCD57+ NK cells from SLE patients exhibited altered phenotypes, increased endogenous apoptosis and higher levels of mtROS and ROS. In addition, when treated with hydrogen peroxide (H2O2), peripheral CD56dimCD57+ NK cell subset was more prone to undergo apoptosis than CD56dimCD57- NK cells. Furthermore, this NK cell subset from SLE patients exhibited impaired cytotoxicity in response to activated CD4+ T cells in vitro.ConclusionOur study demonstrated a selective loss of mature CD56dimCD57+ NK cell subset in SLE patients, which may caused by preferential apoptosis of this subset under increased oxidative stress in SLE. The attenuated in vitro cytotoxicity of CD56dimCD57+ NK cells may contribute to the impaired ability of eliminating pathogenic CD4+ T cells in SLE.

Dataset Information

Single-cell transcriptomics in bone marrow delineates CD56^dimGranzymeK⁺ subset as intermediate stage in NK cell differentiation.

Publications

Single-cell transcriptomics in bone marrow delineates CD56<sup>dim</sup>GranzymeK<sup>+</sup> subset as intermediate stage in NK cell differentiation.

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