Diversity of peripheral blood human NK cells identified by single cell RNA sequencing

Human Natural Killer (NK) cells in peripheral blood perform many functions but classification of specific subsets has been a long-standing problem. Here, we report single-cell RNA sequencing of NK cells from healthy CMV-negative donors, comparing gene expression in unstimulated and IL-2 activated cells. Unsupervised clustering identified seven NK cell subsets. Three resembled well-described populations, CD56brightCD16-, CD56dimCD16+CD57- and CD56dimCD16+CD57+. CD56dimCD16+CD57- cells sub-divided to include a population with higher chemokine mRNA and increased frequency of KIR expression. Three novel human blood NK cell populations were identified as: a population of type I interferon responding NK cells which were CD56neg; a population exhibiting an in-vivo cytokine-induced memory-like phenotype, including increased granzyme B mRNA in response to IL-2, and finally, a small population, with low ribosomal expression, down-regulation of oxidative phosphorylation and high levels of immediate early response genes indicative of cellular activation. Human cytomegalovirus positive donors also included a higher frequency of adaptive NK cells. Together, these data establish an unexpected diversity in blood NK cells and provide a new framework for analysing NK cell responses in health and disease. scRNAseq of sorted NK cells from one healthy individual

外周血中人自然杀伤（Natural Killer, NK）细胞具备多种生理功能，但对其特定亚群的精准分类长期以来都是学界难题。本研究对巨细胞病毒（Cytomegalovirus, CMV）阴性健康供者的NK细胞开展单细胞RNA测序（single-cell RNA sequencing, scRNAseq），对比了未受刺激与白细胞介素-2（Interleukin-2, IL-2）激活状态下的细胞基因表达谱。通过无监督聚类分析，本研究共鉴定出7种NK细胞亚群：其中3种为已被广泛报道的经典亚群，即CD56brightCD16-、CD56dimCD16+CD57-与CD56dimCD16+CD57+；CD56dimCD16+CD57-亚群可进一步细分为一类趋化因子mRNA表达水平更高、杀伤细胞免疫球蛋白样受体（Killer cell immunoglobulin-like receptor, KIR）表达频率更高的细胞群。此外本研究还鉴定出3种全新的人外周血NK细胞亚群：其一为CD56阴性的I型干扰素应答型NK细胞；其二为呈现体内细胞因子诱导的记忆样表型的亚群，该亚群经IL-2刺激后颗粒酶B（Granzyme B）mRNA表达水平显著升高；最后为一类数量稀少的亚群，其核糖体表达水平较低、氧化磷酸化通路受抑制，且高表达指示细胞激活的即刻早期应答基因。巨细胞病毒阳性供者体内同样存在更高比例的适应性NK细胞。综上，本研究揭示了外周血NK细胞此前未被认知的高度多样性，并为分析健康与疾病状态下的NK细胞应答提供了全新的研究框架。本研究数据源自1名健康供者分选所得NK细胞的单细胞RNA测序。