Dazed and Confused: NK Cells

Abstract

Innate lymphoid cells (ILCs) are rapid producers of both proinflammatory and regulatory cytokines in response to local injury, inflammation, pathogen infection, or commensal microbiota perturbation (1). Because most ILCs have been shown to be tissue-resident during homeostasis (with the exception of circulating NK cells) in almost all organs analyzed, their ability to quickly respond to tissue stress and inflammation underpins their critical role in regulating tissue homeostasis and repair during infection or injury (2–4). Recent evidence has suggested that mature ILCs can be further classified into group 1, 2, and 3 ILCs based on different expression of transcription factors, cell surface markers, and effector cytokines (1). Mouse group 1 ILCs, which include natural killer (NK) cells and ILC1, were initially distinguished from other ILCs based on their constitutive expression of the transcription factor Tbx21 (T-bet), co-expression of activating receptors NKp46 and NK1.1, and production of interferon (IFN)-γ following activation (5). In humans, group 1 ILCs are harder to definitively differentiate from other ILCs due to the lack of lineage defining markers and reported functional plasticity amongst group 2 and group 3 ILCs (6). ILC1 are recently discovered tissue-resident sentinels that function to protect the host from bacterial and viral pathogens at initial sites of infection (2, 7, 8). ILC1 rapidly produce IFN-γ following local dendritic cell activation and interleukin (IL)-12 production to limit viral replication and promote host survival before the recruitment of circulating lymphocytes into infected tissue (2). Unlike ILC1, NK cells can be recruited from the circulation into the parenchyma of infected or cancerous tissues where they display potent perforin-dependent cytotoxicity in addition to rapid IFN-γ production (9, 10). However, persistent inflammatory signals can also lead to unrestrained activation of group 1 ILCs during obesity and inflammatory bowel disease (IBD) (3, 11–14). While these studies suggest important roles for group 1 ILCs during host protection and pathology, gaps in evidence have inhibited the ability of recent studies to definitively distinguish between the roles of ILC1 and NK cells in these contexts. NK cells, the founding member of ILCs, were initially defined based on the cell surface expression of NK1.1 in mouse or CD56 in human with the absence of cell surface expression of other lineage (Lin) defining markers including CD3, CD14, CD19, and TCR proteins (15). In subsequent mouse studies over the last 30 years, Lin⁻NK1.1⁺ cells were found to be heterogeneous for the expression of activating and inhibitory Ly49 receptors, cell surface integrins [α1β1 (CD49a), α2β1 (CD49b), αEβ7 (CD103)], cell surface proteins (TRAIL, CD69, CD27, CD11b), transcription factors (Eomes), chemokine receptors (CXCR6), and cytokine receptors (IL-7Rα) in various organs (1, 16). Similarly, human Lin⁻CD56⁺ cells have been reported to be heterogeneous for the expression of transcriptions factors (EOMES and T-BET), cell surface markers (CD49a, CD56, CD16, NKp80, CXCR6, IL-7Rα, CD94, CD69, NKp44), and cytotoxic molecules (Perforin) (1, 16). Early studies concluded that cells with an alternative cell surface or transcription factor phenotype from putative mature NK cells (mouse: Lin⁻NK1.1⁺T-bet⁺Eomes⁺CD49b⁺; human: Lin⁻ IL-7Rα⁻CD56^dimCD16⁺) in peripheral organs and blood likely represented immature NK (iNK) cells (17–21). This hypothesis is supported by studies demonstrating that subsets of developing mouse NK cells can be distinguished based on CD27 and CD11b expression (22, 23). Similarly, previous studies have suggested that CD56^brightCD16⁻ human NK cells in the blood may be immature precursors to CD56^dimCD16⁺ mature NK cells (18, 19). However, whether other phenotypic differences observed in mouse and human group 1 ILCs are due to tissue-specific microenvironments, distinct lineages of cells, or developmental/activation states of NK cells is still under considerable debate and investigation. Insight into these questions came shortly after the identification of Lin⁻IL-7Rα⁺ “helper” ILCs. Specifically, genetic evidence suggested that Tbx21-dependent IL-7Rα⁺Tbet⁺Eomes⁻NK1.1⁺NKp46⁺ “ILC1” in the small intestine did not require Eomes for their development, whereas NK cells did require Eomes (7). A recent study further supported these initial data by using Eomes-GFP reporter mice to generate core transcriptional signatures of Eomes⁻ ILC1 and Eomes⁺ NK cells from 4 independent tissues. The identified core ILC1 signature led to the discovery of the inhibitory receptor CD200r1 as a stable marker expressed by ILC1 but not NK cells during homeostasis and inflammation (2). Additional lineage tracing experiments suggested that CD200r1⁺Eomes⁻CD49b⁻ group 1 ILCs constituted a stable lineage during homeostasis, distinct from CD200r1-Eomes⁺CD49b⁺ mature NK (mNK) cells (2, 7, 24). Functional evidence suggestive of distinct group 1 ILCs in peripheral organs was supported by the findings that T-bet⁺Eomes⁻CD49b⁻ group 1 ILCs (in addition to ILC2 and ILC3) were long-term tissue-resident cells, whereas Eomes⁺CD49b⁺ mNK cells were derived from the circulation in almost all organs tested in mouse parabiosis experiments (2, 4). Similarly, in one human study a subset of donor liver CXCR6⁺ group 1 ILCs was found to be maintained up to...