Tissue-resident Natural Killer Cells Research Articles

Exploring a specific type of tissue-resident natural killer cell involved in the anti-tumor and immunotherapy response in human papillomavirus-positive head and neck squamous cell carcinoma using scRNA-seq.

Human papillomavirus (HPV)-positive head and neck squamous cell carcinoma (HNSCC) is an increasingly common malignancy. We aimed to explore the immune heterogeneity of natural killer (NK) cells in HPV-positive HNSCC. Single-cell RNA-sequencing (scRNA-seq) and bulk RNA-sequencing datasets of HPV-positive HNSCC data were obtained from the Gene Expression Omnibus (GEO) database. "Seurat", "harmony", and "SingleR" were used to perform the scRNA-seq analysis. Subsequently, the "cellphonedb" package was used for the cell crosstalk analysis, and the "clusterProfiler" package was used for the hallmark pathway enrichment analysis. Finally, the "gene set variation analysis" ("GSVA") package was used for the immune cell infiltration, Tumor Immune Dysfunction and Exclusion (TIDE), and risk-score analyses. A total of 30,562 cells were classified into 9 cell clusters that comprised 6 main cell types [i.e., T cells, natural killer T (NKT) cells, NK cells, B cells, plasma cells, and macrophages]. The NK cells were then further clustered into 3 tissue-resident NK (trNK0-2) and 2 tumor-associated NK (taNK0-1) cell types. The trNK0 cell type, which exhibited inhibitory cancer hallmark activity, appeared to exert potential anti-tumor effects via trNK0-macrophage crosstalk. The trNK score could serve as an independent and valuable prognostic classifier, as the patients with high-trNK scores had better outcomes, immune-infiltration levels, and immunotherapy effects. Using an scRNA-seq analysis, we identified a specific type of tissue-resident NK cell (i.e., trNK-0) that was involved in the anti-tumor and immunotherapy response in HPV-positive HNSCC.

Urine scRNAseq reveals new insights into the bladder tumor immune microenvironment.

Due to bladder tumors' contact with urine, urine-derived cells (UDCs) may serve as a surrogate for monitoring the tumor microenvironment (TME) in bladder cancer (BC). However, the composition of UDCs and the extent to which they mirror the tumor remain poorly characterized. We generated the first single-cell RNA-sequencing of BC patient UDCs with matched tumor and peripheral blood mononuclear cells (PBMC). BC urine was more cellular than healthy donor (HD) urine, containing multiple immune populations including myeloid cells, CD4+ and CD8+ T cells, natural killer (NK) cells, B cells, and dendritic cells (DCs) in addition to tumor and stromal cells. Immune UDCs were transcriptionally more similar to tumor than blood. UDCs encompassed cytotoxic and activated CD4+ T cells, exhausted and tissue-resident memory CD8+ T cells, macrophages, germinal-center-like B cells, tissue-resident and adaptive NK cells, and regulatory DCs found in tumor but lacking or absent in blood. Our findings suggest BC UDCs may be surrogates for the TME and serve as therapeutic biomarkers.

Single-cell transcriptome sequencing to explore the infiltration and disappearance of NK cells in liver cancer.

e14520 Background: The emergence of tumor immunotherapy such as immune checkpoint blocking has greatly promoted the development of Hepatocellular carcinoma (HCC) treatment. However, the response rate of HCC patients to immunotherapy is not high, and the probability of adverse reactions is high. Natural killer (NK) cells play a crucial role in inhibiting the proliferation and metastasis of tumor cells and are expected to become a new hope for tumor therapy. Methods: The HCC single-cell transcriptome atlas was constructed by using the tumor tissues, adjacent tissues, peripheral blood of HCC patients. To emulate the hypoxic state within the tumor, NK cells were treated with rotenone, and the post-rotenone treatment changes of NK cells in transcriptome levels were evaluated using qPCR. The proliferation, apoptosis, and cytotoxicity of NK cells were assessed via flow cytometry. The study also investigated the role of c-Jun N-terminal protein kinase (JNK) in NK cell function by applying the JNK inhibitor tanzisertib (CC-930) in vitro. The impact of JNK inhibition on the tumor-killing potential of NK cells was examined in vivo using tumor-bearing mice NCG (NOD/ShiLtJ-Prkdcem26Cd52Il2rgem26Cd22/Gpt). Results: Analysis of the source of NK cell subsets found that the specific NK cells in tumor tissues were tissue-resident NK cells (trNK). Both RNA velocity analysis and pseudo-time sequence analysis suggested that trNK cells differentiated from PBMC-1 NK cells. Upon treatment of PBMC-1 NK cells with Tanzisertib, we observed inhibition of NK cell apoptosis. Through the subcutaneous tumor formation experiment of NCG mice, Tanzisertib-treated NK cells were transferred into tumor-bearing mice. Notably, we observed a significant reduction in tumor weight (Before treatment: 0.19g, 0.19g, 0.22g, 0.24g, 0.26g, 0.27g; After treatment: 0.16g, 0.17g, 0.17g, 0.19g, 0.21g, 0.23g; P value = 0.049). TUNEL assay confirmed a significant increase number of apoptotic cells within the tumor and an increased number of infiltrated NK cells. Conclusions: The NK cells in HCC tumors are likely derived from the infiltration of NK cells in peripheral blood. Importantly, the inhibition of JNK function can significantly inhibit the apoptosis of NK cells and enhance the killing ability of NK cells against tumor cells in vivo.

Natural killer cells display unique, non-cytotoxic profiles in human endometrial cancer

Abstract Endometrial cancer (EC) responds variably to immune checkpoint blockade, warranting study of the tumor microenvironment (TME). Natural killer (NK) cells are cytotoxic innate lymphocytes that can directly kill tumor cells, yet their diversity, locations, and functions in the EC TME are unknown. CITE-seq analysis of fresh EC tumor samples revealed multiple NK cell populations in the TME, with distinct patterns of surface activating and inhibitory receptor expression and less cytolytic granule expression by tumor-derived conventional NK (cNK) and tissue-resident NK (trNK) cells compared to autologous blood cNK. Ex vivo flow cytometry revealed significantly lower expression of cytolytic granules by TME-derived NK cell subsets (granzyme B MFI: blood cNK 1.5e5 +/- 5.4e4, tumor cNK 7.9e4 +/- 2.9e4 p<0.05, tumor trNK 3.7e4 +/- 1.4e4 p<0.05, n=10; perforin MFI: blood cNK 1.3e4 +/- 3.1e3, tumor cNK 1.0e4 +/- 3.2e3 p=0.19, tumor trNK 3.5e3 +/-1.3e3 p<0.01, n=8). To identify NK cell subsets within the TME, we performed immunohistochemistry using CD3, CD56, granzyme B, and CD103 to demarcate cNK and trNK, respectively. We observed granzyme B+ cells primarily in the stroma (78.6 +/- 3.7% Stroma, 21.4 +/- 3.7% Glands, n=10, p<0.05) and CD103+ cells primarily among tumor cells (15.6 +/- 4.5% Stroma, 84.3 +/- 0.64% Glands, n=10, p=0.07). These data suggest that both cNK and trNK cell cytotoxicity is suppressed in the EC TME and ongoing work seeks to elucidate the mechanisms.

Tissue-resident memory NK cells: Homing in on local effectors and regulators.

Natural killer (NK) cells are the prototype innate effector lymphocyte population that plays an important role in controlling viral infections and tumors. Studies demonstrating that NK cells form long-lived memory populations, akin to those generated by adaptive immune cells, prompted a revaluation of the potential functions of NK cells. Recent data demonstrating that NK cells are recruited from the circulation into tissues where they form long-lived memory-like populations further emphasize that NK cells have properties that mirror those of adaptive immune cells. NK cells that localize in non-lymphoid tissues are heterogeneous, and there is a growing appreciation that immune responses occurring within tissues are subject to tissue-specific regulation. Here we discuss both the immune effector and immunoregulatory functions of NK cells, with a particular emphasis on the role of NK cells within non-lymphoid tissues and how the tissue microenvironment shapes NK cell-dependent outcomes.

Tissue-resident Lymphocytes Are Released During Hypothermic and Normothermic Machine Perfusion of Human Donor Kidneys.

Machine perfusion is the preferred preservation method for deceased donor kidneys. Perfusate fluid, which contains a complex mixture of components, offers potential insight into the organ's viability and function. This study explored immune cell release, particularly tissue-resident lymphocytes (TRLs), during donor kidney machine perfusion and its correlation with injury markers. Perfusate samples from hypothermic machine perfusion (HMP; n = 26) and normothermic machine perfusion (NMP; n = 16) of human donor kidneys were analyzed for TRLs using flow cytometry. Residency was defined by expressions of CD69, CD103, and CD49as. TRL release was quantified exclusively in NMP. Additionally, levels of cell-free DNA, neutrophil gelatinase-associated lipocalin, and soluble E-cadherin (sE-cadherin) were measured in NMP supernatants with quantitative polymerase chain reaction and enzyme-linked immunosorbent assay. Both HMP and NMP samples contained a heterogeneous population of TRLs, including CD4 + tissue-resident memory T cells, CD8 + tissue-resident memory T cells, tissue-resident natural killer cells, tissue-resident natural killer T cells, and helper-like innate lymphoid cells. Median TRL proportions among total CD45 + lymphocytes were 0.89% (NMP) and 0.84% (HMP). TRL quantities in NMP did not correlate with donor characteristics, perfusion parameters, posttransplant outcomes, or cell-free DNA and neutrophil gelatinase-associated lipocalin concentrations. However, CD103 + TRL release positively correlated with the release of sE-cadherin, the ligand for the CD103 integrin. Human donor kidneys release TRLs during both HMP and NMP. The release of CD103 + TRLs was associated with the loss of their ligand sE-cadherin but not with general transplant injury biomarkers.

On blood and tissue-resident natural killer cells

Conventional natural killer (cNK) cells patrol the organism via circulation and invade tissues in response to infection or inflammation. In this issue of Immunity, Torcellan etal. report that circulating cNK cells are recruited into infected skin and differentiate into long-lived tissue-resident NK cells capable of mediating an accelerated response upon reinfection.

Circulating NK cells establish tissue residency upon acute infection of skin and mediate accelerated effector responses to secondary infection

Natural killer (NK) cells are present in the circulation and can also be found residing in tissues, and these populations exhibit distinct developmental requirements and are thought to differ in terms of ontogeny. Here, we investigate whether circulating conventional NK (cNK) cells can develop into long-lived tissue-resident NK (trNK) cells following acute infections. We found that viral and bacterial infections of the skin triggered the recruitment of cNK cells and their differentiation into Tcf1hiCD69hi trNK cells that share transcriptional similarity with CD56brightTCF1hi NK cells in human tissues. Skin trNK cells arose from interferon (IFN)-γ-producing effector cells and required restricted expression of the transcriptional regulator Blimp1 to optimize Tcf1-dependent trNK cell formation. Upon secondary infection, trNK cells rapidly gained effector function and mediated an accelerated NK cell response. Thus, cNK cells redistribute and permanently position at sites of previous infection via a mechanism promoting tissue residency that is distinct from Hobit-dependent developmental paths of NK cells and ILC1 seeding tissues during ontogeny.

A Pan-Cancer Single-Cell Transcriptomic Atlas of Natural Killer (NK) Cells Reveals Intrinsic and Extrinsic Mediators of NK Cell Anti-Tumor Functions

Background: Natural killer (NK) cells are critical mediators of anti-tumor immunity, engaging in direct cytotoxic responses and promoting pro-inflammatory immune microenvironments. The molecular mechanisms underlying these functions, however, have yet to be fully elucidated. This is due in part to a lack of understanding of the phenotypic and functional diversity of NK cells across cancer types. Here, we present a single-cell transcriptomic atlas of >250,000 NK cells spanning 340 patients across 17 cancer types to advance our understanding of tumor-infiltrating NK cells. Methods: We re-analyzed publicly available single-cell RNA-sequencing (scRNA-seq) datasets of primary, therapy-naïve tumors, as well as matching peripheral blood samples and healthy donor blood samples. We employed systematic computational analyses to deconvolute the phenotypic and functional diversity of tumor-infiltrating NK cells and define key regulators of NK cell functions in both a cancer type-specific and pan-cancer manner. Feature expression module scoring was used to infer phenotypic and functional characteristics of NK cell clusters in each cancer type. Transcription factor regulon and cell-cell communication prediction algorithms were applied to understand the mechanistic drivers of NK cell populations within tumors. Results: Tumor-infiltrating NK cells display distinct patterns of phenotypic and functional features across cancer types. Nonetheless, we identify consensus NK cell populations across cancers that demonstrate the utility and performance of our gene module-based phenotypic and functional annotations. Interestingly, we observe the enrichment of adaptive-like and CD56 bright NK cell populations in tumors that exhibit features of tissue residency, including the upregulation of ITGA1, ITGAE, and CD69 alongside the downregulation of FCG3RA. Notably, these tissue-resident populations display enhanced chemokine and cytokine activity compared to their recruited counterparts, suggesting the significance of tissue-resident NK cell populations in immune recruitment to tumors. We find the phenotypic composition of intra-tumoral NK cells is vastly different than that of circulating NK cells which harbor predominantly CD56 dim populations, as well as a significant proportion of adaptive-like NK cells in several cancer types. Leveraging the paired nature of several tumor and peripheral blood samples, we performed temporal comparative analyses that revealed the robust expression of KLF2 and S1PR5 in circulation which is then lost in paired tumor tissues. While KLF2 and S1PR5 have been previously implicated in the homing and migration of immune cells, their roles in the trafficking of NK cells to tumors have yet to be defined. In modeling transitions from canonical to stressed NK cell states, we not only identify well-established NK cell checkpoints ( TIM-3, LAG3, and CISH) but also identify novel putative checkpoints yet to be investigated in NK cells, such as the AP-1 family of transcription factors. Additionally, we examine extrinsic factors contributing to intra-tumoral NK cell phenotypes, detecting significant crosstalk between NK cells and immunomodulatory macrophages and CD4+ T cells mediated by galectin-9 and TGFβ interactions. Conclusions: We define the phenotypic and functional pan-cancer landscape of NK cells and uncover both intrinsic and extrinsic factors influencing their anti-tumor functions. We envision our NK cell atlas will facilitate the development of strategies to enhance NK cell-based immunotherapies against cancer.

The immune and metabolic treatment approach of using testosterone on mice models of liver injury.

Background: Natural killer (NK) cells showed an anti-fibrotic effect; however, their function is thought to be impaired in advanced liver injury. In the current study, we aimed to assess the immune and metabolic impact of testosterone on mice models of liver injury. Methods: Carbon-tetrachloride induced liver fibrosis male mice models was i.p injected for 2 weeks (acute) and 4 weeks (chronic) (n = 36). Testosterone (4mg/kg mouse body weight) was injected i.p. following the first week of the acute model of CCl4 and following the second week of the chronic model of CCl4. At the end of the experiments, mice were sacrificed, and serum was collected for assessing liver enzymes of ALT and AST, as well as inflammatory markers of IL-6, metabolic makers of C-peptide levels, and lipid and glucose profiles. Livers were harvested and used for histological assessments for inflammation and fibrosis. Fibrosis profiles from liver extracts, αSMA and Collagen III, were assessed by RT-PCR. Moreover, liver tissue-resident NK cells were isolated and evaluated for their activity by assessing INF-γ and IL-6 receptors using ELISA and flow cytometry, respectively. Results: Serum ALT, AST, and IL-6, as well as metabolic assessments of cholesterol, triglyceride, C-peptide, fasting blood sugar, and fibrotic profiles, were linearly correlated with disease progressions. Histological characterization of the liver was worsened in the chronic model of liver injury. Testosterone-treated mice exhibit a significant reduction in collagen depositions with less dense fibrosis tissue associated with reduced liver injury enzymes and metabolic markers in both the acute and chronic CCl4 mice models in favor of the latter one (p < 0.05). Moreover, testosterone treatments displayed a significant decrease in serum IL-6 of 2.4-fold (p = 0.0001) and 2.3-fold (p = 0.0003) in the acute and chronic models, respectively (p = 0.002), and data showed an increase in INF-γ release from NK associated with a reduction in their IL-6 receptor expressions (p < 0.05). Conclusion: Our results indicated effects of testosterone on mediating a decreased expressions of NK IL-6 receptors and consequently inducing their activation; which in part, could explain the amelioration of liver injury. Our data suggest an anti-inflammatory and anti-fibrotic treatment approach of using testosterone for delaying disease progressions.

Control of human cytomegalovirus replication by liver resident natural killer cells

Natural killer cells are considered to be important for control of human cytomegalovirus– a major pathogen in immune suppressed transplant patients. Viral infection promotes the development of an adaptive phenotype in circulating natural killer cells that changes their anti-viral function. In contrast, less is understood how natural killer cells that reside in tissue respond to viral infection. Here we show natural killer cells resident in the liver have an altered phenotype in cytomegalovirus infected individuals and display increased anti-viral activity against multiple viruses in vitro and identify and characterise a subset of natural killer cells responsible for control. Crucially, livers containing natural killer cells with better capacity to control cytomegalovirus replication in vitro are less likely to experience viraemia post-transplant. Taken together, these data suggest that virally induced expansion of tissue resident natural killer cells in the donor organ can reduce the chance of viraemia post-transplant.

Deciphering the localization and trajectory of human natural killer cell development.

Innate immune cells represent the first line of cellular immunity, comprised of both circulating and tissue-resident natural killer cells and innate lymphoid cells. These innate lymphocytes arise from a common CD34+ progenitor that differentiates into mature natural killer cells and innate lymphoid cells. The successive stages in natural killer cell maturation are characterized by increased lineage restriction and changes to phenotype and function. Mechanisms of human natural killer cell development have not been fully elucidated, especially the role of signals that drive the spatial localization and maturation of natural killer cells. Cytokines, extracellular matrix components, and chemokines provide maturation signals and influence the trafficking of natural killer cell progenitors to peripheral sites of differentiation. Here we present the latest advances in our understanding of natural killer and innate lymphoid cell development in peripheral sites, including secondary lymphoid tissues (i.e. tonsil). Recent work in the field has provided a model for the spatial distribution of natural killer cell and innate lymphoid cell developmental intermediates in tissue and generated further insights into the developmental niche. In support of this model, future studies using multifaceted approaches seek to fully map the developmental trajectory of human natural killer cells and innate lymphoid cells in secondary lymphoid tissues.

Phenotypic diversity of human adipose tissue-resident NK cells in obesity.

Natural killer (NK) cells have emerged as key mediators of obesity-related adipose tissue inflammation. However, the phenotype of NK cell subsets residing in human adipose tissue are poorly defined, preventing a detailed understanding of their role in metabolic disorders. In this study, we applied multicolor flow cytometry to characterize CD56bright and CD56dim NK cells in blood and adipose tissue depots in individuals with obesity and identified surface proteins enriched on adipose tissue-resident CD56bright NK cells. Particularly, we found that adipose tissue harbored clusters of tissue-resident CD56bright NK cells signatured by the expression of CD26, CCR5 and CD63, possibly reflecting an adaptation to the microenvironment. Together, our findings provide broad insights into the identity of NK cells in blood and adipose tissue in relation to obesity.

A single-cell analysis framework allows for characterization of CSF leukocytes and their tissue of origin in multiple sclerosis.

Peripheral central nervous system (CNS)-infiltrating lymphocytes are a hallmark of relapsing-remitting multiple sclerosis. Tissue-resident memory T cells (TRM) not only populate the healthy CNS parenchyma but also are suspected to contribute to multiple sclerosis pathology. Because cerebrospinal fluid (CSF), unlike CNS parenchyma, is accessible for diagnostics, we evaluated whether human CSF, apart from infiltrating cells, also contains TRM cells and CNS-resident myeloid cells draining from the parenchyma or border tissues. Using deep generative models, we integrated 41 CSF and 14 CNS parenchyma single-cell RNA sequencing (scRNAseq) samples from eight independent studies, encompassing 120,629 cells. By comparing CSF immune cells collected during multiple sclerosis relapse with cells collected during therapeutic very late antigen-4 blockade, we could identify immune subsets with tissue provenance across multiple lineages, including CNS border-associated macrophages, CD8 and CD4 TRM cells, and tissue-resident natural killer cells. All lymphocytic CNS-resident cells shared expression of CXCR6 but showed differential ITGAE expression (encoding CD103). A common signature defined CD4 and CD8 TRM cells by expression of ZFP36L2, DUSP1, and ID2. We further developed a user interface-driven application based on this analysis framework for atlas-level cell identity transfer onto new CSF scRNAseq data. Together, these results define CNS-resident immune cells involved in multiple sclerosis pathology that can be detected and monitored in CSF. Targeting these cell populations might be promising to modulate immunopathology in progressive multiple sclerosis and other neuroinflammatory diseases.

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

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. A minor CD56dimGzmK+ subset was identified that shared features with CD56bright and CD56dimGzmK- NK cells based on transcriptome, phenotype (NKG2AhighCD16lowKLRG1highTIGIThigh) and functional analysis in bone marrow and blood, supportive for an intermediate subset. Pseudotime analysis positioned CD56bright, CD56dimGzmK+ and CD56dimGzmK- 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.

Barcode clonal tracking of tissue-resident immune cells in rhesus macaque highlights distinct clonal distribution pattern of tissue NK cells.

Tissue resident (TR) immune cells play important roles in facilitating tissue homeostasis, coordinating immune responses against infections and tumors, and maintaining immunological memory. While studies have shown these cells are distinct phenotypically and functionally from cells found in the peripheral blood (PB), the clonal relationship between these populations across tissues has not been comprehensively studied in primates or humans. We utilized autologous transplantation of rhesus macaque hematopoietic stem and progenitor cells containing high diversity barcodes to track the clonal distribution of T, B, myeloid and natural killer (NK) cell populations across tissues, including liver, spleen, lung, and gastrointestinal (GI) tract, in comparison with PB longitudinally post-transplantation, in particular we focused on NK cells which do not contain endogenous clonal markers and have not been previously studied in this context. T cells demonstrated tissue-specific clonal expansions as expected, both overlapping and distinct from blood T cells. In contrast, B and myeloid cells showed a much more homogeneous clonal pattern across various tissues and the blood. The clonal distribution of TR NK was more heterogenous between individual animals. In some animals, as we have previously reported, we observed large PB clonal expansions in mature CD56-CD16+ NK cells. Notably, we found a separate set of highly expanded PB clones in CD16-CD56- (DN) NK subset that were also contributing to TR NK cells in all tissues examined, both in TR CD56-CD16+ and DN populations but absent in CD56+16- TR NK across all tissues analyzed. Additionally, we observed sets of TR NK clones specific to individual tissues such as lung or GI tract and sets of TR NK clones shared across liver and spleen, distinct from other tissues. Combined with prior functional data that suggests NK memory is restricted to liver or other TR NK cells, these clonally expanded TR NK cells may be of interest for future investigation into NK cell tissue immunological memory, with implications for development of NK based immunotherapies and an understanding of NK memory.

The functional diversity of tissue-resident natural killer cells against infection.

For decades, studies of natural killer (NK) cells have focused on those found in peripheral blood (PBNK cells) as the prototype for NK cell biology. Only recently have researchers begun to explore the diversity of tissue-resident NK (tr-NK) cells. While tr-NK cells were initially identified from mice parabiosis and intravascular staining experiments, they can also be identified by tissue retention markers such as CD69, CD103 and others. More importantly, tr-NK cells have distinct functions compared to PBNK cells. Within the liver, there are diverse subsets of tr-NK cells expressing different combinations of tissue-retention markers and transcription factors, the clinical relevance of which are still unclear. Functionally, liver tr-NK are primed with immediate responsiveness to infection and equipped with regulatory mechanisms to prevent liver damage. When decidual NK (dNK) cells were first discovered, they were mainly characterized by their reduced cytotoxicity and functions related to placental development. Recent studies, however, revealed different mechanisms by which dNK cells prevent uterine infections. The lungs are one of the most highly exposed sites for infection due to their role in oxygen exchange. Upon influenza infection, lung tr-NK cells can degranulate and produce more inflammatory cytokines than PBNK cells. Less understood are gut tr-NK cells which were recently characterized in infants and adults for their functional differences. In this mini-review, we aim to provide a brief overview of the most recent discoveries on how several tr-NK cells are implicated in the immune response against infection.

Recruited and Tissue-Resident Natural Killer Cells in the Lung During Infection and Cancer.

Natural killer (NK) cells are an important component of the innate immune system, and have a key role in host defense against infection and in tumor surveillance. Tumors and viruses employ remarkably similar strategies to avoid recognition and killing by NK cells and so much can be learnt by comparing NK cells in these disparate diseases. The lung is a unique tissue environment and immune cells in this organ, including NK cells, exist in a hypofunctional state to prevent activation against innocuous stimuli. Upon infection, rapid NK cell infiltration into the lung occurs, the amplitude of which is determined by the extent of inflammation and damage. Activated NK cells kill infected cells and produce pro-inflammatory cytokines and chemokines to recruit cells of the adaptive immune system. More recent evidence has shown that NK cells also play an additional role in resolution of inflammation. In lung cancer however, NK cell recruitment is impaired and those that are present have reduced functionality. The majority of lung NK cells are circulatory, however recently a small population of tissue-resident lung NK cells has been described. The specific role of this subset is yet to be determined, but they show similarity to resident memory T cell subsets. Whether resident or recruited, NK cells are important in the control of pulmonary infections, but equally, can drive excessive inflammation if not regulated. In this review we discuss how NK cells are recruited, controlled and retained in the specific environment of the lung in health and disease. Understanding these mechanisms in the context of infection may provide opportunities to promote NK cell recruitment and function in the lung tumor setting.

Natural Killer Cells in Anti-Neutrophil Cytoplasmic Antibody-Associated Vasculitis - A Review of the Literature.

Anti-neutrophil cytoplasmic antibody (ANCA)- associated vasculitis (AAV) is a group of systemic autoimmune diseases characterized by inflammation of small- and medium-sized vessels. The three main types of AAV are granulomatosis with polyangiitis (GPA), microscopic polyangiitis (MPA) and eosinophilic granulomatosis with polyangiitis (EGPA). A growing number of studies focus on natural killer (NK) cells in AAV. NK cells are innate lymphoid cells with important roles in anti-viral and anti-tumor defense, but their roles in the pathogenesis of autoimmunity is less well established. In this review, we will present a summary of what is known about the number, phenotype and function of NK cells in patients with AAV. We review the literature on NK cells in the circulation of AAV patients, studies on tissue resident NK cells and how the treatment affects NK cells.

Methods for Isolating and Defining Single-Cell Transcriptomes of Tissue-Resident Human NK Cells.

Natural killer (NK) cells are innate lymphocytes that control tumors and microbial infections. Human NK cells are transcriptomically and phenotypically heterogeneous. The site where NK cells develop and reside determines their phenotype and effector functions. Our current knowledge about human NK cells is primarily from blood- and bone marrow-derived NK cells. The major limitation in formulating organ-specific clinical therapy is the knowledge gap on how tissue-resident NK cells develop, home, and function. Thus, it is crucial to define the transcriptomic profiles and the transcriptional regulation of tissue-resident NK cells. The major challenges in studying tissue-resident NK cells include their total number and the complexity of the tissue. Additionally, during isolation, keeping them viable and naïve without activation are challenging tasks. Here, we provide methods for isolating and performing transcriptomic analyses of NK cells at the individual cell level. Single-cell RNA sequencing provides a higher resolution of cellular heterogeneity and a better understanding of cell-cell interactions within the microenvironment. Using these methods, we can efficiently identify distinct populations of NK cells in tissues and define their unique transcriptomic profiles.