natural killer cell natural killer cell natural killer cells, also known as nk cells or large granular lymphocytes (lgl), are a type of cytotoxic lymphocyte critical to the innate immune system that belong to the rapidly expanding family of known innate lymphoid cells (ilc) and represent 5–20% of all circulating lymphocytes in humans. the role of nk cells is analogous to that of cytotoxic t cells in the vertebrate adaptive immune response. nk cells provide rapid responses to virus-infected cell and other intracellular pathogens acting at around 3 days after infection, and respond to tumor formation. typically, immune cells detect the antigen presented on major histocompatibility complex (mhc) on infected cell surfaces, triggering cytokine release, causing the death of the infected cell by lysis or apoptosis. nk cells are unique, however, as they have the ability to recognize and kill stressed cells in the absence of antibodies and mhc, allowing for a much faster immune reaction. they were named "natural killers" because of the notion that they do not require activation to kill cells that are missing "self" markers of mhc class 1. this role is especially important because harmful cells that are missing mhc i markers cannot be detected and destroyed by other immune cells, such as t lymphocyte cells. not to be confused with natural killer t cell. natural killer cellhuman natural killer cell, colorized scanning electron micrographdetailssystemimmune systemfunctioncytotoxic lymphocyteidentifiersmeshd007694fma63147anatomical terms of microanatomynk cells can be identified by the presence of cd56 and the absence of cd3 (cd56+, cd3−). nk cells (belonging to the group of innate lymphoid cells) are one of the three kinds of cells differentiated from the common lymphoid progenitor, the other two being b and t lymphocytes. nk cells are known to differentiate and mature in the bone marrow, lymph nodes, spleen, tonsils, and thymus, where they then enter into the circulation. nk cells differ from natural killer t cells (nkts) phenotypically, by origin and by respective effector functions; often, nkt cell activity promotes nk cell activity by secreting interferon gamma. in contrast to nkt cells, nk cells do not express t-cell antigen receptors (tcr) or pan t marker cd3 or surface immunoglobulins (ig) b cell receptors, but they usually express the surface markers cd16 (fcγriii) and cd57 in humans, nk1.1 or nk1.2 in c57bl/6 mice. the nkp46 cell surface marker constitutes, at the moment, another nk cell marker of preference being expressed in both humans, several strains of mice (including balb/c mice) and in three common monkey species. in addition to natural killer cells being effectors of innate immunity, both activating and inhibitory nk cell receptors play important functional roles, including self tolerance and the sustaining of nk cell activity. nk cells also play a role in the adaptive immune response: numerous experiments have demonstrated their ability to readily adjust to the immediate environment and formulate antigen-specific immunological memory, fundamental for responding to secondary infections with the same antigen. the role of nk cells in both the innate and adaptive immune responses is becoming increasingly important in research using nk cell activity as a potential cancer therapy. early history in early experiments on cell-mediated cytotoxicity against tumor target cells, both in cancer patients and animal models, investigators consistently observed what was termed a "natural" reactivity; that is, a certain population of cells seemed to be able to lyse tumor cells without having been previously sensitized to them. the first published study to assert that untreated lymphoid cells were able to confer a natural immunity to tumors was performed by dr. henry smith at the university of leeds school of medicine in 1966, leading to the conclusion that the "phenomenon appear to be an expression of defense mechanisms to tumor growth present in normal mice." other researchers had also made similar observations, but as these discoveries were inconsistent with the established model at the time, many initially considered these observations to be artifacts. by 1973, 'natural killing' activity was established across a wide variety of species, and the existence of a separate lineage of cells possessing this ability was postulated. the discovery that a unique type of lymphocyte was responsible for "natural" or spontaneous cytotoxicity was made in the early 1970s by doctoral student rolf kiessling and postdoctoral fellow hugh pross, in the mouse, and by hugh pross and doctoral student mikael jondal in the human. the mouse and human work was carried out under the supervision of professors eva klein and hans wigzell, respectively, of the karolinska institute, stockholm. kiessling's research involved the well-characterized ability of t lymphocytes to lyse tumor cells against which they had been previously immunized. pross and jondal were studying cell-mediated cytotoxicity in normal human blood and the effect of the removal of various receptor-bearing cells on this cytotoxicity. later that same year, ronald herberman published similar data with respect to the unique nature of the mouse effector cell. the human data were confirmed, for the most part, by west et al. using similar techniques and the same erythroleukemic target cell line, k562. k562 is highly sensitive to lysis by human nk cells and, over the decades, the k562 51chromium-release assay has become the most commonly used assay to detect human nk functional activity. its almost universal use has meant that experimental data can be compared easily by different laboratories around the world. using discontinuous density centrifugation, and later monoclonal antibodies, natural killing ability was mapped to the subset of large, granular lymphocytes known today as nk cells. the demonstration that density gradient-isolated large granular lymphocytes were responsible for human nk activity, made by timonen and saksela in 1980, was the first time that nk cells had been visualized microscopically, and was a major breakthrough in the field. nk cell subsets nk cells can be classified as cd56bright or cd56dim. cd56bright nk cells are similar to t helper cells in exerting their influence by releasing cytokines. cd56bright nk cells constitute the majority of nk cells, being found in bone marrow, secondary lymphoid tissue, liver, and skin. cd56dim nk cells are primarily found in the peripheral blood, and are characterized by their cell killing ability. cd56dim nk cells are always cd16 positive (cd16 is the key mediator of antibody-dependent cellular cytotoxicity (adcc). cd56bright can transition into cd56dim by acquiring cd16. nk cells can eliminate virus-infected cells via cd16-mediated adcc. all coronavirus disease 2019 (covid-19) patients show depleted cd56bright nk cells, but cd56dim is only depleted in patients with severe covid-19. nk cell receptors the hla ligand for kir nk cell receptors can also be differentiated based on function. natural cytotoxicity receptors directly induce apoptosis (cell death) after binding to fas ligand that directly indicate infection of a cell. the mhc-independent receptors (described above) use an alternate pathway to induce apoptosis in infected cells. natural killer cell activation is determined by the balance of inhibitory and activating receptor stimulation. for example, if the inhibitory receptor signaling is more prominent, then nk cell activity will be inhibited; similarly, if the activating signal is dominant, then nk cell activation will result. protein structure of nkg2d nk cell receptor types (with inhibitory, as well as some activating members) are differentiated by structure, with a few examples to follow: protein structure of nkp44 activating receptors ly49 (homodimers), relatively ancient, c-type lectin family receptors, are of multigenic presence in mice, while humans have only one pseudogenic ly49, the receptor for classical (polymorphic) mhc i molecules. ncr (natural cytotoxicity receptors), type 1 transmembrane proteins of the immunoglobulin superfamily, upon stimulation mediate nk killing and release of ifnγ. they bind viral ligands such as hemagglutinins and hemagglutinin neuraminidases, some bacterial ligands and cellular ligands related to tumour growth such as pcna. cd16 (fcγiiia) plays a role in antibody-dependent cell-mediated cytotoxicity; in particular, they bind immunoglobulin g. inhibitory receptors killer-cell immunoglobulin-like receptors (kirs) belong to a multigene family of more recently evolved ig-like extracellular domain receptors; they are present in nonhuman primates, and are the main receptors for both classical mhc i (hla-a, hla-b, hla-c) and nonclassical mamu-g (hla-g) in primates. some kirs are specific for certain hla subtypes. most kirs are inhibitory and dominant. regular cells express mhc class 1, so are recognised by kir receptors and nk cell killing is inhibited. cd94/nkg2 (heterodimers), a c-type lectin family receptor, is conserved in both rodents and primates and identifies nonclassical (also nonpolymorphic) mhc i molecules such as hla-e. expression of hla-e at the cell surface is dependent on the presence of nonamer peptide epitope derived from the signal sequence of classical mhc class i molecules, which is generated by the sequential action of signal peptide peptidase and the proteasome. though indirect, this is a way to survey the levels of classical (polymorphic) hla molecules. ilt or lir (immunoglobulin-like receptor) — are recently discovered members of the ig receptor family. ly49 (homodimers) have both activating and inhibitory isoforms. they are highly polymorphic on the population level; though they are structurally unrelated to kirs, they are the functional homologues of kirs in mice, including the expression pattern. ly49s are receptor for classical (polymorphic) mhc i molecules. function cytolytic granule mediated cell apoptosis nk cells are cytotoxic; small granules in their cytoplasm contain proteins such as perforin and proteases known as granzymes. upon release in close proximity to a cell slated for killing, perforin forms pores in the cell membrane of the target cell, creating an aqueous channel through which the granzymes and associated molecules can enter, inducing either apoptosis or osmotic cell lysis. the distinction between apoptosis and cell lysis is important in immunology: lysing a virus-infected cell could potentially release the virions, whereas apoptosis leads to destruction of the virus inside. α-defensins, antimicrobial molecules, are also secreted by nk cells, and directly kill bacteria by disrupting their cell walls in a manner analogous to that of neutrophils. antibody-dependent cell-mediated cytotoxicity (adcc) infected cells are routinely opsonized with antibodies for detection by immune cells. antibodies that bind to antigens can be recognised by fcγriii (cd16) receptors expressed on nk cells, resulting in nk activation, release of cytolytic granules and consequent cell apoptosis. this is a major killing mechanism of some monoclonal antibodies like rituximab (rituxan), ofatumumab (azzera), and others. the contribution of antibody-dependent cell-mediated cytotoxicity to tumor cell killing can be measured with a specific test that uses nk-92, an immortal line of nk-like cells licensed to nantkwest, inc.: the response of nk-92 cells that have been transfected with a high-affinity fc receptor are compared to that of the "wild type" nk-92 which does not express the fc receptor. cytokine-induced nk and cytotoxic t lymphocyte (ctl) activation cytokines play a crucial role in nk cell activation. as these are stress molecules released by cells upon viral infection, they serve to signal to the nk cell the presence of viral pathogens in the affected area. cytokines involved in nk activation include il-12, il-15, il-18, il-2, and ccl5. nk cells are activated in response to interferons or macrophage-derived cytokines. they serve to contain viral infections while the adaptive immune response generates antigen-specific cytotoxic t cells that can clear the infection. nk cells work to control viral infections by secreting ifnγ and tnfα. ifnγ activates macrophages for phagocytosis and lysis, and tnfα acts to promote direct nk tumor cell killing. patients deficient in nk cells prove to be highly susceptible to early phases of herpes virus infection. missing 'self' hypothesis schematic diagram indicating the complementary activities of cytotoxic t cells and nk cells for nk cells to defend the body against viruses and other pathogens, they require mechanisms that enable the determination of whether a cell is infected or not. the exact mechanisms remain the subject of current investigation, but recognition of an "altered self" state is thought to be involved. to control their cytotoxic activity, nk cells possess two types of surface receptors: activating receptors and inhibitory receptors, including killer-cell immunoglobulin-like receptors. most of these receptors are not unique to nk cells and can be present in some t cell subsets, as well. the inhibitory receptors recognize mhc class i alleles, which could explain why nk cells preferentially kill cells that possess low levels of mhc class i molecules. this mode of nk cell target interaction is known as "missing-self recognition", a term coined by klas kärre and co-workers in the late 90s. mhc class i molecules are the main mechanism