CD56bright natural killer (NK) cells: an important NK cell subset

Human natural killer (NK) cells can be subdivided into different populations based on the relative expression of the surface markers CD16 and CD56. The two major subsets are CD56^bright CD16^dim/− and CD56^dim CD16⁺, respectively. In this review, we will focus on the CD56^bright NK cell subset. These cells are numerically in the minority in peripheral blood but constitute the majority of NK cells in secondary lymphoid tissues. They are abundant cytokine producers but are only weakly cytotoxic before activation. Recent data suggest that under certain conditions, they have immunoregulatory properties, and that they are probably immediate precursors of CD56^dim NK cells. CD56^bright NK cell percentages are expanded or reduced in a certain number of diseases, but the significance of these variations is not yet clear.     

Cytokine production by CD16 CD56bright natural killer cells in the human early pregnancy decidua

Using a cell sorter, CD16^–CD56^bright natural killer (NK)cells were sorted from decidual mononuclear cells at an earlystage of pregnancy. These cells were examined by the reversetranscrlptase-polymerase chain reaction (RT-PCR) method fortheir expression of mRNA coding for the following 12 cytokines:IL-1ß, IL-2, IL-3, IL-4, IL-5, IL-6, granulocyte colony-stimulatingfactor (G-CSF), granulocyte-macrophage colony-stimulating factor(GM-CSF), macrophage colonystimulating factor (M-CSF), tumornecrosis factor- (TNF-), interferond- (IFN-), and leukemia inhibitoryfactor (LIF). Although mRNA coding for every cytokine was detectedin decidual mononuclear cells, mRNAs coding for only G-CSF,GM-CSF, M-CSF, TNF-, IFN-, and LIF were detected In CD16^–CD56^brightNK cells. Also, the supernatant of CD16^–CD56^bright NKcell cultures was found to contain G-CSF, GM-CSF, M-CSF, TNF-,IFN-, and LIF. These findings indicate that CD16^–CD56^brightNK cells produce many different cytokines and that these cytokinesmay play an important role in a successful pregnancy.     

Increased proportion of CD56bright natural killer cells in active and inactive systemic lupus erythematosus

Natural killer (NK) cells belong to the innate immune system but can also affect adaptive immune reactions. This immune regulatory function is often ascribed to the CD56(bright) subpopulation of NK cells that is prevalent in secondary lymphoid tissues and has potent cytokine-producing ability. The NK cells have been described as affecting autoimmune disease and stimulating B-cell production of antibodies, but their role in systemic lupus erythematosus (SLE) pathology has not been extensively studied. We have studied NK cells in SLE, a B-cell-driven systemic autoimmune disease, and phenotypically characterized peripheral blood NK cells in comparison to NK cells from patients with immunoglobulin A nephritis, rheumatoid arthritis and healthy individuals. We have found an increased proportion of CD56(bright) NK cells in SLE, regardless of disease activity. We detected a somewhat increased expression of the activating receptor NKp46/CD335 on NK cells from SLE patients, although neither the percentage of NK cells of all lymphocytes nor the expression of other NK receptors analysed (LIR-1/CD85j, CD94, NKG2C/CD159c, NKG2D/CD314, NKp30/CD337, NKp44/CD336, CD69) differed between patient groups. We show that type I interferon, a proinflammatory cytokine known to be abundant in SLE, can cause increases of CD56(bright) NK cells in vitro. We confirmed that serum levels of interferon-alpha were increased in active, but not in inactive, disease in the SLE patient group. In conclusion, we found an increased proportion of CD56(bright) NK cells in the blood of SLE patients, although it remains to be examined whether and how this relates to the disease process.     

Preferential apoptosis of CD56dim natural killer cell subset in patients with cancer

Natural killer (NK) cells (CD56(+)/CD3(-)) in the circulation of cancer patients were reported to have low NK activity and undergo spontaneous apoptosis. A possible relationship between apoptosis and impaired NK activity was studied by Annexin V-binding and NK-cell assays performed with peripheral blood mononuclear cells of patients with head and neck cancer (HNC), breast cancer (BC) and normal controls (NC). Cells stained with Annexin V (Anx) and antibodies to CD56, CD3, CD95, CD25, CD122 or CD132 were examined by flow cytometry. NK activity was tested against K562 targets in 4-h (51)Cr-release assays. The ratio of CD56(dim)/CD56(bright) NK cells was significantly different in patients vs. controls (10 vs. 16; p<0.01). A significantly greater percentage of CD56(dim) NK cells bound Anx in HNC patients (27+/-17%, median +/- SD) or BC (46+/-18%) than in NC (15+/-18%, p<0.04 and p<0.0002, respectively). CD56(dim) NK cells were preferentially targeted for apoptosis. NK activity was significantly lower in patients with HNC and BC than in NC (p<0.009). An inverse correlation between NK activity and the percent of Anx(+)CD56(dim) NK cells was observed in cancer patients (p =0.002) but not in NC. In patients, circulating CD56(dim) NK cells were targeted for apoptosis, leading to low levels of NK activity.     

A subset of human natural killer cells isolated and characterized by monoclonal antibodies

Monoclonal antibodies were induced against leukemic T cells from a patient with chronic lymphocytic leukemia exhibiting natural killer (NK) activity. Two antibodies, termed T811 and M522, reacted by indirect immunofluorescence with distinct subpopulations of normal human mononuclear blood cells. The antibody T811 defines a surface antigen which is restricted to a subset of the T cell lineage. The antigen recognized by the second antibody, M522, is expressed on monocytes and polymor-phonuclear leukocytes and, in addition, on 9–17% of nonadherent peripheral blood leukocytes (NAL). It is shown that the total NK activity of NAL is confined to the subset of cells expressing the M522-defined antigen. Moreover, the portion of NK cytotoxicity associated with T lymphocytes is mediated by a subpopulation which is characterized by the simultaneous expression of the T811- and the M522-defined antigens. This population comprises about 4% of NAL and could be isolated to a purity of > 85%.     

A new method for in vitro expansion of cytotoxic human CD3-CD56+ natural killer cells

Adoptive transfer of immunocompetent cells may induce anti-tumor effects in vivo. However, a significant obstacle to the development of successful cellular immunotherapy has been the availability of appropriate cytotoxic cells. Among the immunologic effector cells that are considered mediators of anti-tumor effects, those with the highest per-cell cytotoxic capacity express a natural killer (NK) cell phenotype, i.e., CD56(+)CD3(-). However, such cells are normally present only in low numbers in peripheral blood mononuclear cells (PBMCs), lymphokine activated killer (LAK), and cytokine induced killer (CIK) cell preparations. To optimize the expansion of human NK cells, PBMCs were cultured in different serum free medium supplemented with monoclonal anti-CD3 antibodies and interleukin (IL)-2 at varying concentrations. By using Cellgro stem cell growth medium supplemented with 5% human serum and IL-2 (500 U/ml) cells expanded 193-fold (median, range 21-277) after 21 days, and contained 55% (median, range 7-92) CD3(-)CD56(+) cells. The remaining cells were CD3(+) T cells, 22% (median, range 2-68) of which co-expressed CD56. The expanded cell population lysed 26 to 45% of K562 targets in a 1:1 effector to target ratio, signifying substantial cytotoxic efficacy. The described method is a simple and efficient way of expanding and enriching human NK cells. We have termed these high-yield CD3(-)CD56(+) cells cytokine-induced natural killer (CINK) cells.     

Proliferative and cytotoxic capabilities of CD16+CD56- and CD16+/-CD56+ natural killer cells

Natural killer (NK) cells can be divided into several subpopulations according to their expression of the surface antigens CD16 and CD56. The modest quantity of NK cells in the blood available for functional analysis has been a limitation in studies of NK cell subpopulations. In the present study, epinephrine infusion was used to induce lymphocytosis before immunomagnetic methods were applied to isolate CD16+/-CD56+ and CD16+CD56- CD3- NK cells. These subpopulations were compared according to their proliferative and cytotoxic capabilities in 10 human immunodeficiency virus (HIV)-infected individuals and 5 healthy controls. The CD16+CD56- NK cell subgroup had a higher proliferative capacity, whereas the CD16+/-CD56+ NK cell subgroup was mainly cytotoxic, and unaffected by HIV serostatus. This study thus suggests that NK cell phenotypes more strongly predict NK cell function than HIV serostatus. This assertion should be considered when studying NK cell function in subjects with a deviating composition of NK cells.     

Generation of natural killer cells from serum-free, expanded human umbilical cord blood CD34+ cells

Natural killer (NK) cells are important effectors of the innate immune system, which exhibits cytolytic activity against infectious agents and tumor cells. NK cells are derived from CD34(+) hematopoietic stem cells (HSCs). Human umbilical cord blood (UCB) has been recognized as a rich source of HSCs. Previously, we have reported an optimized serum-free medium for ex vivo expansion of CD34(+) cells from UCB. In this study, the serum-free, expanded CD34(+) cells were tested to differentiate into NK cells and their induction kinetics. After 5 weeks of induction, the induced NK cells were characterized by analysis of surface antigens, IFN-gamma secretion, and cytotoxicity against K562 cells. The results indicated that NK cells derived from the serum-free, expanded CD34(+) cells exhibited both characteristics and functions of NK cells. Furthermore, the serum-free, expanded CD34(+) cells showed a significantly higher NK cell differentiation potential than freshly isolated CD34(+) cells. NK cells induced from serum-free, expanded CD34(+) cells showed a higher concentration of IFN-gamma secretion and ability of cytotoxicity than those from freshly isolated CD34(+) cells. Therefore, ex vivo-expanded CD34(+) cells in optimized serum-free medium could differentiate into NK cells and provided a promising cell source for immunotherapeutic approaches.     

CD56bright cells differ in their KIR repertoire and cytotoxic features from CD56dim NK cells

In this study we present new differential characteristics of NK cells expressing CD56 surface antigen in low (CD56dim) or high (CD56bright) density. In contrast to CD56bright NK cells CD56dim cells express killer cell immunoglobulin (Ig)-like receptors (KIR) such as CD158a, CD158b, and NKB1. However, c-type lectin-like receptors (KLR) CD94/NKG2 and CD161 are present on both subsets. The ability to form conjugates with susceptible targets is approximately twice as strongly pronounced in CD56dim vs. CD56bright NK cells. Last but not least, granules of CD56dim cells contain about tenfold more perforin and granzyme A enabling potentially more effective cytolysis compared to CD56bright NK cells. On the other hand, CD56bright NK cells are superior in producing the proinflammatory cytokines IFN-gamma (28.5% vs. 20.8%, p<0.05) and TNF-alpha (28% vs. 15.8%, p<0.001). The different NK cell populations retained their specific phenotype in vitro during culture in the presence of IL-2 contradicting that they simply display different stages of maturity. Taken together our data support the view that CD56bright cells are specialized NK cells that regulate immunological response mechanisms rather by cytokine supply than by their cytotoxic potential. The poor cytolytic capacity of CD56bright NK cells can be explained by weak ability in forming conjugates with target cells and low contents of perforin and granzyme A in their granules.     

Impaired natural killer cell subset phenotypes in human obesity

Obesity is associated with alterations in functionality of immune cells, like macrophages and natural killer (NK) cells, leading to an increased risk for severe infections and several cancer types. This study aimed to examine immune cell populations and functional NK cell parameters focusing on NK cell subset phenotypes in normal-weight and obese humans. Therefore, peripheral blood mononuclear cells (PBMCs) were isolated from normal-weight and obese individuals and analyzed by flow cytometry. Results show no significant changes in the frequency of monocytes, B lymphocytes, or NKT cells but a significantly increased frequency of T lymphocytes in obesity. The frequency of total NK cells was unaltered, whereas the number of low cytotoxic CD56^bright NK cell subset was increased, and the number of high cytotoxic CD56^dim NK cell subset was decreased in obese subjects. In addition, the frequency of CD56^bright NK cells expressing the activating NK cell receptor NKG2D as well as intracellular interferon (IFN)-γ was elevated in the obese study group. In contrast, the frequency of NKG2D- and IFN-γ-positive CD56^dim NK cells was lower in obesity compared to normal-weight individuals. Moreover, the expression of the activation marker CD69 was decreased in NK cells, which can be attributed to a reduction of CD69-positive CD56^dim NK cells in obese subjects. In conclusion, data reveal an impaired NK cell phenotype and NK cell subset alterations in obese individuals. This NK cell dysfunction might be one link to the higher cancer risk and the elevated susceptibility for viral infections in obesity.     

Murine CXCR3+CD27bright NK cells resemble the human CD56bright NK‐cell population

Human NK cells can be subdivided into CD56^dim and CD56^bright NK cells, which exhibit different phenotypical and functional characteristics. As murine NK cells lack CD56 or a distinct correlate, direct comparative studies of NK cells in mice and humans are limited. Although CD27 is currently proposed as a feasible subset marker in mice, we assume that the usage of this marker alone is insufficient. We rather investigated the expression of the chemokine receptor CXCR3 for its suitability for distinguishing murine NK‐cell subsets with simultaneous consideration of CD27. Compared with CXCR3^? NK cells, exerting stronger cytotoxic capability, CXCR3⁺ NK cells displayed an activated phenotype with a lower expression of Ly49 receptors, corresponding to human CD56^bright NK cells. Also in common with human CD56^bright NK cells, murine CXCR3⁺ NK cells exhibit prolific expansion as well as robust IFN‐γ, TNF‐α and MIP‐1α production. We additionally demonstrated changes in both CXCR3 and CD27 expression upon NK‐cell activation. In summary, CXCR3 serves as an additional applicable marker for improved discrimination of functionally distinct murine NK‐cell subsets that comply with those in humans.     

Functional and Vbeta repertoire characterization of human CD8+ T-cell subsets with natural killer cell markers,CD56+ CD57- T cells,CD56+ CD57+ T cells and CD56- CD57+ T cells

We investigated the individual CD8+ populations with natural killer (NK) cell markers (NK-type T cell); CD56 single positive (CD56)-T cells, CD56/CD57 double positive (DP)-T cells and CD57 single positive (CD57)-T cells in the peripheral blood. All NK-type T-cell populations expressed CD122 and intermediate levels of T-cell receptor (TCR; regular CD8+ T cells are CD122- and express high levels of TCR). The number of both DP-T cells and CD57-T cells, but not CD56-T cells, gradually increased with age. All NK-type T-cell populations produced larger amounts of interferon-gamma than did regular CD8+ T cells after stimulation with interleukin (IL)-2, IL-12 and IL-15. However, CD56-T cells and CD57-T cells but not DP-T cells showed a potent antitumour cytotoxity to NK-sensitive K562 cells, whereas only CD56-T cells showed a potent cytotoxity to NK-resistant Raji cells. Furthermore, although NK-type T cells produced large amounts of soluble Fas-ligands, their cytotoxic activities appeared to be mediated by the perforin/granzyme pathway. The oligoclonal or pauciclonal expansions of certain VbetaT cells were found in each NK-type T-cell population. The non-variant CDR3 region(s) for the TCRbeta chain(s) showed CD57-T cells and CD56-T cells to be derived from distinct origins, while the DP-T cell population consisted of a mixture of the clones seen in both CD56-T cells and CD57-T cells. Our results suggest that CD57-T cells and CD56-T cells are functionally and ontogenically different populations while DP-T cells appear to originate from both CD56-T cells and CD57-T cells.     

CD56bright natural killer cell subsets: Characterization of distinct functional responses to interleukin-2 and the c-kit ligand

Natural killer (NK) cells are bone marrow-derived large granular lymphocytes that express the CD56 surface antigen. The CD56^bright NK subset represents approximately 10 % of all NK cells and is thought to be the least differentiated NK cell component in blood. The most mature NK cell expresses CD56 at low density and CD16 (FcRγIII) at high density, whereas CD56^bright NK cells either lack CD 16 (CD56^brightCD16^?) or express it at low density (CD56^brightCD16^dim). c-kit is a tyrosine kinase receptor which is expressed on both CD34⁺ hemato-poietic precursor cells and CD56^bright NK cells. In the current study, we characterize interleukin (IL)-2 receptor (IL-2R) and c-kit expression in each of the CD56^bright subsets. Both the CD56^brightCD16^? and CD56^brightCD16^dim NK subsets express the high-affinity IL-2R and the c-kit receptor when isolated from fresh blood. However, each CD56^bright NK cell subset has distinct functional responses to IL-2, the c-kit ligand (KL), or both. Activation of the high-affinity IL-2R on CD56^brightCD16^? NK cells induces a proliferative response that is significantly weaker than that observed in the CD56^brightCD16^dim NK cell subset. Incubation of the CD56^brightCD16^? NK cell subset with KL significantly enhances IL-2-induced proliferation, while KL has no such effect on the CD56^brightCD16^dim NK subset. Activation of the high-affinity IL-2R in both CD56^bright subsets induces lymphokine-activated killer (LAK) activity, but the addition of KL has no effect on LAK activity. Co-stimulation of either CD56^bright subset with IL-12 and concentrations of IL-2 that only saturate the high-affinity IL-2R induces substantial interferon (IFN)-γ production. The addition of KL to this co-stimulatory signal enhances IFN-γ production in both CD56^bright NK subsets. The distinct functional responses to IL-2 and KL seen in the CD56^brightCD16^? and CD56^brightCD16^dim NK subsets provide insight into IL-2R signaling and suggest that each phenotype identifies a discrete stage of NK cell differentiation.     

Functional expression of CD43 on human natural killer cells

CD43 is the major leukocyte sialoglyco-protein that plays important functional roles in neutrophils and lymphocytes. However, the expression of CD43 on human natural killer (NK) cells and its participation in the regulation of NK activity has not been studied. We have therefore investigated the expression of CD43 isoforms on human NK cell subpopulations as well as the role of this molecule in NK cell activation and cytotoxicity. We found that CD56bright and CD56dim NK cells express different sialylated forms of CD43, observing that activation of the CD56bright NK cells induces the change of tetrasaccharide O-glycans to hexasaccharide O-glycans on CD43. Cross-linking of the molecule with mAbs results in a metalloprotease-dependent loss of CD43 from the NK cell surface, whereas soluble anti-CD43 mAbs induce a vigorous NK cell proliferation. This property is distinct from T cells, which proliferate after CD43 cross-linking only in the presence of monocytes. Occupancy of the CD43 receptor on NK cells transduces specific signals, leading to enhanced killing activity and tyrosine phosphorylation and de-phosphorylation of several substrates. We therefore propose that CD43 significantly contributes to the regulation of the NK cell function by participating in the control of effector/target interactions and, if pertinent, by transducing activation signals.     

beta-cell antigen-specific CD56(+) NKT cells from type 1 diabetic patients: autoaggressive effector T cells damage human CD56(+) beta cells by HLA-restricted and non-HLA-restricted pathways

Studies of type 1 diabetes indicate that autoaggressive T cells specific to beta-cell antigens, reaching certain threshold levels, may play critical roles in the development of the disease. Flow cytometric analyses found that autoreactive T-cell lines from patients induced by beta-cell antigens consisted of four major subsets (CD4(+)CD56(-), CD4(+)CD56(+), CD8(+)CD56(-), and CD8(+)CD56(+)) and that CD56(+) NKT cells might be derived from CD56(-) T cells. Moreover, the proportion of CD56(+) NKT cells in the T-cell lines was influenced by time course of repeated antigen stimulation. beta-cell antigen-specific CD56(+) NKT (CD4(+) or CD8(+)) cells were more aggressive (HLA-restricted and -unrestricted) effector cells lysing target cells such as K562, Jurkat, P815 plus anti-CD3 antibody, and autologous B cells sensitized by beta-cell peptides, when compared with their CD56(-) counterparts. beta-cell antigen- specific CD4(+)CD56(+) NKT cells showed non-HLA-restricted cytotoxicity to human beta cells, insulinoma cell line CM, and to islet cell lines TRM-6 and HP62 expressing CD56 but not to four CD56(-) pancreatic cell lines of non- islet origin. The CD4(+)CD56(+) NKT cells showed stronger cytotoxicity to CM, TRM-6 and HP62 cells than did CD4(+)CD56(-) T cells. Moreover, isotope-unlabelled CD56(+) cells and anti-CD56 antibodies were able to inhibit cytotoxicity of CD4(+)CD56(+) NKT to CD56(+) target cells. These results suggest that CD56(+) NKT cells are aggressive cytotoxic cells to beta cells and that CD56 expression might be associated with the aggressiveness of effector T cells and the susceptibility of target cells.     

Abundance of unconventional CD8(+) natural killer T cells in the large intestine.

Natural killer T (NKT) cells are mainly present in the liver and thymus, and the majority of these T cells express either a CD4(+) or a double-negative (DN) CD4(-)8(-) phenotype. In the present study, we examined whether such NKT cells were present in the intestine. NKT cells were rare in all sites of the small intestine, including an intraepithelial site. However, a considerable number of NKT cells were found at an intraepithelial site in the large intestine. This result was confirmed by both immunofluorescence and immunohistochemistry. In contrast to conventional NKT cells, NKT cells in the large intestine were CD8(+) or DN CD4(-)8(-). In the case of conventional NKT cells, their existence is known to depend on non-classical MHC class I-like antigens (i. e. CD1d) but not on classical MHC class I antigens. However, the NKT cells in the large intestine were independent of the presence of both CD1d and classical MHC class I antigens. These results were obtained using knockout mice lacking the corresponding genes and molecules. NKT cells in the large intestine were mainly alpha betaTCR(+) (> 75 %) but did not use an invariant chain of Valpha14Jalpha281, which is preferentially used by conventional NKT cells. These NKT cells did not bias the TCR-Vbeta usage toward Vbeta8. These findings suggest that the large intestine is a site in which unconventional NKT cells carrying the CD8(+) phenotype (or DN CD4(-)8(-)) are abundant and that these cells are independent of MHC and MHC-like antigens.     

A multi-step isolation scheme for obtaining CD16+ human natural killer cells

A multi-step isolation scheme capitalizing on negative selection protocols is described for obtaining an enriched population of CD16+ human natural killer (NK) cells. The isolation scheme consists of incubating peripheral blood mononuclear cells (MNC) on nylon wool, rosetting the nylon wool non-adherent cells with sheep red blood cells (SRBCs) for 1 h at 29 degrees C and then utilizing a 'panning' technique to remove CD3+, non-rosetting cells. The final working cell population contained 70-80% CD16+ cells, 15% CD2+ cells, 1-3% CD3+ cells, 5-7% SIg+ cells and no detectable MO2+ cells. In comparing the final NK cell population from the multi-step isolation protocol to NK cells obtained by the Percoll density gradient centrifugation technique, the multistep method: (1) yielded a higher percentage of CD16+ cells, (2) mediated a greater degree of cytotoxicity at a 25:1 E:T ratio, and (3) contained fewer contaminating monocytes/macrophages (none were detectable). In addition, the multi-step scheme allowed recovery of 30% of the total CD16+ cells present compared to only 7% recovered by the Percoll density gradient technique. Pretreatment of the enriched NK cells, obtained from the multi-step scheme, with interleukin-2 (3.5 and 7.0 U/ml of activity) resulted in an increase in NK cell-mediated cytotoxicity. In addition, these cells were as effective at synthesizing the cytotoxin, NKCF, at a 25:1 E:T ratio as at 50:1 and 100:1 E:T ratios. This multi-step isolation scheme consistently yields a high percentage of CD16+ NK cells and thus may greatly facilitate studies on the mechanism(s) involved in NK cell-mediated cytotoxicity and may further the study of the cytotoxins involved.     

Systematic characterization of human CD8+ T cells with natural killer cell markers in comparison with natural killer cells and normal CD8+ T cells

We investigated the function of CD56+ CD8+ T cells (CD56+ T cells) and CD56- CD57+ CD8+ T cells (CD57+ T cells; natural killer (NK)-type T cells) and compared them with those of normal CD56- CD57- CD8+ T cells (CD8+ T cells) and CD56+ NK cells from healthy volunteers. After the stimulation with immobilized anti-CD3 antibodies, both NK-type T cells produced much larger amounts of interferon-gamma (IFN-gamma) than CD8+ T cells. Both NK-type T cells also acquired a more potent cytotoxicity against NK-sensitive K562 cells than CD8+ T cells while only CD56+ T cells showed a potent cytotoxicity against NK-resistant Raji cells. After the stimulation with a combination of interleukin (IL)-2, IL-12 and IL-15, the IFN-gamma amounts produced were NK cells > or = CD56+ T cells > or = CD57+ T cells > CD8+ T cells. The cytotoxicities against K562 cells were NK cells > CD56+ T cells > or = CD57+ T cells > CD8+ T cells while cytotoxicities against Raji cells were CD56+ T cells > CD57+ T cells > or = CD8+ T cells > or = NK cells. However, the CD3-stimulated proliferation of both NK-type T cells was smaller than that of CD8+ T cells partly because NK-type T cells were susceptible to apoptosis. In addition to NK cells, NK-type T cells but not CD8+ T cells stimulated with cytokines, expressed cytoplasmic perforin and granzyme B. Furthermore, CD3-stimulated IFN-gamma production from peripheral blood mononuclear cells (PBMC) correlated with the proportions of CD57+ T cells in PBMC from donors. Our findings suggest that NK-type T cells play an important role in the T helper 1 responses and the immunological changes associated with ageing.