PD-1 Expression on Natural Killer Cells and CD8+ T Cells During Chronic HIV-1 Infection
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Abstract:
Programmed death receptor 1 (PD-1) is an important marker of T-cell exhaustion during HIV-1 infection. Natural killer (NK) cells lose their functional capacity during HIV-1 infection, and PD-1 is expressed on NK cells during other chronic viral and bacterial infections. Here, PD-1 expression was increased on NK cells from both viremic and aviremic HIV-1-seropositive individuals, compared to seronegative controls. However, PD-1 was expressed on a small subset of NK cells and at lower frequency than that observed for CD8+ T cells. PD-1 was also induced on a minor fraction of NK cells and CD8+ T cells after long-term culture with IL-15. Raised levels of PD-1 were associated with limited NK cell proliferation, which may have consequences for their maintenance during chronic HIV-1 infection.Keywords:
Chronic infection
Abstract NK cell populations were derived from murine splenocytes stimulated by IL-2, IL-15, or the combination of IL-12 and IL-18. Whereas NK cells derived with the latter cytokines consisted of an homogeneous population of NK cells (DX5+CD3−), those derived with IL-2 or IL-15 belonged to two different populations, namely NK cells (DX5+CD3−) and T-NK cells (DX5+CD3+). Among NK cells, only those derived with IL-12/IL-18 produced detectable levels of cytokines, namely IFN-γ, IL-10, and IL-13 (with the exception of IL-13 production by NK cells derived with IL-2). As for T-NK cells, IL-2-stimulated cells produced a wide range of cytokines, including IL-4, IL-5, IL-9, IL-10, and IL-13, but no IFN-γ, whereas IL-15-derived T-NK cells failed to produce any cytokine. Switch-culture experiments indicated that T-NK cells derived in IL-2 and further stimulated with IL-12/IL-18 produced IFN-γ and higher IL-13 levels. Next, we observed that NK/T-NK cell populations exerted distinct effects on Ig production by autologous splenocytes according to the cytokines with which they were derived. Thus, addition of NK cells derived in IL-12/IL-18 inhibited Ig production and induced strong cytotoxicity against splenocytes, whereas addition of NK or T-NK cells grown in IL-2 or IL-15 did not. Experiments performed in IFN-γR knockout mice demonstrated that IFN-γ was not involved in the killer activity of IL-12/IL-18-derived NK cells. The hypothesis that their cytotoxic activity was related to the induction of target apoptosis was confirmed on murine A20 lymphoma cells. Experiments performed in MRL/lpr mice indicated that IL-12/IL-18-derived NK cells displayed their distinct killer activity through a Fas-independent pathway. Finally, perforin was much more expressed in IL-12/IL-18-derived NK cells as compared with IL-2- or IL-15-derived NK cells, an observation that might explain their unique cytotoxicity.
Janus kinase 3
Lymphokine-activated killer cell
Interleukin 15
CD49b
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Abstract Natural Killer (NK) cells are an important component of the innate immune system, capable of providing a fast and effective response against virally infected cells. NK cells are mainly characterized by their cytotoxic function and their ability to secrete cytokines. It has been shown that infant NK cells have decreased cytotoxicity and cytokine-secreting function, suggesting hyporesponsiveness of NK cells during the first year of life. Because NK cell activation is dependent on cytokine stimulation, we hypothesized that infant NK cells are hyporesponsive due to deficiencies in signaling following cytokine stimulation. We examined the activation of human infant and adult NK cells in response to IL2, IL12 or IFNα stimulation, cytokines important in NK survival and function. Using Phosflow analysis, we measured the phosphorylation of transcription factors, STATs, specific for the distinct cytokines. Cord blood NK cells showed significantly lower pSTAT activation when compared to adult NK cells when treated with IL2 or IFNα but not IL12. However, despite pSTAT4 activation in response to IL12, no nuclear translocation was observed by ImageStreamX Mark II analysis. NK cells in 6 and 12-month-old infants showed similar frequencies of pSTAT NK cells compared to adults when treated with IL2, IL12 or IFNα, suggesting that cytokine signaling in NK cells is age-dependent. Surprisingly, pSTAT activation was restored in cord blood NK cells when treated with a cocktail of IL2, IL12 and IL15. These results suggest that JAK/STAT signaling in infant NK cells is impaired at different steps in the pathway in response to specific cytokines.
Janus kinase 3
Lymphokine-activated killer cell
Interleukin 15
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Interleukin 15
Janus kinase 3
Lymphokine-activated killer cell
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Janus kinase 3
Lymphokine-activated killer cell
NK-92
CD49b
Interleukin 3
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Objective: To study the effects of IL-2 and IL-15 on the expression of NKG2D and the cytotoxicity of edited-NK cells against human nasopharyngeal carcinoma cell line CNE2. Methods: NK cells were purified by anti-CD56 MACS and were divided into four groups: non-edited-NK cells group (NK cells treated with 100 U/ml IL-2), edited-NK cells group (NK cells co-cultured with CNE2 cells at a ratio of 10:1 and then treated with 100 U/ml IL-2), edited-NK cells retreated with 1 000 U/ml IL-2 group, and edited-NK cells retreated with 10 ng/ml IL-15 group. Expression of NKG2D in each group was determined by FACS 24 h later. Cytotoxicity of NK cells against CNE2 cells (NK:CNE2 being 20:1) was measured by LDH releasing assay. Results: The expression of NKG2D in non-edited-NK cells, edited-NK cells, edited-NK cells retreated with IL-2, and edited-NK cells retreated with IL-15 were (97.63±0.83)%, (53.50±1.25)%, (94.47±1.00)%, and (98.07±0.21)%, respectively. The expression of NKG2D on edited-NK cells retreated with IL-2 or IL-15 was significantly increased than that on edited-NK cells (P＜0.01). The cytotoxicity of non-edited-NK cells, edited-NK cells, edited-NK cells retreated with IL-2, and edited-NK cells retreated with IL-15 against CNE2 cells were (35.90±3.27)%, (4.70±2.30)%, (31.70±3.56)% and (40.18±2.94)%, respectively. The cytotoxicity of edited-NK cells was significantly enhanced after retreated with IL-2 or IL-15 (P＜0.01), with those retreated with IL-15 being stronger than those retreated with IL-2. Conclusion: High dose IL-2 and IL-15 can up-regulate the expression of NKG2D on edited-NK cells and restore their cytotoxicity against CNE2 cells, with the efficacy of IL-15 stronger than that of IL-2.
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Summary 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‐γ (IFN‐γ) 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‐γ amounts produced were NK cells ≥ CD56 + T cells ≥ CD57 + T cells > CD8 + T cells. The cytotoxicities against K562 cells were NK cells > CD56 + T cells ≥ CD57 + T cells > CD8 + T cells while cytotoxicities against Raji cells were CD56 + T cells > CD57 + T cells ≥ CD8 + T cells ≥ 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‐γ 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.
Interleukin 3
Janus kinase 3
Lymphokine-activated killer cell
CD49b
Granzyme
K562 cells
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Abstract The role of IL-4 in proliferation and differentiation of human NK cells was studied using newly established sublines of an IL-4-dependent NK cell clone (IL4d-NK cells) and an IL-2-dependent NK cell clone (IL2d-NK cells) derived from a parental conditioned medium-dependent NK cell clone (CM-NK cells). IL-4 induced the higher proliferation of CM-NK cells, but abolished their NK activity and decreased CD16 and CD56 Ag expression. In contrast, IL-2 induced the higher NK activity and increased CD16 and CD56 Ag expression. Addition of anti-IL-4 antibody to the culture of CM-NK cells with CM inhibited the proliferation, but slightly increased NK activity, and largely increased CD56 Ag expression. Addition of anti-IL-2 antibody to the culture of CM-NK cells with CM inhibited both proliferation and cytotoxicity. Proliferation of IL4d-NK cells, which is totally dependent on rIL-4, is greater than that of IL2d-NK cells, which was greater than parental CM-NK cells. Morphologically, IL4d-NK cells are small and round, whereas IL2d-NK cells are large and elongated. Anti-IL-4 antibody inhibited proliferation of IL4d-NK but not IL2d-NK cells, whereas anti-IL-2 antibody inhibited that of IL2d-NK but not IL4d-NK cells. IL-2 was not detected in the supernatant from IL4d-NK cells, nor was IL-2-mRNA expressed in IL4d-NK cells. In contrast, IFN-gamma production and protein expression in IL4d- and IL2d-NK cells were detected. NK cell activation markers (CD16 and CD56) were expressed on IL2d-NK cells but not IL4d-NK cells. IL4d-NK cells were not cytotoxic to any tumor cells tested, whereas IL2d-NK cells displayed potent NK activity and lymphokine-activated killer activity. IL4d-NK cells failed to bind K562 tumor cells, whereas one-third of the IL2d-NK cells did. IL4d-NK cells responded to rIL-2, proliferated, and differentiated into cytotoxic NK cells, whereas IL2d-NK cells failed to respond to rIL-4 and died. These results raise a possibility that IL4d-NK cells or IL2d-NK cells primarily represent the immunologic properties of immature or activated types of human NK cells, respectively. Our results provide the first evidence of the capability of IL-4 to support continuous proliferation of a lymphocyte clone with immature NK cell characteristics and to stimulate IFN-gamma production in the clone. IL-4 is suggested as a potential growth factor for certain types of human NK cell progenitors.
Janus kinase 3
Lymphokine-activated killer cell
clone (Java method)
CD16
Interleukin 15
NK-92
CD49b
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Janus kinase 3
NK-92
Lymphokine-activated killer cell
CD49b
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The NK-1.1(-) mouse was constructed by weekly injections of monoclonal anti-NK-1.1 antibody from birth through adulthood. Spleen cells from these mice have decreased NK-1.1+ cells and null (Thy-1- and B220-) cells. Their splenic NK activity to YAC targets was low and was not enhanced by IFN-alpha or IFN-beta. Bone marrow (BM) of these NK-1.1(-) mice have normal precursors to NK cells: 1) NK activity could be generated from NK-1.1(-) BM cells cultured in rIL 2 for 5 to 6 days. These cultured BM cells expressed Qa-5, Thy-1, AsGm-1, and NK-1.1 antigens. The precursor cells of these BM cytotoxic cells are NK-1.1-; 2) transfer of BM cells from the NK-1.1(-) mice reconstituted the NK activity of irradiated, NK-depleted recipients. Lymphokine-activated killer cells could also be generated from spleens of these NK-1.1(-) mice. Therefore, the NK-1.1(-) mice were specifically depleted of mature cytotoxic NK cells, but not the NK-1.1- precursors of NK cells. This mouse model is valuable to study ontogeny and physiologic relevance of NK cells.
Janus kinase 3
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NK-92
CD49b
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U1B3.3 monoclonal antibody (mAb) was established by immunizing a rat with the tMK-2U lymphoma cell line, derived from athymic nude mice. U1B3.3 mAb recognizes some T cell populations, B cells and natural killer (NK) cells. U1B3.3+ T cells express CD3/T cell receptor (TCR) complex at a unique intensity: between that of TCR-intermediate cells (TCRint cells) and TCRbright cells. TCRint cells strongly express the interleukin-2 receptor β chain (IL-2Rβ), but almost all U1B3.3+ T cells express high or low levels of IL-2Rβ. When purified U1B3.3+ NK1.1-T cells were cultured with IL-2, approximately 14% of the cells acquired expression of NK1.1 molecules, which are expressed on NK cells and natural killer T (NKT) cells. Furthermore, U1B3.3+ NK1.1-T cells killed tumor cells after culture with IL-2. These results indicate that U1B3.3 mAb recognizes a precursor of NK1.1+ cytotoxic T cells generated by IL-2.
Lymphokine-activated killer cell
Interleukin 3
Janus kinase 3
ZAP70
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