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The corticosteroid-induced inhibitory effect on NK cell function reflects down-regulation and/or dysfunction of triggering receptors involved in natural cytotoxicity
Abstract
Corticosteroids are known to inhibit NK cell functions. However no information is available on whether such inhibition may affect the expression and/or the function of receptors involved in NK cell activation. In an attempt to analyze this point, we studied peripheral blood NK cells isolated from pediatric patients undergoing allogeneic BM transplantation. NK cells were analyzed before, during and after methylprednisolone administration to treat acute graft-versus-host disease. In NK cells freshly isolated from peripheral blood during methylprednisolone treatment, the surface expression of activating receptors, particularly NKp46 and NKp30, was consistently reduced. Such impaired expression could also be detected after 5 days of culture in IL-2. Such cultured NK cells also failed to express the IL-2-inducible NKp44 receptor. Accordingly, cytotoxicity against different tumor target cell lines was sharply reduced. The effect on NK cells isolated from healthy individuals and cultured in the presence of methylprednisolone was also analyzed. A similar inhibitory effect occurred in the expression of activating NK receptors. In addition, a sharp impairment of NK cytotoxicity against different tumor target cell lines or immature DC was detected.
... Several studies have examined the effect of ISD on NK cells obtained from healthy donors in vitro (36)(37)(38)(39)(40)(41)(42)(43)(44)(45) and a few studies ex vivo, using blood samples from immunosuppressed patients (46)(47)(48)(49)(50)(51). However, the current evidence has never been consolidated and remains somewhat inconclusive, mostly due to different experimental designs ( Table 2). ...
... In contrast, the group of Mingari (36,40) reported that MePRD significantly down-regulates the expression of NKp30 and NKp44, perforin and CD69 in a dose-dependent and reversible manner; NKG2D, and 2B4 were partially inhibited, whereas NKp46, NKp80, NTBA, KIR and NKG2A remained unchanged. Ohira et al. showed a reduction of NKp44, TRAIL, and CD69 by several ISD (MePRD>calcineurin inhibitors and RAPA>MPA) whereas NKp30, NKG2D, and CD132 expression FIGURE 7 | Inhibition of NK cell IFNγ production by MePRD and calcineurin inhibitors but not by IVIg. ...
... In addition, IVIg was the strongest inhibitor of ADCC, even at doses used in replacement therapy to treat immunodeficiencies. These results are in line with previous reports showing that IVIg suppress NK cytotoxicity and ADCC (22,66), almost equally effective as prednisolone (40). Finally, early reports by the groups of Good and Herberman had also shown inhibition of NK cytotoxicity by IVIg without providing a mechanism (44,67); and more recently, Jacobi et al demonstrated suppression of K562 killing by NK cells, both in vitro and ex vivo, in neurological patients treated with high-dose intravenous IVIg, paralleled by the induction of degranulation and IFNγ release (54). ...
Small-molecule immunosuppressive drugs (ISD) prevent graft rejection mainly by inhibiting T lymphocytes. Therapeutic immunoglobulins (IVIg) are used for substitution, antibody-mediated rejection (AbMR) and HLA-sensitized recipients by targeting distinct cell types. Since the effect of ISD and IVIg on natural killer (NK) cells remains somewhat controversial in the current literature, the aim of this comparative study was to investigate healthy donor's human NK cell functions after exposure to ISD and IVIg, and to comprehensively review the current literature. NK cells were incubated overnight with IL2/IL12 and different doses and combinations of ISD and IVIg. Proliferation was evaluated by 3[H]-thymidine incorporation; phenotype, degranulation and interferon gamma (IFNγ) production by flow cytometry and ELISA; direct NK cytotoxicity by standard 51[Cr]-release and non-radioactive DELFIA assays using K562 as stimulator and target cells; porcine endothelial cells coated with human anti-pig antibodies were used as targets in antibody-dependent cellular cytotoxicity (ADCC) assays. We found that CD69, CD25, CD54, and NKG2D were downregulated by ISD. Proliferation was inhibited by methylprednisolone (MePRD), mycophenolic acid (MPA), and everolimus (EVE). MePRD and MPA reduced degranulation, MPA only of CD56bright NK cells. MePRD and IVIg inhibited direct cytotoxicity and ADCC. Combinations of ISD demonstrated cumulative inhibitory effects. IFNγ production was inhibited by MePRD and ISD combinations, but not by IVIg. In conclusion, IVIg, ISD and combinations thereof differentially inhibit NK cell functions. The most potent drug with an effect on all NK functions was MePRD. The fact that MePRD and IVIg significantly block NK cytotoxicity, especially ADCC, has major implications for AbMR as well as therapeutic strategies targeting cancer and immune cells with monoclonal antibodies.
... Glucocorticoids, like dexamethasone, bind glucocorticoid receptors irreversibly forming a complex, which interacts with cellular DNA in the nucleus and thereby changing gene transcription [11,12]. Steroids impinge on various stages of antigen presentation, cytokine production Transplant Immunology 37 (2016) [1][2][3][4][5][6][7][8][9] and proliferation, leading to a systemic suppression of inflammation and immune response [13,14]. ...
... Already we, amongst others, reported on the effects of ISD and MSCs on NK-cells and demonstrated that depending on the drug, both NK-cell proliferation and function can be impaired after 5-7 days of culture [13,[20][21][22]. IL-2 and IL-12 are critical cytokines involved in the maturation, activation, and survival of NK-cells [23]. ...
... Already we, amongst others, reported on the long term in vitro effects of ISD and MSCs on NK-cells and demonstrated that depending on the drug, both NK-cell proliferation and function is impaired after 5-7 days of culture [13,[20][21][22]. Further to our previous data, here we focused on early NK-cell activation markers, as in vivo contact with MSCs is likely limited due to rapid disappearance of these cells after infusion, and cytokine induced NK-cell responses are almost immediate [24,25]. ...
Graft rejection and graft-versus-host disease are leading causes of transplant related mortality despite advancements in immunosuppressive therapy. Mesenchymal stem cells (MSCs) offer a promising addition to immunosuppressive drugs (ISD), while NK-cells are increasingly used to combat graft-versus-leukemia. Combined therapy of ISD, NK-cells and/or MSCs is used in clinical practice. Here, we examined the effects of MSCs and selected ISD (tacrolimus, cyclosporin A, mycophenolic acid, dexamethasone) treatment on early NK-cell activation. We assessed STAT4 and STAT5 phosphorylation triggered by IL-12 and IL-2, respectively. Furthermore, we determined IFNγ, perforin production and the expression pattern of selected NK-cell receptors. Of all drugs tested, only dexamethasone inhibited NK-cell STAT4 and STAT5 phosphorylation. All ISD, with the exception of MPA, significantly inhibited IFNγ, only dexamethasone also inhibited upregulation of early activation markers CD69 and CD25 (IL-2 condition only). MSCs inhibited IL-2 induced NK cell STAT5 phosphorylation, IFNγ production and CD69 upregulation, and IL-12 induced IFNγ and perforin production. While MSCs mediated inhibition of CD69 expression was cell contact dependent, inhibition of IFNγ and perforin production, as well as STAT5 phosphorylation was cell-contact independent. Importantly, dexamethasone augmented MSCs mediated inhibition of both IL-12 and IL-2 induced CD69 expression and IFNγ production, as well as IL-2 induced STAT5 phosphorylation. Taken together, these novel insights may help the design of future NK-cell and MSCs based immunotherapy.
... Additionally, another study showed that imatinib induced the production of interferon gamma by natural killer cells, which correlated with its anti-tumor e cacy [14,15]. Considering that systemic steroids may inhibit T cell immunity [16] and suppress NK cell activity [17][18][19], systemic steroids might affect the immunomodulatory effects of imatinib, thereby decreasing its e cacy. ...
... These results suggest that systemic steroids may counteract the anti-tumor e cacy of ICIs. Given the potential immunomodulatory effects of imatinib [16][17][18][19], there are concerns regarding the potential adverse impact of systemic steroids on the immunomodulatory effects of imatinib by decreasing the activation of CD8 + T cells and NK cells. ...
Background: Effective management of adverse events is required to maintain sufficient imatinib dosing when treating patients with gastrointestinal stromal tumors (GISTs). Skin rash is a common adverse event of imatinib, which can be effectively controlled by systemic steroid treatment without imatinib dose modification or interruption. However, the impact of the use of systemic steroids on the efficacy of imatinib treatment remains unclear.
Methods: Between October 2014 and February 2022, 277 consecutive patients from a prospective registry of GIST patients were included as the study population. Patients who started systemic steroids due to grade ≥ 3 skin rash or grade 2 skin rash with grade 2 pruritis were classified as the steroid group, whereas patients who did not develop a skin rash or those who did not require steroids for a mild skin rash were classified as the control group. Efficacy outcomes were compared between the two groups.
Results: Among the 277 patients, 30 (10.8%) were treated with systemic steroids for skin rash. There was no significant difference in PFS or OS between the steroid and control groups (3-year PFS, 67.7% vs. 65.1%, p=0.53; 3-year OS, 91% vs. 89.9%, p=0.67, respectively). The use of systemic steroids was not an independent factor associated with PFS (HR 0.87, 95% CI 0.46–1.65, p=0.664) and OS (HR 0.53, 95% CI 0.19–1.46, p=0.218). In the steroid group, patients who successfully maintained the imatinib dosage showed a trend toward more favorable survival outcomes than those who did not (3-year PFS, 73.3% vs. 44.4%, p=0.34; 3-year OS, 95.8% vs. 75.0%, p=0.15, respectively).
Conclusions: The use of systemic steroids for the control of imatinib induced severe skin rash did not adversely affect the efficacy outcomes of imatinib in patients with advanced GIST.
... Furthermore, accumulating evidence indicated the effect of steroid hormones on NK cells [26][27][28][29][30][31][32][33][34][35]. Krukowski and her colleagues described that the release of glucocorticoids leads to the suppression of NK cell activity and the alteration of their cytokine production [26]. ...
... Glucocorticoids regulate NK cell function at least in part via epigenetic mechanisms, e.g., by decreasing the accessibility of promoters of interferon-gamma (IFNγ), perforins, and granzyme B [26][27][28]. In another study, the administration of exogenous glucocorticoids also decreased the surface expression of NK cell-activating receptors NKp30 and NKp46 [29]. Importantly, endogenous glucocorticoids upregulate checkpoint receptor PD-1 expression on NK cells during pathologic conditions and in disease progression [30,31]. ...
Steroid hormone production via the adrenal cortex, gonads, and placenta (so-called glandular steroidogenesis) is responsible for the endocrine control of the body’s homeostasis and is organized by a feedback regulatory mechanism based on the hypothalamus–pituitary–steroidogenic gland axis. On the other hand, recently discovered extraglandular steroidogenesis occurring locally in different tissues is instead linked to paracrine or autocrine signaling, and it is independent of the control by the hypothalamus and pituitary glands. Bone cells, such as bone-forming osteoblasts, osteoblast-derived osteocytes, and bone-resorbing osteoclasts, respond to steroid hormones produced by both glandular and extraglandular steroidogenesis. Recently, new techniques to identify steroid hormones, as well as synthetic steroids and steroidogenesis inhibitors, have been introduced, which greatly empowered steroid hormone research. Based on recent literature and new advances in the field, here we review the local role of steroid hormones in regulating bone homeostasis and skeletal lesion formation. The novel idea of extraglandular steroidogenesis occurring within the skeletal system raises the possibility of the development of new therapies for the treatment of bone diseases.
... Further, accumulating evidence indicated the effect of steroid hormones on NK cells [26][27][28][29][30][31][32][33][34][35]. Krukowski and her collegaues described, that release of glucocorticoids lead to suppression of NK cell activity and alteration of their cytokine production [26]. ...
... Glucocorticoids regulate NK cell function at least in part via epigenetic mechanisms, e.g. by decreasing the accessibility of promoters of interferon gamma (IFNγ), perforins and granzyme B [26][27][28]. In another study, administration of exogenous glucocorticoids also decreased the surface expression of NK cell activating receptors NKp30 and NKp46 [29]. Importantly, endogenous glucocorticoids upregulate checkpoint receptor PD-1 expression on NK cells during pathologic conditions and in disease progression [30][31]. ...
Steroid hormone production by adrenal cortex, gonads and placenta (so called glandular steroidogenesis) is responsible for the endocrine control of the body’s homeostasis and organized by a feedback regulatory mechanism based on the hypothalamus-pituitary-steroidogenic gland axis. On the other hand, recently discovered extraglandular steroidogenesis occurring locally in different tissues is rather linked to paracrine or autocrine signaling and it is independent of the control by the hypothalamus and pituitary. Bone cells, such as bone-forming osteoblast, osteoblast-derived osteocytes and bone-resorbing osteoclasts respond to steroid hormones produced by both glandular and extraglandular steroidogenesis. Recently, new techniques to identify steroid hormones as well as synthetic steroids and steroidogenesis inhibitors have been introduced, which greatly empowered steroid hormone research. Based on recent literature and new advances in the field, here we review the local role of steroid hormones in regulating bone homeostasis and skeletal lesion formation. The novel idea of extraglandular steroidogenesis occurring within the skeletal system raises the possibility of the development of new therapies for the treatment of bone diseases.
... The possible mechanisms of the interaction between high cortisol and low NKA are based on several direct effects of glucocorticoids on the immune system. Previous studies have reported the inhibitory effect of glucocorticoids on NK function [18][19][20][21][22][39][40][41][42]. After binding to the glucocorticoid receptor, glucocorticoids alter gene transcription, reducing the expression of genes important for NK function [21,41], like integrin LFA-1, which allows NK cells to adhere to target cells, or effector molecules such as perforin, granzyme A and B, which reduce the cytotoxicity of NK cells [20,22]. ...
... Previous studies have reported the inhibitory effect of glucocorticoids on NK function [18][19][20][21][22][39][40][41][42]. After binding to the glucocorticoid receptor, glucocorticoids alter gene transcription, reducing the expression of genes important for NK function [21,41], like integrin LFA-1, which allows NK cells to adhere to target cells, or effector molecules such as perforin, granzyme A and B, which reduce the cytotoxicity of NK cells [20,22]. The surface expression of the NK cytotoxicity receptors, such as NKp64 and NKp30, are down-regulated by glucocorticoids [43]. ...
The adrenal steroid hormones, cortisol and dehydroepiandrosterone sulfate (DHEAS), are associated with the immune system in opposite actions. This study aimed to investigate the relationship between cortisol and DHEAS serum concentrations, their ratio (CDR), and natural killer cell activity (NKA). This cross-sectional study included 2275 subjects without current infection or inflammation in the final analyses. NKA was estimated by measuring the amount of interferon-gamma (IFN-γ) released by activated natural killer cells; low NKA was defined as IFN-γ level < 500 pg/mL. Cortisol, DHEAS levels, and CDRs were categorized by quartiles in men, premenopausal women, and postmenopausal women. Compared with the lowest quartile as reference, the adjusted odd ratios (ORs) and 95% confidence intervals (CIs) for low NKA of the highest cortisol and CDR group were 1.66 (1.09–2.51) and 1.68 (1.11–2.55) in men, 1.58 (1.07–2.33) and 2.33 (1.58–3.46) in premenopausal women, and 2.23 (1.28–3.87) and 1.85 (1.07–3.21) in postmenopausal women. Only in premenopausal women, the highest DHEAS group showed significantly lower risk of low NKA (OR: 0.51, 95% CI: 0.35–0.76). HPA axis activation indicated as high cortisol level, CDR was significantly associated with low NKA, while high DHEAS levels were inversely associated with low NKA in premenopausal women.
... For concentration-response experiments, NK cells were exposed to four-to five-log increasing concentrations of dexamethasone (100 pM-1 mM) or LXA 4 (100 pM-100 nM). These concentrations were chosen based on prior published literature evaluating the effect of dexamethasone or other corticosteroids on human NK cells (14,(16)(17)(18)(19) and LXA 4 or other SPMs on human NK cells, monocytes, or PBMC (9,12,(20)(21)(22)(23)(24). ...
... Expression of NK cell-activating receptors was altered from baseline after 48 h of in vitro culture. Surface expression of NKp46 (Supplemental Fig. 2C) and NKG2D (Supplemental Fig. 2D) proteins were both lower in dexamethasone-exposed NK cells relative to vehicle control, consistent with prior reports for both human and murine NK cells exposed to corticosteroids in vitro (18,19,27). In contrast, NKp46 and NKG2D expression was preserved in LXA 4 -exposed NK cells with surface levels equivalent to vehicle-exposed NK cells (Supplemental Fig. 2C, 2D). ...
NK cells provide immune surveillance and host protection against viruses and tumors through their cytotoxic effector function. Cytoskeletal rearrangement is necessary for NK cell lytic granule trafficking and immune synapse formation to trigger apoptosis of targeted cells. LIM kinase (LIMK) regulates F-actin remodeling by phosphorylating cofilin to inhibit actin severing and depolymerization. In this study, in human NK cells, the glucocorticoid dexamethasone downregulated LIMK expression, F-actin accumulation at the immune synapse, lytic granule trafficking, and cytotoxicity. In contrast, the specialized proresolving mediator lipoxin A4 promoted NK cell LIMK expression, lytic granule polarization to the immune synapse and cytotoxicity. Using a LIMK inhibitor, we show that LIMK activity is necessary for NK cell cytotoxicity, including lipoxin A4's proresolving actions. Together, our findings identify LIMK as an important control mechanism for NK cell cytoskeletal rearrangement that is differentially regulated by glucocorticoids and specialized proresolving mediators to influence NK cell cytotoxicity.
... [33][34][35] This finding has been confirmed by many labs, and more details about the effect of GCs on NK cell reactivity have been reported. 34,[36][37][38][39][40][41] Through binding to GC receptors, GCs typically alter gene transcription. This change results in reduced expression of several genes that are important for NK cell functions. ...
... This change results in reduced expression of several genes that are important for NK cell functions. 37,39 Via reduced expression of the integrin LFA-1, adhesion to target cells is reduced by GCs. 38,41 In addition, effector molecules, such as perforin, granzyme B, and granzyme A, are inhibited in their expression, resulting in reduced cytotoxic activity of NK cells. ...
The immune system and the nervous system are highly complex organs composed of various different cells that must interact with each other for proper function of the system. This communication can be mediated by soluble factors. The factors released by the nervous system (neurotransmitters) differ from those released by the immune system (cytokines). Nevertheless, the nervous and immune systems can influence each other’s activity because immune cells express neurotransmitter receptors, and neurons express cytokine receptors. Moreover, immune cells can synthesize and release neurotransmitters themselves, thus using neurotransmitter-mediated pathways via autocrine and paracrine mechanisms. Natural killer (NK) cells are innate lymphocytes that are important for early and effective immune reactions against infections and cancer. Many studies have shown the strong influence of stress and the nervous system on NK cell activity. This phenomenon may be one reason why chronic stress leads to a higher incidence of infections and cancer. Here, we review the effects of neuroendocrine factors on the different activities of NK cells. Understanding the effects of neuroendocrine factors on NK cell activities during physiological and pathophysiological conditions may result in novel therapeutic strategies to enhance NK cell functions against tumors.
... This NCR has been shown to play important roles in crosstalk between NK cells and dendritic cells (DCs) through promoting both the maturation of and the cytotoxicity of immature DC (24,25). Surface expression levels of NKp30 and NKp46 can be upregulated by IFN-α, IL-2, and prolactin and downregulated by cortisol and methylprednisolone (26)(27)(28). In addition, both receptors are also commonly downregulated in "adaptive" or "memory-like" NK cells that are found in some cytomegalovirus-infected individuals (29,30). ...
... IL-2induced upregulation of NKp44 on NK cells can be inhibited by prostaglandin E2, which is readily produced by tumor-associated fibroblasts, especially when exposed to NK cells in culture (34). Similarly, prednisolone can suppress IL-2-mediated upregulation of NKp44 (26). ...
The natural cytotoxicity receptor (NCR) family is constituted by NKp46, NKp44, and NKp30 in humans, which are expressed mainly on natural killer (NK) cells and are encoded by the ncr1, ncr2, and ncr3 genes, respectively. NCRs have classically been defined as activating receptors that trigger cytotoxicity and cytokine responses by NK cells upon engaging with ligands on tumor cells. Several new findings, however, have challenged this model and identified alternative mechanisms regulating the function of NCRs. Recent reports indicate that ligand matters, since the interaction of NKp44 with distinct ligands on target cells can either activate or inhibit NK cells. Also, the NCRs have been found to interact with distinct specificities to various heparan sulfate glycosaminogly-cans, which are complex polysaccharides found in extracellular matrix or on cell surface heparan sulfate proteoglycans (HSPGs). The NCRs can engage with HSPGs in trans as a co-ligand on the target cells or in cis on the NK cell surface to regulate receptor–ligand interactions and NK cell activation. A number of splice variants of ncr2 and ncr3 have also been identified, and a predominant expression of certain variants results in inhib-itory signaling through NKp44 and NKp30. Several recent studies have found that the selective expression of some of these inhibitory splice variants can significantly influence outcome in the contexts of cancer, infection, and pregnancy. These findings establish that NCR functions are more diverse than originally thought, and better understanding of their splice variant expression profiles and ligand interactions are needed to establish their functional regulation in the context of human health.
... We know more about the effect of SAIDs, and especially DEX, on human NK cells. It has been demonstrated that these drugs reduce NKCA [10][11][12][13] , lower production of IFN-g by human NK cells 13,14 , and also show proapoptotic and anti-proliferative effect on these lymphocytes 15,16 and reduce their count 10,17 . The cited results indicate that SAIDs significantly affect the functioning of human NK cells, which certainly contribute to the production of immunosuppressive and anti-inflammatory properties of these drugs. ...
... What is surprising is the fact that although the available literature contains many results of studies on the effect of glucocorticosteroids on NKCA [10][11][12][13][29][30][31] , data on the effect of these compounds on counts of NK cells are very rare. Van Ierssel et al. 10 demonstrated that treatment with budesonide and prednisolone (SAIDs) significantly decreased the number of NK cells in patients with active ileocaecal Crohn's disease. ...
Abstract Due to the unrecognized effect of dexamethasone (DEX) and meloxicam (MEL) on bovine natural killer (NK) cells, studies have been undertaken in order to determine whether the above medications can affect these cells in respect of their counts, apoptosis, proliferation and production of selected cytokines. Peripheral blood mononuclear cells (PBMCs) were treated with the drugs in concentrations reflecting their plasma levels achieved in vivo at therapeutic doses and in 10-fold lower concentrations. The effect of DEX and MEL on percentages and absolute counts of NK cells was determined 6, 12, 48 and 168 h after the exposure of PBMCs to the drugs. At each time point, it was found out that DEX reduced the absolute count of NK cells, an effect attributed to the proapoptotic and anti-proliferative influence of the drug on these cells. DEX lowered the production of IFN-γ by the analyzed cells and raised the percentage of IL-10-producing cells. Thus, the above effects are important elements contributing to the complex mechanism responsible for the anti-inflammatory and immunosuppressive properties of the drug. MEL neither affects the apoptosis of NK cells nor did it reduce their count. Moreover, one-week exposure to MEL raised the absolute count of these cells, which was the result of their more intense proliferation in the presence of the drug. Thus, the influence of MEL with respect to the proliferation and count of NK cells was immunostimulating. On the other hand, MEL reduced the percentage of IFN-γ-producing NK cells, which in turn is an immunosuppressive effect.
... On the other hand, the levels of natural killer (NK) cells, which kill unwanted cells, are known to be negatively correlated with chronic pain conditions [104][105][106]. Of interest are the findings that NK cell numbers or cytotoxicity are increased by acute stress hormones or acute pain [107][108][109][110], whereas NK cell cytotoxicity is inhibited by chronic pain or chronic stress hormones through the hypothalamic-pituitaryadrenal axis [106,111,112]. There are reports that NK cells express LPAR 1 -3 [113,114], LPA decreases the cytotoxicity of NK cells [115], and secretes interferon-γ from NK cells [113]. ...
Fibromyalgia (FM) is a chronic pain condition characterized by widespread musculoskeletal pain and other frequent symptoms such as fatigue, sleep disturbance, cognitive impairment, and mood disorder. Based on the view that intermittent stress would be the most probable etiology for FM, intermittent cold- and intermittent psychological stress-induced generalized pain (ICGP and IPGP) models in mice have been developed and validated as FM-like pain models in terms of the patho-physiological and pharmacotherapeutic features that are shared with clinical versions. Both models show long-lasting and generalized pain and female-predominant sex differences after gonadectomy. Like many other neuropathic pain models, ICGP and IPGP were abolished in lysophosphatidic acid receptor 1 (LPAR1) knock-out mice or by LPAR1 antagonist treatments, although deciding the clinical importance of this mechanism depends on waiting for the development of a clinically available LPAR1 antagonist. On the other hand, the nonsteroidal anti-inflammatory drug diclofenac with morphine did not suppress hyperalgesia in these models, and this is consistent with the clinical findings. Pharmacological studies suggest that the lack of morphine analgesia is associated with opioid tolerance upon the stress-induced release of endorphins and subsequent counterbalance through anti-opioid NMDA receptor mechanisms. Regarding pharmacotherapy, hyperalgesia in both models was suppressed by pregabalin and duloxetine, which have been approved for FM treatment in clinic. Notably, repeated treatments with mirtazapine, an α2 adrenergic receptor antagonist-type antidepressant, and donepezil, a drug for treating Alzheimer’s disease, showed potent therapeutic actions in these models. However, the pharmacotherapeutic treatment should be carried out 3 months after stress, which is stated in the FM guideline, and many preclinical studies, such as those analyzing molecular and cellular mechanisms, as well as additional evidence using different animal models, are required. Thus, the ICGP and IPGP models have the potential to help discover and characterize new therapeutic medicines that might be used for the radical treatment of FM, although there are several limitations to be overcome.
... NK cells comprise a crucial barrier of the innate immune system by their early production of cytokines and chemokines and cell lysis potential irrespective of prior sensitization (44). Previous publications have demonstrated that GCs influence NK cell function through epigenetic modifications resulting in dysregulation of effector protein expression and hence inhibition of NK cell cytotoxicity, at least partially explaining the effect of chronic stress on incidence of infections and cancer (44)(45)(46). GCs however can alsodependent on the cytokine profile presentsupport NK cell proliferation and reactivity (35). Our data supports this hypothesis. ...
Introduction
Patients with primary adrenal insufficiency (PAI) suffer from increased risk of infection, adrenal crises and have a higher mortality rate. Such dismal outcomes have been inferred to immune cell dysregulation because of unphysiological cortisol replacement. As the immune landscape of patients with different types of PAI has not been systematically explored, we set out to immunophenotype PAI patients with different causes of glucocorticoid (GC) deficiency.
Methods
This cross-sectional single center study includes 28 patients with congenital adrenal hyperplasia (CAH), 27 after bilateral adrenalectomy due to Cushing’s syndrome (BADx), 21 with Addison’s disease (AD) and 52 healthy controls. All patients with PAI were on a stable GC replacement regimen with a median dose of 25 mg hydrocortisone per day. Peripheral blood mononuclear cells were isolated from heparinized blood samples. Immune cell subsets were analyzed using multicolor flow cytometry after four-hour stimulation with phorbol myristate acetate and ionomycin. Natural killer (NK-) cell cytotoxicity and clock gene expression were investigated.
Results
The percentage of T helper cell subsets was downregulated in AD patients (Th1 p = 0.0024, Th2 p = 0.0157, Th17 p < 0.0001) compared to controls. Cytotoxic T cell subsets were reduced in AD (Tc1 p = 0.0075, Tc2 p = 0.0154) and CAH patients (Tc1 p = 0.0055, Tc2 p = 0.0012) compared to controls. NKCC was reduced in all subsets of PAI patients, with smallest changes in CAH. Degranulation marker CD107a expression was upregulated in BADx and AD, not in CAH patients compared to controls (BADx p < 0.0001; AD p = 0.0002). In contrast to NK cell activating receptors, NK cell inhibiting receptor CD94 was upregulated in BADx and AD, but not in CAH patients (p < 0.0001). Although modulation in clock gene expression could be confirmed in our patient subgroups, major interindividual-intergroup dissimilarities were not detected.
Discussion
In patients with different etiologies of PAI, distinct differences in T and NK cell-phenotypes became apparent despite the use of same GC preparation and dose. Our results highlight unsuspected differences in immune cell composition and function in PAI patients of different causes and suggest disease-specific alterations that might necessitate disease-specific treatment.
... Relevant for our study, corticosteroid treatment of pediatric patients was shown to reduce surface expression of NKG2D, NKp30, and NKp46 and lower the activity of NK cells. 47,48 It is thus difficult to conclude whether the phenotypic changes observed at day 79 are due to intrinsic defects in the NK cells or to an effect of the chemotherapeutic regimen. By the end of treatment, we observed a higher abundance of NKG2A + CD57 − KIR − immature NK cells (data not shown), indicating that newly developed NK cells may dominate the circulating pool of NK cells at this point. ...
Natural killer (NK) cells are important for early tumor immune surveillance. In patients with hematological cancers, NK cells are generally functional deficient and display dysregulations in their receptor repertoires. Acute leukemia is the most common cancer in children, and we here performed a comparative phenotypic profiling of NK cells from B-cell precursor acute lymphoblastic leukemia (BCP-ALL) patients to identify aberrant NK cell phenotypes. NK cell phenotypes, maturation, and function were analyzed in matched bone marrow and blood NK cells from BCP-ALL patients at diagnosis, during treatment, and at end of treatment and compared with age-matched pediatric control subjects. Expression of several markers were skewed in patients, but with large interindividual variations. Undertaking a multiparameter approach, we found that high expression levels of NKG2A was the single predominant marker distinguishing NK cells in BCP-ALL patients compared with healthy control subjects. Moreover, naïve CD57−NKG2A NK cells dominated in BCP-ALL patients at diagnosis. Further, we found dysregulated expression of the activating receptor DNAM-1 in resident bone marrow CXCR6+ NK cells. CXCR6+ NK cells lacking DNAM-1 expressed NKG2A and had a tendency for lower degranulation activity. In conclusion, high expression of NKG2A dominates NK cell phenotypes from pediatric BCP-ALL patients, indicating that NKG2A could be targeted in therapies for this patient group.
... Methylprednisolone on purified NK cells significantly inhibited cytotoxicity, proliferation, IFN-γ production and the expression of NK cell activation markers, showing a direct effect of corticosteroids [130]. Finally, methylprednisolone administration in vivo decreases the expression of natural cytotoxicity receptors (NKp30, NKp44 and NKp46) on peripheral blood NK cells [131]. ...
Asthma is a chronic inflammatory airway disorder whose pathophysiological and immunological mechanisms are not completely understood. Asthma exacerbations are mostly driven by respiratory viral infections and characterised by worsening of symptoms. Despite current therapies, asthma exacerbations can still be life-threatening. Natural killer (NK) cells are innate lymphoid cells well known for their antiviral activity and are present in the lung as circulating and resident cells. However, their functions in asthma and its exacerbations are still unclear. In this review, we will address NK cell activation and functions, which are particularly relevant for asthma and virus-induced asthma exacerbations. Then, the role of NK cells in the lungs at homeostasis in healthy individuals will be described, as well as their functions during pulmonary viral infections, with an emphasis on those associated with asthma exacerbations. Finally, we will discuss the involvement of NK cells in asthma and virus-induced exacerbations and examine the effect of asthma treatments on NK cells.
... The biological reason for the poor prognosis in patients with high-dose mPSL use may be the attenuation of NK-cells. 34 Moreover, non-CAR lymphocytes expand more in the tumor microenvironment (TME) than CAR T-cells 5 days after CAR T-cell infusion. 35 Corticosteroid therapy may suppress non-CAR lymphocytes in infiltrating TME. ...
Background:
The prognostic impact of corticosteroid therapy for patients with tisagenlecleucel (tisa-cel) treatment who are more likely to develop cytokine release syndrome (CRS) still remains unclear.
Objective:
This study aimed to evaluate the clinical impact and lymphocyte kinetics of corticosteroid administration for CRS in 45 patients with relapsed and/or refractory B-cell lymphoma treated with tisa-cel.
Study design:
This was a retrospective evaluation of all consecutive patients diagnosed with relapsed and/or refractory diffuse large B-cell lymphoma, follicular lymphoma with histologic transformation to large B-cell lymphoma, or follicular lymphoma who received commercial-based tisa-cel treatment.
Results:
The best overall response rate, complete response rate, median progression-free survival (PFS), and median overall survival (OS) were 72.7%, 45.5%, 6.6 months, and 15.3 months, respectively. CRS (predominantly grade 1/2) occurred in 40 patients (88.9%), while ICANS (all grade) occurred in 3 patients (6.7%). No grade 3 or higher ICANS occurred. Patients with high-dose (≥ 524 mg, methylprednisolone equivalent, n = 12) or long-term (≥ 8 days, n = 9) corticosteroid use had inferior PFS and OS to patients with low or no corticosteroid use (both p < 0.05). The prognostic impact remained even in 23 patients with stable (SD) or progressive disease (PD) before tisa-cel infusion (p = 0.015) but not in patients with better disease status (p = 0.71). The timing of corticosteroid initiation did not have a prognostic impact. Multivariate analysis revealed that high-dose or long-term corticosteroid use were the independent prognostic factors of PFS and OS, respectively, after adjusting for elevated LDH before lymphodepletion chemotherapy and disease status (SD or PD). Lymphocyte kinetics analysis demonstrated that after methylprednisolone administration, the proportion of regulatory T-cells (Tregs), CD4+ central memory T (TCM) cells, and NK-cells were decreased, while that of CD4+ effector memory T (TEM) cells was increased. Patients with the higher proportion of Tregs at day 7 had a lower incidence of CRS, but this did not affect prognosis, indicating that early elevation of Tregs may serve as a biomarker for CRS development. Furthermore, patients with the higher number of CD4+ TCM cells and NK-cells at various time points had significantly better PFS and OS, whereas the number of CD4+ TEM cells did not impact prognostic outcomes.
Conclusion:
This study suggested that high-dose or long-term corticosteroid use attenuates the efficacy of tisa-cel, especially in patients with SD or PD. Additionally, patients with the high level of CD4+ TCM cells and NK-cells after tisa-cel infusion have longer PFS and OS.
... Dexamethasone (Dex) is a strong synthetic member of the glucocorticoid class of steroid drugs that act as an anti-in ammatory and immunosuppressant molecule [4]. To achieve systemic inhibition of in ammatory and immune response, the steroids in uence various levels of antigen presentation, immune cell proliferation, and cytokine production [5]. Therefore, dexamethasone impacts cellular DNA, thereby changing gene transcription [6]. ...
Background: Human Mesenchymal Stem Cells (hMSCs) represent a promising cell source for cell-based therapy in autoimmune diseases and other degenerative disorders due to their immunosuppressive, anti-inflammatory and regenerative potentials. Belonging to a glucocorticoid family, Dexamethasone (Dex) is a powerful anti-inflammatory compound that is widely used as therapy in autoimmune disease conditions or allogeneic transplantation. However, minimal immunomodulatory effect of hMSCs may limit their therapeutic uses. Moreover, the effect of glucocorticoids on the immunomodulatory molecules or other regenerative properties of tissue-specific hMSCs remains unknown.
Method: Herein, we evaluated the in vitro effect of Dex at various dose concentrations and time intervals, 1000 ng/ml, 2000 ng/ml, 3000 ng/ml and 24 h, 48 h respectively, on the basic characteristics and immunomodulatory properties of Bone marrow derived MSC (BM-MSCs), Adipose tissue derived MSCs (AD-MSCs), Dental Pulp derived MSC (DP-MSCs) and Umbilical cord derived MSCs (UC-MSCs).
Results: The present study indicated that the concentration of Dex did not ramify the cellular morphology nor showed cytotoxicity as well as conserved the basic characteristics of tissue specific hMSCs including cell proliferation and surface marker profiling. However, quite interestingly it was observed that the stemness markers (Oct-4, Sox-2, Nanog & Klf-4) showed a significant upregulation in DP-MSCs and AD-MSCs followed by UC-MSCs and BM-MSCs. Additionally, immunomodulatory molecules, Prostaglandin E-2 (PGE-2), Indoleamine- 2,3-dioxygenase (IDO) and Human Leukocyte Antigen-G (HLA-G) were seen to be upregulated in a dose-dependent manner. Moreover, there was a differential response of tissue specific hMSCs after pre-conditioning with Dex during mixed lymphocyte reaction, wherein UC-MSCs and DP-MSCs showed enhanced immunosuppression as compared to AD-MSCs and BM-MSCs, thereby proving to be a better candidate for therapeutic applications in immune-related diseases.
Conclusion: Dex preconditioning ameliorates the hMSCs immunomodulatory property and may void the challenge associated with minimal potency and strengthen their therapeutic efficacy.
... An immunophenotyping study by Chitadze et al. found that CD56 bright NK cells were significantly downregulated in glioblastoma patients treated with steroids, whereas steroid had no apparent impact on CD56 dim NK cells [45]. Vitale further indicated that the surface density of various activating NK receptors declined during methylprednisolone treatment, which is recognized as NK cell dysfunction, and then returned to normal levels shortly after steroid discontinuation or low-dose use [46]. Therefore, in this study, the decline of NKA and CD56 bright NK cells on POD3 after surgical removal glioblastoma may not be or slightly due to the steroid use. ...
Glioblastoma is the most common primary malignant brain tumor, and median survival is relatively short despite aggressive standard treatment. Natural killer (NK) cell dysfunction is strongly associated with tumor recurrence and metastasis but is unclear in glioblastoma. NK activity (NKA) represents NK cell-secreted interferon-γ (IFN-γ), which modulates immunity and inhibits cancer progression. This study aimed to analyze NKA in glioblastoma patients to obtain a clearer overview of immunity surveillance. From 2020 to 2021, a total of 20 patients and six healthy controls were recruited. Peripheral blood samples were collected preoperatively and on postoperative days (POD) 3 and 30. Then, NKA was measured using the NK VUE kit. Although NKA decreased on POD3, it recovered and further significantly enhanced on POD30, with a nearly five-fold increase compared to baseline (p = 0.004). Furthermore, the percentage of CD56brightCD16− NK cells decreased significantly on POD3 (p = 0.022) and further recovered on PO30. Subgroup analysis of extent surgical resection further revealed that the recovery of impaired NKA was attributable to gross total resection (GTR) rather than subtotal resection (STR). In conclusion, NKA is significantly impaired in glioblastoma, and GTR has demonstrated superior benefit in improving the suppressed NKA and increased CD56brightCD16− NK subset in glioblastoma patients, which may be associated with subsequent patients’ prognosis. Therefore, the goal of performing GTR for glioblastoma should be achieved when possible since it appears to increase NKA cell immunity.
... Frontiers in Oral Health | www.frontiersin.org [163,188,[205][206][207][208][209][210] and dendritic cell maturation, (3) lymphopenia, (4) reduction of CD4 T cell function, (5) an increase in regulatory T and B cells able to dampen immune responses, and (6) a suppressed NK phenotype through a decreased number of activating receptors and cytotoxic potential as well as increased number of inhibitory receptor numbers [244]. Interestingly, surgical resection of the tumor can reverse various of these peripheral immune perturbations in multiple murine breast and colon cancer models, suggesting the observed immune effects to indeed be induced by the tumor itself [245]. ...
Candida albicans and Staphylococcus aureus account for most invasive fungal and bacterial bloodstream infections (BSIs), respectively. However, the initial point of invasion responsible for S. aureus BSIs is often unclear. Recently, C. albicans has been proposed to mediate S. aureus invasion of immunocompromised hosts during co-colonization of oral mucosal surfaces. The status of the oral immune system crucially contributes to this process in two distinct ways: firstly, by allowing invasive C. albicans growth during dysfunction of extra-epithelial immunity, and secondly following invasion by some remaining function of intra-epithelial immunity. Immunocompromised individuals at risk of developing invasive oral C. albicans infections could, therefore, also be at risk of contracting concordant S. aureus BSIs. Considering the crucial contribution of both oral immune function and dysfunction, the aim of this review is to provide an overview of relevant aspects of intra and extra-epithelial oral immunity and discuss predominant immune deficiencies expected to facilitate C. albicans induced S. aureus BSIs.
... Furthermore, GCs have also long been known to inhibit NK cell activity. GCs were shown to alter the expression of surface receptors critical for NK cell functions, including CD16, DNAM-1, NKp46 and NKp30 [125,126]. Treatment of isolated NK cells with the synthetic GC dexamethosone was shown to inhibit mTORC1 leading to reduced IFNγ production following IL-2/IL-12 sitmulation [126]. In addition, GCs have been shown to impair NK cell cytoxiticty as evidenced by reduced perforin and granzyme A/B release [124,127,128]. ...
Surgical resection is the foundation for the curative treatment of solid tumors. However, metastatic recurrence due to the difficulty in eradicating micrometastases remain a feared outcome. Paradoxically, despite the beneficial effects of surgical removal of the primary tumor, the physiological stress resulting from surgical trauma serves to promote cancer recurrence and metastasis. The postoperative environment suppresses critical anti-tumor immune effector cells, including Natural Killer (NK) cells. The literature suggests that NK cells are critical mediators in the formation of metastases immediately following surgery. The following review will highlight the mechanisms that promote the formation of micrometastases by directly or indirectly inducing NK cell suppression following surgery. These include tissue hypoxia, neuroendocrine activation, hypercoagulation, the pro-inflammatory phase, and the anti-inflammatory phase. Perioperative therapeutic strategies designed to prevent or reverse NK cell dysfunction will also be examined for their potential to improve cancer outcomes by preventing surgery-induced metastases.
... Dexamethasone (Dex) is a strong synthetic member of the glucocorticoid class of steroid drugs that act as an antiinflammatory and immunosuppressant molecule [4]. To achieve systemic inhibition of inflammatory and immune response, the steroids influence various levels of antigen presentation, immune cell proliferation, and cytokine production [5]. Therefore, dexamethasone impacts cellular DNA, thereby changing gene transcription [6]. ...
Background
Human Mesenchymal Stem Cells (hMSCs) represent a promising cell source for cell-based therapy in autoimmune diseases and other degenerative disorders due to their immunosuppressive, anti-inflammatory and regenerative potentials. Belonging to a glucocorticoid family, Dexamethasone (Dex) is a powerful anti-inflammatory compound that is widely used as therapy in autoimmune disease conditions or allogeneic transplantation. However, minimal immunomodulatory effect of hMSCs may limit their therapeutic uses. Moreover, the effect of glucocorticoids on the immunomodulatory molecules or other regenerative properties of tissue-specific hMSCs remains unknown.
Method
Herein, we evaluated the in vitro effect of Dex at various dose concentrations and time intervals, 1000 ng/ml, 2000 ng/ml, 3000 ng/ml and 24 h, 48 h respectively, on the basic characteristics and immunomodulatory properties of Bone marrow derived MSC (BM-MSCs), Adipose tissue derived MSCs (AD-MSCs), Dental Pulp derived MSC (DP-MSCs) and Umbilical cord derived MSCs (UC-MSCs).
Results
The present study indicated that the concentration of Dex did not ramify the cellular morphology nor showed cytotoxicity as well as conserved the basic characteristics of tissue specific hMSCs including cell proliferation and surface marker profiling. However, quite interestingly it was observed that the stemness markers (Oct-4, Sox-2, Nanog and Klf-4) showed a significant upregulation in DP-MSCs and AD-MSCs followed by UC-MSCs and BM-MSCs. Additionally, immunomodulatory molecules, Prostaglandin E-2 (PGE-2), Indoleamine- 2,3-dioxygenase (IDO) and Human Leukocyte Antigen-G (HLA-G) were seen to be upregulated in a dose-dependent manner. Moreover, there was a differential response of tissue specific hMSCs after pre-conditioning with Dex during mixed lymphocyte reaction, wherein UC-MSCs and DP-MSCs showed enhanced immunosuppression as compared to AD-MSCs and BM-MSCs, thereby proving to be a better candidate for therapeutic applications in immune-related diseases.
Conclusion
Dex preconditioning improved the hMSCs immunomodulatory property and may have reduced the challenge associated with minimal potency and strengthen their therapeutic efficacy.
Graphical Abstract
Preconditioning of tissue specific hMSCs with dexamethasone biomanufacturers the enhanced potential hMSCs with better stemness and immunomodulatory properties for future therapeutics.
... Synthetic corticosteroids methylprednisolone treatment causes reduction of natural cytotoxicity receptors including NKp46, NKp30 and inhibits the expression of IL-2inducible NKp44 receptor. Methylprednisolone-treated normal NK cells exhibit a reduced level of intra-cytoplasmic perforin, granzyme A and B [47]. Estrogen inhibits NK cell cytotoxicity, possibly through estrogen receptor (ER)-β [48]. ...
Historically tools and technologies facilitated scientific discoveries. Steroid hormone research is not an exception. Unfortunately, the dramatic advancement of the field faded this research area and flagged it as a solved topic. However, it should have been the opposite. The area should glitter with its strong foundation and attract next-generation scientists. Over the past century, a myriad of new facts on biochemistry, molecular biology, cell biology, physiology and pathology of the steroid hormones was discovered. Several innovations were made and translated into life-saving treatment strategies such as synthetic steroids, and inhibitors of steroidogenesis and steroid signaling. Steroid molecules exhibit their diverse effects on cell metabolism, salt and water balance, development and function of the reproductive system, pregnancy, and immune-cell function. Despite vigorous research, the molecular basis of the immunomodulatory effect of steroids is still mysterious. The recent excitement on local extra-glandular steroidogenesis in regulating inflammation and immunity is revitalizing the topic with a new perspective. Therefore, here we review the role of steroidogenesis in regulating inflammation and immunity, discuss the unresolved questions, and how this area can bring another golden age of steroid hormone research with the development of new tools and technologies and advancement of the scientific methods.
... Barkin J et al. [21] found that this method was used to detect NK cell activity, and the results showed that subjects with low NK cell activity values were more likely to have positive results in prostate cancer biopsy. Tae BS et al. [34] also proved that the NK cell activity [38], proton pump inhibitors [39], long-term smoking [40], drinking [41] and foods rich in zinc and selenium [42] [43], etc., will temporarily decrease or increase the value of NK cell activity high. Therefore, the clinical application of this method needs to be further verified, and the influence of more exclusionary factors needs to be considered. ...
... NK cells are the first lymphoid lineage cell type reconstituted after allo-HSCT and remain the dominant circulating lymphocyte population during the first three months post-transplant [3]. However, CMV infection, corticosteroids, or the presence of GvHD can affect the reconstitution of NK cells [4,5]. ...
Killer-immunoglobulin-like receptors (KIRs) are critical natural killer (NK) cell regulators. The expression of KIRs is a dynamic process influenced by many factors. Their ligands—HLA(Human Leukocyte Antigen) class I molecules—are expressed on all nucleated cells that keep NK cells under control. In hematopoietic stem cell transplantation (HSCT), NK cells play an essential role in relapse protection. In the presented pilot study, we characterized the dynamic expression of inhibitory KIRS (iKIRs), which protect cells against untoward lysis, in donors and patients during the first three months after HSCT using flow cytometry. The expression of all iKIRs was highly variable and sometimes correlated with patients’ clinical presentation and therapy regiment. Cyclophosphamide (Cy) in the graft-versus-host disease (GvHD) prevention protocol downregulated KIR2DL1 to just 25% of the original donor value, and the FEAM (Fludarabine + Etoposid + Ara-C + Melphalan) conditioning protocol reduced KIR2DL3. In lymphoid neoplasms, there was a slightly increased KIR2DL3 expression compared to myeloid malignancies. Additionally, we showed that the ex vivo activation of NK cells did not alter the level of iKIRs. Our study shows the influence of pre- and post-transplantation protocols on iKIR expression on the surface of NK cells and the importance of monitoring their cell surface.
... In a study, 400 mg/kg IVIG was given for at least 6 months with four weeks interval to ten immunodeficient children with ASD; improvements in social interactions, behavioral issues, echolalia, speech, eye contact, and reduced hyperactivity were seen in them [103]. Corticosteroids reduce NK cells activity and cytotoxicity by reducing the expression of activating receptors [106]. Effectiveness of corticosteroid therapy in children with regressive ASD has been investigated; a group of 20 children with ASD treated with 2 mg/kg/day oral prednisolone was compared with another group of 24 children with ASD not treated with steroids. ...
ntroduction: Autism spectrum disorder (ASD), as a wide spectrum of neurodevelopmental disorders, is characterized by early-onset impairments in social-communication, repetitive behaviors, and restrictive interests.
Areas covered: Although still unknown, there are some pieces of evidence suggesting altered immune function in the etiology of ASD. This review aims to summarize studies linking Natural Killer (NK) cells to ASD by searching through databases like MEDLINE and Scopus up to October 2020. NK cells play important roles in the innate immune system and immune regulation. As parts of the immune system, they interact with the neural system as well. Immune dysregulations such as autoimmunity and improper immune responses to both internal and external stimulations, especially in early developmental stages of the brain, may induce neurodevelopmental disorders. NK cells’ dysfunction in children with ASD as well as their parents have been highlighted in many studies.
Expert opinion: Changes in the frequency, gene expressions, cytotoxicity features, and receptors of NK cells are reported in children with ASD. Immune therapy for children with ASD with immune abnormality has shown promising results. However, further studies are needed to elucidate the exact role of NK cells in the pathogenesis of ASD providing future treatment options for these children.
... Inhaled corticosteroids (ICS) are potent immunomodulators and high doses of ICS are routinely administered in response to COPD exacerbations. Indeed, high doses of ICS use has been shown, both in vivo and in vitro, to be associated with alterations to NK surface phenotype and function 10,11 . Therefore, patients with an exacerbation and possible ICS use in the month prior to enrollment were excluded. ...
Chronic Obstructive Pulmonary Disease (COPD) is the third leading cause of death worldwide. COPD is frequently punctuated by acute exacerbations that are precipitated primarily by infections, which increase both morbidity and mortality and inflates healthcare costs. Despite the significance of exacerbations, little understanding of immune function in COPD exacerbations exists. Natural killer (NK) cells are important effectors of innate and adaptive immune responses to pathogens and NK cell function is altered in smokers and COPD. Using high-dimensional flow cytometry, we phenotyped peripheral blood NK cells from never smokers, smokers, and COPD patients and employed a non-supervised clustering algorithm to define and detect changes in NK cell populations. We identified greater than 1,000 unique NK cell subpopulations across patient groups and describe 13 altered NK populations in patients who experienced prior exacerbations. Based upon cluster sizes and associated fluorescence data, we generated a logistic regression model to predict patients with a history of exacerbations with high sensitivity and specificity. Moreover, highly enriched NK cell subpopulations implicated in the regression model exhibited enhanced effector functions as defined by in vitro cytotoxicity assays. These novel data reflect the effects of smoking and disease on peripheral blood NK cell phenotypes, provide insight into the potential immune pathophysiology of COPD exacerbations, and indicate that NK cell phenotyping may be a useful and biologically relevant marker to predict COPD exacerbations.
... The surface expression of NKp46 can be perturbed by different factors released in tumor microenvironment, including the l-kynurenine, which is a tryptophan catabolism product generated by the indoleamine 2,3-dioxygenase (IDO) enzyme [77], corticosteroid [78] and by chronic interaction with tumor cells (Fig. 1) [79]. Together with NKp30, NKp46 is also down-regulated on "adaptive" or "memory-like" NK cells during the course of Cytomegalovirus (HCMV) infection/reactivation and even more on the PD-1 pos NK cell subset found in HCMV pos healthy donors [11,23,80,81]. ...
Natural Killer (NK) cells are innate lymphocytes able to mediate immune-surveillance and clearance of viral infected and tumor-transformed cells. Growing experimental and clinical evidence highlighted a dual role of NK cells either in the control of cancer development/progression or in promoting the onset of immune-suppressant tumor microenvironments. Indeed, several mechanisms of NK cell-mediated tumor escape have been described and these includes cancer-induced aberrant expression of activating and inhibitory receptors (i.e. NK cell immune checkpoints), impairments of NK cell migration to tumor sites and altered NK cell effector-functions. These phenomena highly contribute to tumor progression and metastasis formation.
In this review, we discuss the latest insights on those NK cell receptors and related molecules that are currently being implemented in clinics either as possible prognostic factors or therapeutic targets to unleash NK cell anti-tumor effector-functions in vivo. Moreover, we address here the major recent advances in regard to the genetic modification and ex vivo expansion of anti-tumor specific NK cells used in innovative adoptive cellular transfer approaches.
... Moreover, the source of CD34 + precursors used for hematopoietic stem cell transplantation (HSCT) influences the prevalent type of progeny (mainly NK cells vs ILC3) [28]. In addition, drugs, including steroids [29,30] or tyrosin kynase inhibitors (Damele, Frontiers in Immunology 2018, in press) may also cause a differential recovery of NK cells and/or ILC3 in vitro, suggesting the possibility of preferential growth of one or another subset in vivo. A remarkable characteristic of ILC is their plasticity, i.e. their ability to convert into one another as a consequence of signals delivered by the tissue microenvironment, cytokines or pathogens [16]. ...
Innate lymphoid cells (ILC) including NK cells (cytotoxic) and the recently identified “helper” ILC1, ILC2 and ILC3, play an important role in innate defenses against pathogens. Notably, they mirror analogous T cell subsets, regarding the pattern of cytokine produced, while the timing of their intervention is few hours vs days required for T cell-mediated adaptive responses. On the other hand, the effectiveness of ILC in anti-tumor defenses is controversial. The relevance of NK cells in the control of tumor growth and metastasis has been well documented and they have been exploited in the therapy of high risk leukemia in the haploidentical hematopoietic stem cell transplantation setting. In contrast, the actual involvement of helper ILCs remains contradictory. Thus, while certain functional capabilities of ILC1 and ILC3 may favor anti-tumor responses, other functions could rather favor tumor growth, neo-angiogenesis, epithelial-mesenchymal transition and metastasis. In addition, ILC2, by secreting type-2 cytokines, are thought to induce a prevalent pro-tumorigenic effect. Finally, the function of both NK cells and helper ILCs may be inhibited by the tumor microenvironment, thus adding further complexity to the interplay between ILC and tumors.
... The mechanisms of T and B cell inhibition mediated by conventional immunosuppressive drugs (ISD) and biologicals used in transplantation medicine are well known. As to NK cells, the literature reports a variety of specific effects of ISD on NK cells with to some extent conflicting conclusions [152][153][154][155][156][157][158][159][160]. A comprehensive comparative study confirmed that corticosteroids are potent inhibitors of NK functions including ADCC, direct NK cytotoxicity, and IFNγ production (own unpublished data). ...
Recruitment of human NK cells to porcine tissues has been demonstrated in pig organs perfused ex vivo with human blood in the early 1990s. Subsequently, the molecular mechanisms leading to adhesion and cytotoxicity in human NK cell-porcine endothelial cell (pEC) interactions have been elucidated in vitro to identify targets for therapeutic interventions. Specific molecular strategies to overcome human anti-pig NK cell responses include (1) blocking of the molecular events leading to recruitment (chemotaxis, adhesion, and transmigration), (2) expression of human MHC class I molecules on pECs that inhibit NK cells, and (3) elimination or blocking of pig ligands for activating human NK receptors. The potential of cell-based strategies including tolerogenic dendritic cells (DC) and regulatory T cells (Treg) and the latest progress using transgenic pigs genetically modified to reduce xenogeneic NK cell responses are discussed. Finally, we present the status of phenotypic and functional characterization of nonhuman primate (NHP) NK cells, essential for studying their role in xenograft rejection using preclinical pig-to-NHP models, and summarize key advances and important perspectives for future research.
... With regard to the triggering receptors, we observed no effect on the expression of NKG2A, but a moderate decrease in the expression of both NKG2D and NKp46 by treatment with 10 lg of Dex. The observed inhibitory effect of Dex on NKp46 is in line with a previous report, demonstrating reduction in NKp46 by methylprednisolone (Vitale et al. 2004). ...
Glucocorticoids (GCs) act via the intracellular glucocorticoid receptor (GR), which can regulate the expression of target genes. With regard to the immune system, GCs may affect both innate and adaptive immunity. Our study analyzed the immunoregulatory effects of dexamethasone (Dex) treatment on splenic T, Treg, NK and NKT cells by treating C57Bl6 mice with various doses of Dex. We observed that treatment with Dex decreased the number of NK cells in the spleen and suppressed their activity. In particular, the expression of both Ly49G and NKG2D receptors was decreased by Dex. However, Dex did not affect the population of NKT cells. With regard to splenic T cells, our results show a dose-dependent reduction in CD3⁺, CD4⁺, CD8⁺, CD44⁺ and CD8⁺CD122⁺ T cells, but a stimulatory effect on CD4⁺CD25⁺ regulatory T cells by Dex treatment. In addition, treatment with Dex suppressed anti-tumor immune response in a mouse EG7 tumor model. We conclude that Dex may suppress both T- and NK-mediated immunity.
... Corticosteroids, the most effective drugs in the treatment of many acute and chronic inflammatory diseases, are able to control many cellular functions involved in the pathogenesis of the damage induced by viral infections ( Figure 5). 19,[47][48][49][50][51][52] Despite the poverty of clinical studies on this subject, the above considerations suggest that a short-term treatment with inhaled steroids can represent an effective therapeutic option in the treatment of persistent post-infection cough, especially if associated with bronchoconstriction. 38,53 In support of this statement is: 1) the indication to use inhaled steroids in children with non-productive cough, sustained by an evidence-based review on the management of non-specific cough in children, 38 and 2) the suggestion, reported by the Evidence-Based Clinical Practice Guidelines of the American College of Chest Physicians, to consider the use of inhaled corticosteroids when the post-infectious cough adversely affects the patient's quality of life and/or when persists despite the use of other drugs, such as ipratropium inhalation. 53 To limit the damage induced by the host's inflammatory response, and its impact on the possible associated cough and bronchoconstriction, treatment with inhaled steroids should be initiated early in the course of the infection, evaluating the opportunity to associate bronchodilators. ...
Post-infectious cough is a common symptom associated with common colds and/or upper respiratory tract infection. This cough is expected to last for only for few days and resolve spontaneously, whilst when persists for longer than three weeks is defined "persistent" and is associated tickling or an irritating sensation in the throat which often leads to paroxysms of coughing. Persistent post-infectious cough can cause morbidity since it may interfere with usual living. Despite the recent advances in understanding the mechanisms that regulate cough, in physiological and pathological conditions, current therapeutic options for post-infectious cough are little or only moderately effective.
... NK cells, resulting in their widespread use in the treatment of inflammatory disorders and in cancer (21)(22)(23)(24)(25)(26)(27)(28). However, there is also some evidence to indicate that GCs could have a stimulatory effect on immune cells, including macrophages and T cells (25,29,30). ...
Glucocorticoids (GCs) have long been known to be immune suppressive and synthetic variants are widely used in the treatment of inflammatory disorders. Here, we report that, while suppressing the initial production of interferon-γ (IFN-γ), the synthetic GC dexamethasone (Dex) enhances the proliferation and survival of natural killer (NK) cells stimulated with interleukin-2 (IL-2) + interleukin-12 (IL-12). Inhibition of mTOR complex 1 by rapamycin revealed the immunosuppressive activity of Dex was independent from the effect of enhancing NK cell proliferation. In the presence of IL-2 + IL-12, Dex also increased the percentage of NK cells that were CD16+ and DNAM1bright, increased the level of expression of CD94 or NKG2A, and improved mitochondrial function of NK cells. Moreover, NK cells treated with cytokines IL-2 and IL-12 + Dex, followed by a 7-day rest, displayed an increased IFN-γ response upon restimulation. Thus, there is a dichotomic effect of GCs on NK cell function dependent on the local cytokine milieu; the NK cell effector response is initially suppressed, but, dependent on the cytokines present, Dex can also augment the proliferation, survival, and reactivity of human NK cells in a secondary recall response.
... 49 Methylprednisolone has been shown to induce not only NK cell maturation but also to impair their cytotoxicity in vitro. 50,51 In vivo, prophylactic corticosteroids seemed to alter NK cell recovery after HSCT. 52 The similar results between related and unrelated HSCT, which we observed, suggest that antithymocyte globulin has a minor role in GvL effect, confirming previous reports. ...
There is currently a major concern regarding the optimal immunosuppression therapy to be administered after hematopoietic stem cell transplantation (HSCT) to reduce both the toxicity of GvHD and the rate of relapse. We report the outcome of high-risk leukemia children transplanted with a new way of managing cyclosporine (CsA)-based GvHD prophylaxis. A total of 110 HSCT in 109 ALL or AML children who received CsA without mycophenolate or methotrexate in matched related as well as in matched or mismatched unrelated stem cell transplantation were included. CsA dosage regimens were individualized to obtain specific trough blood concentrations values. The incidences of grade I-II and III-IV acute GvHD were 69.1% and 1.8%, respectively, and 8.4% for chronic GvHD. GvHD was neither more frequent nor severe in unrelated than in related HSCT. GvHD occurred in 87% of patients with a mean CsA trough concentration ⩽120 ng/mL versus 43% with concentration >120 ng/mL (P<0.0001). Five-year disease-free survival (DFS) and overall survival were 78% and 83.6%, respectively. DFS was 76.9% for ALL and 80.4% for AML patients. There was no difference in DFS between matched siblings and matched unrelated or mismatched unrelated HSCT. DFS in patients with minimal residual disease (MRD) ⩾10(-3) and in those with MRD <10(-3) before SCT was comparable. Our results indicate that a GvHD prophylaxis regimen based on CsA without mycophenolate or methotrexate is safe and effective whatever the donor compatibility is. These results suggest that GvL effect may be enhanced by this strategy of GvHD prophylaxis.Bone Marrow Transplantation advance online publication, 25 January 2016; doi:10.1038/bmt.2015.350.
... Therefore, it is important to define factors that may influence and modulate NK cells or other ILCs differentiation from CD34 + precursors early after transplant. Notably, the conditioning regimen, graft manipulation, residual leukemia cells, as well as opportunistic infections induce an inflammatory reaction that can interfere with hematopoiesis and in particular with NK-cell development [15,[35][36][37]. ...
NK cells are innate lymphocytes characterized by the expression of Nuclear factor interleukin 3 regulated (NFIL3 or E4BP4), Eomesodermin (EOMES) transcription factors (TF) and by the ability to exert cytolytic activity and to release IFN-γ. In the haploidentical-hematopoietic stem cell transplantation setting (haplo-HSCT), CD34(+) donor-derived NK cells play a major role in the control of leukemic relapses. Therefore, it is important to better define cytokines that influence NK-cell differentiation from CD34(+) precursors. We analyzed the effects of IL-1β on NK-cell differentiation from umbilical cord blood (UCB) CD34(+) cells. While IL-1β inhibited CD161(+) CD56(+) cell proliferation, an increased expression of LFA-1, CD94/NKG2A, KIRs, Perforin on CD56(+) cells was detected. In addition, within the CD161(+) CD56(+) IL-1RI(+) LFA-1(-) cell fraction (representing group 3 Innate Lymphoid Cells, ILC3-like cells) a significant increase of EOMES, NKp46 and CD94/NKG2A receptors, cytolytic granules and IFN-γ was detected. This increase was paralleled by a decrease of related orphan receptors (RORγt) TF, NKp44 expression and IL-22 production. These data suggest that IL-1β inhibits ILC3- while favoring NK-cell maturation. Since in haplo-HSCT conditioning regimen, infections or residual leukemia cells may induce IL-1β production, this may influence the NK/ILC3 development from donor-derived CD34(+) precursors. This article is protected by copyright. All rights reserved.
This article is protected by copyright. All rights reserved.
... ± 66.098), Table 10. Our result is in agreement with Chiara et al. [4] who found that methylprednisolone-treated normal NK cells display reduced levels of intracytoplasmic granzyme B that was done on 10 patients receiving methylprednisolone. In the present study there was no statistically significant correlation found between granzyme B and pulmonary function tests (FEV1%, FVC%, FEV1/FVC, FEF25-75%) in asthmatic patients. ...
Background: Asthma is a major public health problem with a high economic burden. It involves several inflammatory cells and multiple mediators. Granzyme B is an inflammatory mediator expressed and secreted by both immune and non immune cells. Recently it was found to play a role in the pathogenesis of asthma.
The aim of this work: was to evaluate the effect of both inhaled and oral corticosteroids on sputum granzyme B in asthmatic patients with moderate severity.
Methods: The study included 25 patients with moderate persistent asthma plus 15 healthy subjects as a control group. Granzyme B was measured before treatment with corticosteroids then after inhalation therapy and oral therapy.
Results: It was found that expected pulmonary function parameters were significantly lower in asthmatic patients than in controls. Sputum granzyme B levels were significantly higher in asthmatic patients than in controls. Sputum granzyme B levels were significantly lower after treatment with inhaled corticosteroids than basal levels. Oral corticosteroids further significantly lowered granzyme B, but the lowering effect of inhaled steroids was significantly higher than that of oral drugs. There was no statistically significant correlation between granzyme B and PFTs in asthmatic patients.
Conclusion: Granzyme B levels are elevated in bronchial asthma. Granzyme B could play a role in the pathogenesis of bronchial asthma. Both inhaled and oral corticosteroids lowered granzyme B levels significantly. The lowering effect of inhaled corticosteroids on sputum granzyme B is more than that of the oral corticosteroids.
... This could be one of the reasons that make treatment with corticosteroids inefficient in patients who have had the disease for many years. In addition to this, two things must be taken into account: both kinds of drugs can interfere with the action of B cells, although they usually do not reduce their numbers and corticosteroids produce an inhibitory effect in NK cells [42] but these cells have a lower sensitivity to these kinds of drugs [43]. These data coincide with our results where more time is needed to reduce their activation. ...
This study aimed to search the correlation among immunological profiles and clinical phenotypes of scleroderma in well-characterized groups of scleroderma patients, comparing forty-nine scleroderma patients stratified according to specific clinical phenotypes with forty-nine healthy controls. Five immunological cell subpopulations (B, CD4(+) and CD8(+) T-cells, NK, and monocytes) and their respective stages of apoptosis and activation were analyzed by flow cytometry, in samples of peripheral blood mononuclear cells (PBMCs). Analyses of results were stratified according to disease stage, time since the diagnosis, and visceral damage (pulmonary fibrosis, pulmonary hypertension, and cardiac affliction) and by time of treatment with corticosteroids. An increase in the percentages of monocytes and a decrease in the B cells were mainly related to the disease progression. A general apoptosis decrease was found in all phenotypes studied, except in localized scleroderma. An increase of B and NK cells activation was found in patients diagnosed more than 10 years ago. Specific cell populations like monocytes, NK, and B cells were associated with the type of affected organ. This study shows how, in a heterogeneous disease, proper patient's stratification according to clinical phenotypes allows finding specific cellular profiles. Our data may lead to improvements in the knowledge of prognosis factors and to aid in the analysis of future specific therapies.
... The appearance of KIR + NK cells (containing the alloreactive subset) requires four to six additional weeks. Therefore, it is conceivable that an efficient NK-mediated anti-leukemic effect occurs only after this time interval from transplantation (87,(89)(90)(91) (Figure 1C). ...
Natural Killer (NK) cells are major effector cells of the innate immunity. The discovery, over two decades ago, of major histocompatibility complex-class I-specific inhibitory NK receptors and subsequently of activating receptors, recognizing ligands expressed by tumor or virus-infected cells, paved the way to our understanding of the mechanisms of selective recognition and killing of tumor cells. Although NK cells can efficiently kill tumor cells of different histotypes in vitro, their activity may be limited in vivo by their inefficient trafficking to tumor lesions and by the inhibition of their function induced by tumor cells themselves and by the tumor microenvironment. On the other hand, the important role of NK cells has been clearly demonstrated in the therapy of high risk leukemias in the haploidentical hematopoietic stem cell (HSC) transplantation setting. NK cells derived from donor HSC kill leukemic cells residual after the conditioning regimen, thus preventing leukemia relapses. In addition, they also kill residual dendritic cells and T lymphocytes, thus preventing both GvH disease and graft rejection.
... However, the study of [38] has demonstrated the possibility that NCR expression could also be regulated by PKC in NK cells. Prolactin induces upregulation of NCR expression augmenting NK cytotoxicity against tumour cells, and vice versa corticosteroids or TGF-1 reduces NK cytotoxicity [39,40]. A recent study of [37] revealed that the conformation of PKC-binding sites might Mediators of Inflammation 9 result in posttranscriptional regulation of NKp46 expression. ...
NK cells represent a potential tool for adoptive immunotherapy against tumors. Membrane-bound Hsp70 acts as a tumor-specific marker enhancing NK cell activity. Using flow cytometry the effect of in vitro stimulation with IL-2 or IL-15 alone or in combination with Hsp70-derived 14-mer peptide (TKD) on cell surface expression of NK activatory receptors (CD16, NKG2D, NKG2C, NKp46, NKp44, NKp30, KIR2DL4, DNAM-1, and LAMP1) and NK inhibitory receptors (NKG2A, KIR2DL2/L3, LIR1/ILT-2, and NKR-P1A) in healthy individuals was studied. Results were expressed as the percentage of receptor expressing cells and the amount of receptor expressed by CD3CD56 cellular population. CD94, NKG2D, NKp44, NKp30, KIR2DL4, DNAM-1, LAMP1, NKG2A, and NKR-P1A were upregulated after the stimulation with IL-2 or IL-15 alone or in combination with TKD. KIR2DL2/L3 was upregulated only by IL-15 and IL-15/TKD. Concurrently, an increase in a number of NK cells positive for CD94, NKp44, NKp30, KIR2DL4, and LAMP1 was observed. IL-15 and IL-15/TKD caused also cell number rise positive for KIR2DL2/L3 and NKR-P1A. Cell number positive for NKG2C and NKG2A was increased only by IL-2 and IL-2/TKD. The diverse effect of IL-2 or IL-15 w or w/o TKD on cell surface expression was observed in CD16, NKp46, and LIR1/ILT-2.
Background
Allogeneic hematopoietic stem cell transplantation (HSCT) remains the standard of care for chemotherapy-refractory leukemia patients, but cure rates are still dismal. To prevent leukemia relapse following HSCT, we aim to improve the early graft-versus-leukemia effect mediated by natural killer (NK) cells. Our approach is based on the adoptive transfer of Therapeutic Inducers of Natural Killer cell Killing (ThINKK). ThINKK are expanded and differentiated from HSC, and exhibit blood plasmacytoid dendritic cell (pDC) features. We previously demonstrated that ThINKK stimulate NK cells and control acute lymphoblastic leukemia (ALL) development in a preclinical mouse model of HSCT for ALL. Here, we assessed the cellular identity of ThINKK and investigated their potential to activate allogeneic T cells. We finally evaluated the effect of immunosuppressive drugs on ThINKK-NK cell interaction.
Methods
ThINKK cellular identity was explored using single-cell RNA sequencing and flow cytometry. Their T-cell activating potential was investigated by coculture of allogeneic T cells and antigen-presenting cells in the presence or the absence of ThINKK. A preclinical human-to-mouse xenograft model was used to evaluate the impact of ThINKK injections on graft-versus-host disease (GvHD). Finally, the effect of immunosuppressive drugs on ThINKK-induced NK cell cytotoxicity against ALL cells was tested.
Results
The large majority of ThINKK shared the key characteristics of canonical blood pDC, including potent type-I interferon (IFN) production following Toll-like receptor stimulation. A minor subset expressed some, although not all, markers of other dendritic cell populations. Importantly, while ThINKK were not killed by allogeneic T or NK cells, they did not increase T cell proliferation induced by antigen-presenting cells nor worsened GvHD in vivo. Finally, tacrolimus, sirolimus or mycophenolate did not decrease ThINKK-induced NK cell activation and cytotoxicity.
Conclusion
Our results indicate that ThINKK are type I IFN producing cells with low T cell activation capacity. Therefore, ThINKK adoptive immunotherapy is not expected to increase the risk of GvHD after allogeneic HSCT. Furthermore, our data predict that the use of tacrolimus, sirolimus or mycophenolate as anti-GvHD prophylaxis regimen will not decrease ThINKK therapeutic efficacy. Collectively, these preclinical data support the testing of ThINKK immunotherapy in a phase I clinical trial.
Novel disease-modifying treatments for neuropathic pain are urgently required. The cellular immune response to nerve injury represents a promising target for therapeutic development. Recently, the role of natural killer (NK) cells in both CNS and PNS disease has been the subject of growing interest. In this opinion article, we set out the case for NK cell-based intervention as a promising avenue for development in the management of neuropathic pain. We explore the potential cellular and molecular targets of NK cells in the PNS by contrasting with their reported functional roles in CNS diseases, and we suggest strategies for using the beneficial functions of NK cells and immune-based therapeutics in the context of neuropathic pain.
Improvements in supportive care, use of reduced intensity conditioning regimens, and the widespread availability of haploidentical donors and cord blood units have led to steady increases in the number of patients undergoing hematopoietic cell transplantations (HCT). With their potent immunosuppressive capabilities, calcineurin inhibitors (CNIs) cause adverse effects including nephrotoxicity, electrolyte abnormalities, neurotoxicity, hypertension, and endocrinopathies. Sirolimus is a macrolide antibiotic, rapamycin, derived from Streptomyces hygroscopicus that was discovered to demonstrate immunosuppressive activity. Graft‐versus‐host disease (GVHD) is a complex immunologic process driven principally by allo‐reactive donor lymphocytes. Post‐HCT patients, treated with corticosteroids, often develop short‐ and long‐term effects including infectious, metabolic, psychiatric, and other complications that are a direct result of, or exacerbated by, corticosteroid use. Alemtuzumab is lymphodepleting serotherapy that has been used for GVHD treatment with variable response rates of 50‐80%. Tumor necrosis factor‐α is a cytokine in the inflammatory cascade that causes acute GVHD.
Natural killer (NK) cells are part of the innate immune system. These cells recover quickly after blood and marrow transplantation and have been implicated in graft-versus-leukemia reactions, engraftment, rejection and infection protection. NK-cell activation is controlled by a complex series of receptors that recognize MHC or MHC-like molecules that are present on the surface of potential target cells. NK effector function includes direct cytotoxicity and cytokine production. NK cells interact with a variety of components of the immune system, including dendritic cells. Recent studies suggest that it may be possible to choose related or unrelated donors based on the presence or absence of certain NK-cell genes (i.e., killer-cell immunoglobulin-like receptor) and that this might influence the risk of relapse. Lastly, adoptive cellular therapy studies with purified and cultured NK cells are ongoing and hold promise as a non-toxic, effective anticancer therapy.
Natural killer (NK) cells represent a distinct lineage of lymphocyte that is conserved in all mammals. Their dominant role is to protect against infection prior to the appearance of effective T cell responses. They may also be important, however, to antitumor responses, and to coordination of adaptive immunity. Functionally, they are much like cytotoxic T cells in using granules containing perforin and granzymes to kill target cells; they also secrete many immune-coordinating cytokines, foremost of which is interferon-?. Recent studies show NK cells to be more complex than previously suspected - transiting a multisite and multistage developmental process, engaging in critical interactions with accessory cells, and using a panoply of receptors to recognize targets. This chapter summarizes these elements of NK biology, in addition to NK cell assays and chemical agents that modulate NK function.
The evidence of NK cell implication in human diseases has initially been shown for antiviral immunity and tumor surveillance. Nowadays, increased attention is being paid to the aspect of NK cells and innate immunity in studies on autoimmune diseases. However, despite a growing knowledge on NK cell function and regulation, their role in human autoimmune disease still remains controversial. In animal models, studies of NK cells have shown conflicting results toward a disease-promoting or disease-protective role. Similarly, in human diseases, available data suggest an unequivocal role of NK cells. Yet, the understanding of NK cell implication in human diseases is far from being achieved. We review here the current knowledge on NK cell biology in human autoimmune and inflammatory diseases and discuss their possible mechanisms of action in these complex pathologies.
The effects of prednisolone on pregnancy outcome are open to debate. This meta-analysis was performed to evaluate the efficiency of prednisolone administration on unexplained recurrent miscarriage (RM) and the process of assisted reproductive technology (ART). Relevant publications were searched from databases and the included randomized controlled trials (RCTs) investigated effects of prednisolone administration in women with unexplained RM or during ART. The outcomes in our analysis were measured in terms of risk ratios (RRs) with 95% confidence intervals (CI) using random effect models. This meta-analysis was performed based on pregnancy outcomes following prednisolone treatment. This meta-analysis provides evidence that prednisolone therapy improves pregnancy outcomes in women with idiopathic RM (live birth rate: RR 1.58, 95% CI 1.23-2.02; successful pregnancy outcome: RR 7.63, 95% CI 3.71-15.69; miscarriage rate: RR 0.42, 95% CI 0.28-0.61). Our meta-analysis revealed a non-significant effect of prednisolone on pregnancy outcome during intracytoplasmic sperm injection (ICSI) cycles (pregnancy rate: RR 1.02, 95% CI 0.84-1.24; clinical pregnancy rate: RR 1.01, 95% CI 0.82-1.24; implantation rate: RR 1.04, 95% CI 0.85-1.28). Prednisolone administration may improve pregnancy outcomes in women with idiopathic RM; its efficacy in women undergoing ICSI is not significant.
© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.
Abstract Glucocorticoids (GCs) strongly impact on different T cell subsets inducing generally immunosuppressive effects, whereas much less is known about the effect of GC on natural killer (NK) cells. The aims of this study were to investigate the effects of GC on T cell functions, including T cell-mediated anti-tumor immune response, and on NK cells. We have used lck-GR mice, which overexpress a transgenic rat GR in both T and NK cells. These mice were found to have decreased both CD4(+) and CD8(+) T cell populations in the periphery. In contrast, both NK and NKT cells were found in normal numbers in lck-GR mice. To identify genes and pathways affected by GR overexpression in our system in T cells, we have compared gene expression profiles in wild-type and lck-GR T cells. Among the genes upregulated in T cells from lck-GR mice, the microarray analysis has identified genes regulating expansion of regulatory T cells. The analysis of genes downregulated in lck-GR mice has identified genes and gene associated with the regulation of immune response. With regard to the effects on T cell functions in lck-GR mice, transgenic expression of GR had a suppressive effect on killer cell activity in vitro. In addition, lck-GR mice showed an increased tumor growth in murine tumor model in vivo, which may be a possible consequence of reduced T cell numbers and activity. We conclude that an increased expression of the GR strongly affects numbers and possibly functions of T cell subsets, but has little effect on NK cells.
Natural killer (NK) cells are a key component of the innate immune system and play pivotal roles as inflammatory regulators and in tumor surveillance. Human aspartyl β-hydroxylase (HAAH) is a plasma membrane and endoplasmic reticulum protein with hydroxylation activity, which is over-expressed in many malignant neoplasms and can be detected from the sera of tumor patients. HAAH is involved in regulating tumor cell infiltration and metastasis. Escaping from immune surveillance may help tumor cell infiltration and metastasis. However, the effects of HAAH on tumor immune surveillance have not yet been investigated carefully. The present study investigated the potential use of HAAH as an immune regulator of human NK cells. We assessed the effects of recombinant HAAH (r-HAAH) on primary human NK cell morphology, viability, cytotoxicity, apoptosis, receptors expression and cytokine/cytolytic proteins production. Our results demonstrated that r-HAAH negatively affects NK cell activity in a time and dose-dependent manner. It noticeably reduces the viability of the NK cells by increasing apoptosis and necrosis via caspase signaling pathways. Moreover, r-HAAH reduces the NK cell cytotoxicity by inhibiting surface expression of NKG2D, NKp44 and IFN-γ secretion. These findings suggest that one of the ways by which HAAH actively promotes tumor formation and proliferation is by inhibiting NK cell-surveillance activity.
In a clinical phase I/II trial, pediatric patients with high-risk malignancies were treated with ex vivo IL-2-stimulated donor natural killer (NK) cells after transplantation with haploidentical stem cells. To evaluate the potential negative effects of the immunosuppressive drug mycophenolate mofetil (MMF) used for immunotherapy, the functionality and signaling of ex vivo NK cells was investigated. Our results show that during NK cell expansion, long-term (9 days) incubation with mycophenolic acid (MPA), the active metabolite of MMF, in therapeutically relevant concentrations led to the severe inhibition of NK cell proliferation. This correlated with a significantly reduced cytokine/chemokine secretion and the inhibited acquisition of surface receptors regarding cytotoxicity (e.g., NKp30, NKp44, NKp46, NKG2D), adhesion/migration (e.g., ICAM-1/CD54, LFA-1/CD11a, CD62L, CXCR3) and activation (e.g., CD25). Moreover, MPA prevented phosphorylation of the central signaling molecules STAT-3/-4/-5, AKT and ERK1/2. In contrast, short-term (24 h) MPA incubation of IL-2-stimulated NK cells had no or only marginal effects on the activated NK cell phenotype, including receptor expression, cytokine/chemokine secretion and intracellular signaling. Further, short-term MPA incubation only moderately affected the highly cytotoxic activity of previously IL-2-stimulated NK cells. In conclusion, while long-term MPA incubation significantly compromised ex vivo NK cell functionality, previously IL-2-activated NK cells seemed to be rather resistant to short-term MPA treatment. This finding supports the use of IL-2-activated NK cells as immunotherapy, especially for patients treated with MMF after haploidentical stem cell transplantation.
Lung transplantation is a recognised treatment for patients with end stage pulmonary disease. Transplant recipients receive life-long administration of immunosuppressive drugs that target T cell mediated graft rejection. However little is known of the impact on NK cells, which have the potential to be alloreactive in response to HLA-mismatched ligands on the lung allograft and in doing so, may impact negatively on allograft survival. NK cells from 20 healthy controls were assessed in response to Cyclosporine A, Mycophenolic acid (MPA; active form of Mycophenolate mofetil) and Prednisolone at a range of concentrations. The impact of these clinically used immunosuppressive drugs on cytotoxicity (measured by CD107a expression), IFN-γ production and CFSE proliferation was assessed in response to various stimuli including MHC class-I negative cell lines, IL-2/IL-12 cytokines and PMA/Ionomycin. Treatment with MPA and Prednisolone revealed significantly reduced CD107a expression in response to cell line stimulation. In comparison, addition of MPA and Cyclosporine A displayed reduced CD107a expression and IFN-γ production following PMA/Ionomycin stimulation. Diminished proliferation was observed in response to treatment with each drug. Additional functional inhibitors (LY294002, PD98059, Rottlerin, Rapamycin) were used to elucidate intracellular pathways of NK cell activation in response to stimulation with K562 or PMA-I. CD107a expression was significantly decreased with the addition of PD98059 following K562 stimulation. Similarly, CD107a expression significantly decreased following PMA-I stimulation with the addition of LY294002, PD98059 and Rottlerin. Ten lung transplant patients, not receiving immunosuppressive drugs pre-transplant, were assessed for longitudinal changes post-transplant in relation to the administration of immunosuppressive drugs. Individual patient dynamics revealed different longitudinal patterns of NK cell function post-transplantation. These results provide mechanistic insights into pathways of NK cell activation and show commonly administered transplant immunosuppression agents and clinical rejection/infection events have differential effects on NK cell function that may impact the immune response following lung transplantation.
A role in pulmonary immunity has been ascribed to Natural Killer (NK) cells and several in vitro studies have shown a corticosteroid-induced inhibition of NK cells mediated cytotoxicity. Several clinical trials on chronic obstructive pulmonary disease (COPD) have suggested a relationship between COPD treatment and occurrence of respiratory infections. Aims of our study were to investigate if real life COPD treatment affects peripheral blood NK cells total count and their receptors expression and to assess if different doses of formoterol and budesonide, administered alone or in combination, are able to modulate the surface expression of activating (NKp30, NKp44, NKp46 and NKG2D) and inhibitory (KIR2DL2/L3, KIR3DL1 and NKG2A) receptors on peripheral blood NK cells of COPD patients. Moreover, we evaluated the potential effect of treatment with budesonide and/or formoterol on IFN-γ secretion in vitro. NK cells were isolated from peripheral blood of 7 healthy volunteers, 9 chronic bronchitis (CB) and 11 COPD patients. Total NK cells count and activating and inhibitory receptors expression were evaluated. NK cells were cultured for 20h in 96-well plates with IL-2 (100IU/ml)+IL-12 (2.5ng/ml), with or without budesonide (Bud; 1 and 0.01μM) and formoterol (For; 30 and 0.3nM) alone or in combination. Cells were analyzed by flow cytometry and IFN-γ was measured in cell supernatants by ELISA test. No difference between real life treated COPD, CB and healthy subjects was found concerning NK total count and NK cell receptors expression. When cells were stimulated over night with cytokines and treated with drugs, only NKG2D receptor was modulated. Its expression was significantly downregulated by budesonide alone and in combination with formoterol in COPD patients. IFN-γ production induced by stimulation with IL-2+IL-12 was decreased in a highly significant way (p<0.01) by all treatments in all groups. Even if in vitro experiments with budesonide, alone or in combination with formoterol, showed a modulation of NKG2D receptor expression and IFN-γ production, our ex vivo results show that real life LABA and ICS treatment does not influence peripheral NK cells count and their receptors phenotype.
Many patients with melanoma show spontaneous T cell responses against tumor antigens, and induction or amplification of these
T cells responses can frequently be achieved through vaccination or adoptive T cell transfer. However, tumor responses as
measured by tumor shrinkage remain infrequent. These observations have argued for analysis of the tumor microenvironment in
metastatic melanoma for potential mechanisms of resistance to immune effector function at the level of the target tumor site.
This review discusses two categories of regulation at the level of the tumor microenvironment, chemokine-mediated migration
of effector T cells and active immune suppression, that have been identified through gene expression profiling of human specimens.
Melanoma cell-intrinsic apoptosis also is discussed. The identification of these mechanisms points toward new strategies of
intervention to consider for improving the clinical efficacy of T cell-based immunotherapy for cancer, and also suggest that
molecular profiling of tumors might be used as a strategy for stratifying patients enrolled on immunotherapy clinical trials.
KeywordsMelanoma-gene expression profiling-chemokines-T cells-immune suppression
Stress-inducible MICA, a distant homolog of major histocompatibility complex (MHC) class I, functions as an antigen for γδ
T cells and is frequently expressed in epithelial tumors. A receptor for MICA was detected on most γδ T cells, CD8+αβ T cells, and natural killer (NK) cells and was identified as NKG2D. Effector cells from all these subsets could be stimulated
by ligation of NKG2D. Engagement of NKG2D activated cytolytic responses of γδ T cells and NK cells against transfectants and
epithelial tumor cells expressing MICA. These results define an activating immunoreceptor-MHC ligand interaction that may
promote antitumor NK and T cell responses.
Glucocorticoids inhibit the proinflammatory activities of transcription factors such as AP-1 and NF-B as well as that of diverse cellular signaling molecules. One of these signaling molecules is the extracellular signal-regulated kinase (Erk-1/2) that controls the release of allergic mediators and the induction of proinflammatory cytokine gene expression in mast cells. The mechanism of inhibition of Erk-1/2 activity by glucocorticoids is unknown. Here we report a novel dual action of glucocorticoids for this inhibition. Glucocorticoids increase the expression of the MAP kinase phosphatase-1 (MKP-1) gene at the promoter level, and attenuate proteasomal degradation of MKP-1, which we report to be triggered by activation of mast cells. Both induction of MKP-1 expression and inhibition of its degradation are necessary for glucocorticoid-mediated inhibition of Erk-1/2 activation. In NIH-3T3 fibroblasts, although glucocorticoids up-regulate the MKP-1 level, they do not attenuate the proteasomal degradation of this protein and consequently they are unable to inhibit Erk-1/2 activity. These results identify MKP-1 as essential for glucocorticoid-mediated control of Erk-1/2 activation and unravel a novel regulatory mechanism for this anti-inflammatory drug.
IL-2 potentiates both growth and cytotoxic function of T lymphocytes and NK cells. Resting peripheral blood NK cells can respond directly to rIL-2, without requirement for accessory cells or cofactors, and enhanced cytotoxicity can be measured within a few hours after exposure to this lymphokine. In this study, we describe an activation antigen, Leu-23, that is rapidly induced and phosphorylated after IL-2 stimulation of NK cells and a subset of low buoyant density T lymphocytes. Previously, it has been uncertain whether all NK cells or only a subset are responsive to IL-2. Since within 18 h after exposure to IL-2, essentially all NK cells express Leu-23, these findings indicate that all peripheral blood NK cells are responsive to stimulation by IL-2. The Leu-23 antigen is a disulfide-bonded homodimer, composed of 24-kD protein subunits with two N-linked oligosaccharides. Appearance of this glycoprotein on NK cells is IL-2 dependent and closely parallels IL-2-induced cytotoxicity against NK-resistant solid tumor cell targets.
Lymphoid cells with natural killer (NK)-like function, morphology, and antigenic phenotype have been identified in a mixed lymphocyte culture generated by co-culture of human peripheral blood mononuclear cells with an allogeneic human B lymphoblastoid cell line CCRF-SB. The majority of these mixed lymphocyte (MLR)-response activated NK cells express the Leu-11 surface antigen, but do not express certain T cell-associated antigens (Leu-1, Leu-3, and Leu-4) or the mature monocyte specific antigen, Leu-M3. Unlike most freshly isolated Leu-11+ human NK cells, the MLR-activated Leu-11+ cells expressed class II major histocompatibility antigens, DR and DC. Concomitant with expression of class II gene products, the Leu-11+,DR+ NK cells demonstrate enhanced cytotoxicity against the NK-sensitive tumor cell line K562. The presence of mitotic cells in the Leu-11+,DR+ population and the acquisition of increased levels of transferrin receptor on the cell surface were further indicators of activation of these cells. The direct precursors of the MLR-activated Leu-11+,DR+ cell were Leu-11+ cells that lacked expression of another NK-associated antigen Leu-7, i.e., Leu-7-,11+. These studies provided a definitive identification of the "NK-like" cell in MLR cultures and thus allow quantitation and isolation of these cells for further study.
The ability of glucocorticoids to suppress cellular immune functions, including the cytotoxic activity of natural killer cells, is well known. However, the molecular mechanism(s) of glucocorticoid-mediated suppression of cellular cytotoxicity mediated by natural killer cells is not understood. We have investigated the effects of cortisol on protein expression and cytotoxic function of natural killer cells using NK3.3, a well-characterized, cloned human natural killer cell line. Cortisol, at concentrations up to 2 microM, does not significantly alter the viability or proliferative capacity of NK3.3 cells. However, micromolar concentrations of cortisol induce the expression of a small set of proteins which are not synthesized by NK3.3 cells in the absence of cortisol, and repress the synthesis of another set of proteins including several phenotypic determinants and cytokines. In the presence of added cortisol, the synthesis of perforin mRNA was partially repressed. However, the most striking effect of cortisol on this NK clone was its repression of granzyme A synthesis. In conjunction with the downregulation of adhesion proteins, NK3.3 cells cultured in the presence of cortisol exhibit a reduced capacity to form conjugates with K562 target cells. Whereas cortisol treatment of NK3.3 cells causes an approximately 50% decrease in their ability to form conjugates with K.562 target cells, the cytotoxic function of these cells is completely abolished under the same conditions. This first report of hormonal regulation of granzyme expression and the strong correlation between granzyme A repression and cytotoxic function suggests that cortisol may regulate NK function by repression of granzyme A synthesis. In addition to demonstrating the significant influence of cortisol on natural killer cell function, these studies provide a model system for elucidation of molecular mechanism(s) whereby glucocorticoids repress cellular immune function, especially with respect to natural killer cells.
The activity of c-Jun, the major component of the transcription factor AP-1, is potentiated by amino-terminal phosphorylation on serines 63 and 73 (Ser-63/73). This phosphorylation is mediated by the Jun amino-terminal kinase (JNK) and required to recruit the transcriptional coactivator CREB-binding protein (CBP). AP-1 function is antagonized by activated members of the steroid/thyroid hormone receptor superfamily. Recently, a competition for CBP has been proposed as a mechanism for this antagonism. Here we present evidence that hormone-activated nuclear receptors prevent c-Jun phosphorylation on Ser-63/73 and, consequently, AP-1 activation, by blocking the induction of the JNK signaling cascade. Consistently, nuclear receptors also antagonize other JNK-activated transcription factors such as Elk-1 and ATF-2. Interference with the JNK signaling pathway represents a novel mechanism by which nuclear hormone receptors antagonize AP-1. This mechanism is based on the blockade of the AP-1 activation step, which is a requisite to interact with CBP. In addition to acting directly on gene transcription, regulation of the JNK cascade activity constitutes an alternative mode whereby steroids and retinoids may control cell fate and conduct their pharmacological actions as immunosupressive, anti-inflammatory, and antineoplastic agents.
NK cells are regulated by opposing signals from receptors that activate and inhibit effector function. While positive stimulation may be initiated by an array of costimulatory receptors, specificity is provided by inhibitory signals transduced by receptors for MHC class I. Three distinct receptor families, Ly49, CD94/NKG2, and KIR, are involved in NK cell recognition of polymorphic MHC class I molecules. A common pathway of inhibitory signaling is provided by ITIM sequences in the cytoplasmic domains of these otherwise structurally diverse receptors. Upon ligand binding and activation, the inhibitory NK cell receptors become tyrosine phosphorylated and recruit tyrosine phosphatases, SHP-1 and possibly SHP-2, resulting in inhibition of NK cell-mediated cytotoxicity and cytokine expression. Recent studies suggest these inhibitory NK cell receptors are members of a larger superfamily containing ITIM sequences, the inhibitory receptor superfamily (IRS).
After culture in interleukin (IL)-2, natural killer (NK) cells acquire an increased capability of mediating non-major histocompatibility complex (MHC)-restricted tumor cell lysis. This may reflect, at least in part, the de novo expression by NK cells of triggering receptors involved in cytolysis. In this study we identified a novel 44-kD surface molecule (NKp44) that is absent in freshly isolated peripheral blood lymphocytes but is progressively expressed by all NK cells in vitro after culture in IL-2. Different from other markers of cell activation such as CD69 or VLA.2, NKp44 is absent in activated T lymphocytes or T cell clones. Since NKp44 was not detected in any of the other cell lineages analyzed, it appears as the first marker specific for activated human NK cells. Monoclonal antibody (mAb)-mediated cross-linking of NKp44 in cloned NK cells resulted in strong activation of target cell lysis in a redirected killing assay. This data indicated that NKp44 can mediate triggering of NK cell cytotoxicity. mAb-mediated masking of NKp44 resulted in partial inhibition of cytolytic activity against certain (FcgammaR-negative) NK-susceptible target cells. This inhibition was greatly increased by the simultaneous masking of p46, another recently identified NK-specific triggering surface molecule. These data strongly suggest that NKp44 functions as a triggering receptor selectively expressed by activated NK cells that, together with p46, may be involved in the process of non-MHC-restricted lysis. Finally, we show that p46 and NKp44 are coupled to the intracytoplasmic transduction machinery via the association with CD3zeta or KARAP/DAP12, respectively; these associated molecules are tyrosine phosphorylated upon NK cell stimulation.
Antigen stimulation of IgE-sensitized rat basophilic leukemia RBL-2H3 cells induced activation of c-Jun N-terminal kinase (JNK) within a few minutes with maximum activity attained 40 min later. The increase in JNK activity was accompanied with an increase in phosphorylation of c-Jun in the cells. The Ag-induced JNK activation was inhibited by the phosphatidylinositol 3-kinase inhibitors wortmannin (10-100 nM) and LY 294002 (100 microM) but not by the protein kinase C inhibitors calphostin C (1 and 3 microM) and Ro 31-8425 (1 and 3 microM). Pretreatment with dexamethasone (10 and 100 nM) for 18 h inhibited the Ag-induced increase in JNK activity in a concentration-dependent manner. At least 6 h of preincubation with dexamethasone was necessary to inhibit the Ag-induced JNK activation. The phosphorylation of c-Jun induced by the Ag stimulation was reduced by pretreatment with dexamethasone without reduction of the content of c-Jun protein. The Ag-induced activation of the JNK kinase kinase mitogen-activated protein kinase-extracellular signal-regulated kinase kinase-1 was also inhibited by pretreatment with dexamethasone at 10 and 100 nM. These findings indicate that dexamethasone reduces JNK protein level and inhibits the Ag-induced activation of JNK resulting in the inhibition of c-Jun phosphorylation.
Surface receptors involved in natural killer (NK) cell triggering during the process of tumor cell lysis have recently been identified. Of these receptors, NKp44 is selectively expressed by IL-2- activated NK cells and may contribute to the increased efficiency of activated NK cells to mediate tumor cell lysis. Here we describe the molecular cloning of NKp44. Analysis of the cloned cDNA indicated that NKp44 is a novel transmembrane glycoprotein belonging to the Immunoglobulin superfamily characterized by a single extracellular V-type domain. The charged amino acid lysine in the transmembrane region may be involved in the association of NKp44 with the signal transducing molecule killer activating receptor-associated polypeptide (KARAP)/DAP12. These molecules were found to be crucial for the surface expression of NKp44. In agreement with data of NKp44 surface expression, the NKp44 transcripts were strictly confined to activated NK cells and to a minor subset of TCR-gamma/delta+ T lymphocytes. Unlike genes coding for other receptors involved in NK cell triggering or inhibition, the NKp44 gene is on human chromosome 6.
Major histocompatibility complex class I-specific inhibitory receptors on natural killer cells prevent the lysis of healthy autologous cells. The outcome of this negative signal is not anergy or apoptosis of natural killer cells but a transient abortion of activation signals. The natural killer inhibitory receptors fulfill this function by recruiting the tyrosine phosphatase SHP-1 through a cytoplasmic immunoreceptor tyrosine-based inhibition motif. This immunoreceptor tyrosine-based inhibition motif has become the hallmark of a growing family of receptors with inhibitory potential, which are expressed in various cell types such as monocytes, macrophages, dendritic cells, leukocytes, and mast cells. Most of the natural killer inhibitory receptors and two members of a monocyte inhibitory-receptor family bind major histocompatibility complex class I molecules. Ligands for many of the other receptors have yet to be identified. The inhibitory-receptor superfamily appears to regulate many types of immune responses by blocking cellular activation signals.
We evaluated 18,014 patients who underwent allogeneic bone marrow transplantation (BMT) at 235 centers worldwide to examine the incidence of and risk factors for posttransplant lymphoproliferative disorders (PTLD). PTLD developed in 78 recipients, with 64 cases occurring less than 1 year after transplantation. The cumulative incidence of PTLD was 1.0% +/- 0.3% at 10 years. Incidence was highest 1 to 5 months posttransplant (120 cases/10,000 patients/yr) followed by a steep decline to less than 5/10,000/yr among >/=1-year survivors. In multivariate analyses, risk of early-onset PTLD (<1 year) was strongly associated (P <.0001) with unrelated or human leukocyte antigen (HLA) mismatched related donor (relative risk [RR] = 4.1), T-cell depletion of donor marrow (RR = 12.7), and use of antithymocyte globulin (RR = 6.4) or anti-CD3 monoclonal antibody (RR = 43.2) for prophylaxis or treatment of acute graft-versus-host disease (GVHD). There was a weaker association with the occurrence of acute GVHD grades II to IV (RR = 1.9, P =.02) and with conditioning regimens that included radiation (RR = 2.9, P =.02). Methods of T-cell depletion that selectively targeted T cells or T plus natural killer (NK) cells were associated with markedly higher risks of PTLD than methods that removed both T and B cells, such as the CAMPATH-1 monoclonal antibody or elutriation (P =.009). The only risk factor identified for late-onset PTLD was extensive chronic GVHD (RR = 4.0, P =.01). Rates of PTLD among patients with 2 or >/=3 major risk factors were 8.0% +/- 2.9% and 22% +/- 17.9%, respectively. We conclude that factors associated with altered immunity and T-cell regulatory mechanisms are predictors of both early- and late-onset PTLD.
Two major receptors involved in human natural cytotoxicity, NKp46 and NKp44, have recently been identified. However, experimental
evidence suggested the existence of additional such receptor(s). In this study, by the generation of monoclonal antibodies
(mAbs), we identified NKp30, a novel 30-kD triggering receptor selectively expressed by all resting and activated human natural
killer (NK) cells. Although mAb-mediated cross-linking of NKp30 induces strong NK cell activation, mAb-mediated masking inhibits
the NK cytotoxicity against normal or tumor target cells. NKp30 cooperates with NKp46 and/or NKp44 in the induction of NK-mediated
cytotoxicity against the majority of target cells, whereas it represents the major triggering receptor in the killing of certain
tumors. This novel receptor is associated with CD3ζ chains that become tyrosine phosphorylated upon sodium pervanadate treatment
of NK cells. Molecular cloning of NKp30 cDNA revealed a member of the immunoglobulin superfamily, characterized by a single
V-type domain and a charged residue in the transmembrane portion. Moreover, we show that NKp30 is encoded by the previously
identified 1C7 gene, for which the function and the cellular distribution of the putative product were not identified in previous
studies.
2B4 is a surface molecule involved in activation of the natural killer (NK) cell-mediated cytotoxicity. It binds a protein termed Src homology 2 domain-containing protein (SH2D1A) or signaling lymphocyte activation molecule (SLAM)-associated protein (SAP), which in turn has been proposed to function as a regulator of the 2B4-associated signal transduction pathway. In this study, we analyzed patients with X-linked lymphoproliferative disease (XLP), a severe inherited immunodeficiency characterized by critical mutations in the SH2D1A gene and by the inability to control Epstein-Barr virus (EBV) infections. We show that, in these patients, 2B4 not only fails to transduce triggering signals, but also mediates a sharp inhibition of the NK-mediated cytolysis. Other receptors involved in NK cell triggering, including CD16, NKp46, NKp44, and NKp30, displayed a normal functional capability. However, their activating function was inhibited upon engagement of 2B4 molecules. CD48, the natural ligand of 2B4, is highly expressed on the surface of EBV(+) B cell lines. Remarkably, NK cells from XLP patients could not kill EBV(+) B cell lines. This failure was found to be the consequence of inhibitory signals generated by the interaction between 2B4 and CD48, as the antibody-mediated disruption of the 2B4-CD48 interaction restored lysis of EBV(+) target cells lacking human histocompatibility leukocyte antigen (HLA) class I molecules. In the case of autologous or allogeneic (HLA class I(+)) EBV(+) lymphoblastoid cell lines, restoration of lysis was achieved only by the simultaneous disruption of 2B4-CD48 and NK receptor-HLA class I interactions. Molecular analysis revealed that 2B4 molecules isolated from either XLP or normal NK cells were identical. As expected, in XLP-NK cells, 2B4 did not associate with SH2D1A, whereas similar to 2B4 molecules isolated from normal NK cells, it did associate with Src homology 2 domain-containing phosphatase 1.
In humans, natural killer (NK) cell function is regulated by a series of receptors and coreceptors with either triggering or inhibitory activity. Here we describe a novel 60-kD glycoprotein, termed NTB-A, that is expressed by all human NK, T, and B lymphocytes. Monoclonal antibody (mAb)-mediated cross-linking of NTB-A results in the induction of NK-mediated cytotoxicity. Similar to 2B4 (CD244) functioning as a coreceptor in the NK cell activation, NTB-A also triggers cytolytic activity only in NK cells expressing high surface densities of natural cytotoxicity receptors. This suggests that also NTB-A may function as a coreceptor in the process of NK cell activation. Molecular cloning of the cDNA coding for NTB-A molecule revealed a novel member of the immunoglobulin superfamily belonging to the CD2 subfamily. NTB-A is characterized, in its extracellular portion, by a distal V-type and a proximal C2-type domain and by a cytoplasmic portion containing three tyrosine-based motifs. NTB-A undergoes tyrosine phosphorylation and associates with the Src homology 2 domain-containing protein (SH2D1A) as well as with SH2 domain-containing phosphatases (SHPs). Importantly, analysis of NK cells derived from patients with X-linked lymphoproliferative disease (XLP) showed that the lack of SH2D1A protein profoundly affects the function not only of 2B4 but also of NTB-A. Thus, in XLP-NK cells, NTB-A mediates inhibitory rather than activating signals. These inhibitory signals are induced by the interaction of NTB-A with still undefined ligands expressed on Epstein-Barr virus (EBV)-infected target cells. Moreover, mAb-mediated masking of NTB-A can partially revert this inhibitory effect while a maximal recovery of target cell lysis can be obtained when both 2B4 and NTB-A are simultaneously masked. Thus, the altered function of NTB-A appears to play an important role in the inability of XLP-NK cells to kill EBV-infected target cells.
During the innate response to many inflammatory and infectious stimuli, dendritic cells (DCs) undergo a differentiation process termed maturation. Mature DCs activate antigen-specific naive T cells. Here we show that both immature and mature DCs activate resting human natural killer (NK) cells. Within 1 wk the NK cells increase two-- to fourfold in numbers, start secreting interferon (IFN)-gamma, and acquire cytolytic activity against the classical NK target LCL721.221. The DC-activated NK cells then kill immature DCs efficiently, even though the latter express substantial levels of major histocompatibility complex (MHC) class I. Similar results are seen with interleukin (IL)-2--activated NK cell lines and clones, i.e., these NK cells kill and secrete IFN-gamma in response to immature DCs. Mature DCs are protected from activated NK lysis, but lysis takes place if the NK inhibitory signal is blocked by a human histocompatibility leukocyte antigen (HLA)-A,B,C--specific antibody. The NK activating signal mainly involves the NKp30 natural cytotoxicity receptor, and not the NKp46 or NKp44 receptor. However, both immature and mature DCs seem to use a NKp30 independent mechanism to act as potent stimulators for resting NK cells. We suggest that DCs are able to control directly the expansion of NK cells and that the lysis of immature DCs can regulate the afferent limb of innate and adaptive immunity.
The cytolytic function of natural killer (NK) cells is induced by the engagement of a series of activating receptors and coreceptors some of which have recently been identified and collectively termed natural cytotoxicity receptors (NCRs). Here, we analyzed the cytolytic function of NK cells obtained from patients with acute myeloid leukemia (AML). In sharp contrast with healthy donors, in most (16 of 18) patients with AML the majority of NK cells displayed low NCR surface density (NCR(dull)). This phenotype correlated with a weak cytolytic activity against autologous leukemic cells that could not be reversed by the monoclonal antibody-mediated disruption of HLA class I/killer immunoglobulinlike receptor interaction. The remaining 2 patients were characterized by NK cells having an NCR(bright) phenotype. Surprisingly, although displaying NCR-mediated cytolytic activity, these NCR(bright) NK cells were unable to kill autologous leukemic blasts. Importantly, the leukemic blasts from these 2 patients were also resistant to lysis mediated by normal NCR(bright) allogeneic NK cells. Our study suggests that in most instances the inability of NK cells to kill autologous leukemic blasts is consequent to low NCR surface expression. In few cases, however, this failure appears to involve a mechanism of tumor escape based on down-regulation of ligands relevant for NCR-mediated target cell recognition.
The surface density of the triggering receptors responsible for the natural killer (NK)-mediated cytotoxicity is crucial for the ability of NK cells to kill susceptible target cells. In this study, we show that transforming growth factor beta1 (TGFbeta1) down-regulates the surface expression of NKp30 and in part of NKG2D but not that of other triggering receptors such as NKp46. The TGFbeta1-mediated inhibition of NKp30 surface expression reflects gene regulation at the transcriptional level. NKp30 has been shown to represent the major receptor involved in the NK-mediated killing of dendritic cells. Accordingly, the TGFbeta1-dependent down-regulation of NKp30 expression profoundly inhibited the NK-mediated killing of dendritic cells. On the contrary, killing of different NK-susceptible tumor cell lines was variably affected, reflecting the differential usage of NKp30 and/or NKG2D in the lysis of such tumors. Our present data suggest a possible mechanism by which TGFbeta1-producing dendritic cells may acquire resistance to the NK-mediated attack.
The occurrence of post-transplant lymphoproliferative disorder (PTLD) in relation to immunosuppressive treatment was determined in 257 patients treated with non-T-cell-depleted allogeneic stem cell transplantation from an HLA-matched sibling (173 patients) or unrelated donor (84 patients). The conditioning consisted of total body irradiation and cyclophosphamide (myeloablative conditioning, 250 patients), or fludarabine combined with cyclophosphamide or a single 2 Gy dose of TBI (nonmyeloablative conditioning, seven patients). In transplantations from an unrelated donor, the patients also received antithymocyte globulin (ATG). The prophylaxis against graft-versus-host disease (GVHD) consisted of cyclosporine A, methotrexate, and methylprednisolone. The autopsy reports of deceased patients were systematically reviewed, and the autopsy materials of cases suggestive of PTLD were re-examined histologically for Epstein-Barr virus (EBV). Nineteen patients with EBV-positive PTLD were identified, of whom six had been transplanted from a sibling donor and 13 from an unrelated donor. All the patients who developed PTLD had been given ATG either for the treatment of steroid-resistant acute GVHD (all PTLD patients with a sibling donor and one with an unrelated donor), or as part of the conditioning (all patients with an unrelated donor). In conclusion, in transplantations from an HLA-identical donor with a non-T-cell-depleted graft, the risk of PTLD correlated strongly with the intensity of the immunosuppressive treatment.
Human natural killer (NK) cells express a series of activating receptors and coreceptors that are involved in recognition and killing of target cells. In this study, in an attempt to identify the cellular ligands for such triggering surface molecules, mice were immunized with NK-susceptible target cells. On the basis of a functional screening, four mAbs were selected that induced a partial down-regulation of the NK-mediated cytotoxicity against the immunizing target cells. As revealed by biochemical analysis, three of such mAbs recognized molecules of approximately 70 kD. The other mAb reacted with two distinct molecules of approximately 65 and 60 kD, respectively. Protein purification followed by tryptic digestion and mass spectra analysis, allowed the identification of the 70 kD and the 65/60 kD molecules as PVR (CD155) and Nectin-2 delta/alpha (CD112), respectively. PVR-Fc and Nectin-2-Fc soluble hybrid molecules brightly stained COS-7 cells transfected with the DNAM-1 (CD226) construct, thus providing direct evidence that both PVR and Nectin-2 represent specific ligands for the DNAM-1 triggering receptor. Finally, the surface expression of PVR or Nectin-2 in cell transfectants resulted in DNAM-1-dependent enhancement of NK-mediated lysis of these target cells. This lysis was inhibited or even virtually abrogated upon mAb-mediated masking of DNAM-1 (on NK cells) or PVR or Nectin-2 ligands (on cell transfectants).
The surface density of the triggering receptors responsible for the natural killer (NK)-mediated cytotoxicity is crucial for the ability of INK cells to kill susceptible target cells. In this study, we show that transforming growth factor beta1 (TGFbeta1) down-regulates the surface expression of NKp30 and in part of NKG2D but not that of other triggering receptors such as NKp46. The TGFbeta1-mediated inhibition of NKp30 surface expression reflects gene regulation at the transcriptional level. NKp30 has been shown to represent the major receptor involved in the NK-mediated killing of dendritic cells. Accordingly, the TGFbeta1-dependent down-regulation of NKp30 expression profoundly inhibited the NK-mediated killing of dendritic cells. On the contrary, killing of different NK-susceptible tumor cell lines was variably affected, reflecting the differential usage of NKp30 and/or NKG2D in the lysis of such tumors. Our present data suggest a possible mechanism by which TGFbeta1producing dendritic cells may acquire resistance to the NK-mediated attack.
1. Glucocorticoids are widely used for the suppression of inflammation in chronic inflammatory diseases such as asthma, rheumatoid arthritis, inflammatory bowel disease and autoimmune diseases, all of which are associated with increased expression of inflammatory genes. The molecular mechanisms involved in this antiinflammatory action of glucocorticoids is discussed, particularly in asthma, which accounts for the highest clinical use of these agents.
2. Glucocorticoids bind to glucocorticoid receptors in the cytoplasm which then dimerize and translocate to the nucleus, where they bind to glucocorticoid response elements (GRE) on glucocorticoid-responsive genes, resulting in increased transcription. Glucocorticoids may increase the transcription of genes coding for antiinflammatory proteins, including lipocortin-1, interleukin-10, interleukin-1 receptor antagonist and neutral endopeptidase, but this is unlikely to account for all of the widespread anti-inflammatory actions of glucocorticoids.
3. The most striking effect of glucocorticoids is to inhibit the expression of multiple inflammatory genes (cytokines, enzymes, receptors and adhesion molecules). This cannot be due to a direct interaction between glucocorticoid receptors and GRE, as these binding sites are absent from the promoter regions of most inflammatory genes. It is more likely to be due to a direct inhibitory interaction between activated glucocorticoid receptors and activated transcription factors, such as nuclear factor-κB and activator protein-1, which regulate the inflammatory gene expression.
4. It is increasingly recognized that glucocorticoids change the chromatin structure. Glucocorticoid receptors also interact with CREB-binding protein (CBP), which acts as a co-activator of transcription, binding several other transcription factors that compete for binding sites on this molecule. Increased transcription is associated with uncoiling of DNA wound around histone and this is secondary to acetylation of the histone residues by the enzymic action of CBP. Glucocorticoids may lead to deacetylation of histone, resulting in tighter coiling of DNA and reduced access of transcription factors to their binding sites, thereby suppressing gene expression.
5. Rarely patients with chronic inflammatory diseases fail to respond to glucocorticoids, although endocrine function of steroids is preserved. This may be due to excessive formation of activator protein-1 at the inflammatory site, which consumes activated glucocorticoid receptors so that they are not available for suppressing inflammatory genes.
6. This new understanding of glucocorticoid mechanisms may lead to the development of novel steroids with less risk of side effects (which are due to the endocrine and metabolic actions of steroids). ‘Dissociated’ steroids which are more active in transrepression (interaction with transcription factors) than transactivation (GRE binding) have now been developed. Some of the transcription factors that are inhibited by glucocorticoid, such as nuclear factor-κB, are also targets for novel anti-inflammatory therapies.
Lymphoid cells with natural killer (NK)-like function, morphology, and antigenic phenotype have been identified in a mixed lymphocyte culture generated by co-culture of human peripheral blood mononuclear cells with an allogeneic human B lymphoblastoid cell line CCRF-SB. The majority of these mixed lymphocyte (MLR)-response activated NK cells express the Leu-11 surface antigen, but do not express certain T cell-associated antigens (Leu-1, Leu-3, and Leu-4) or the mature monocyte specific antigen, Leu-M3. Unlike most freshly isolated Leu-11+ human NK cells, the MLR-activated Leu-11+ cells expressed class II major histocompatibility antigens, DR and DC. Concomitant with expression of class II gene products, the Leu-11+,DR+ NK cells demonstrate enhanced cytotoxicity against the NK-sensitive tumor cell line K562. The presence of mitotic cells in the Leu-11+,DR+ population and the acquisition of increased levels of transferrin receptor on the cell surface were further indicators of activation of these cells. The direct precursors of the MLR-activated Leu-11+,DR+ cell were Leu-11+ cells that lacked expression of another NK-associated antigen Leu-7, i.e., Leu-7-,11+. These studies provided a definitive identification of the "NK-like" cell in MLR cultures and thus allow quantitation and isolation of these cells for further study.
NKG2D is a recently described activating receptor expressed by both NK cells and CTL. In this study we investigated the role of NKG2D in the natural cytolysis mediated by NK cell clones. The role of NKG2D varied depending on the type of target cells analyzed. Lysis of various tumors appeared to be exclusively natural cytotoxicity receptors (NCR) dependent. In contrast, killing of anothergroup of target cells, including not only the epithelial cell lines HELA and IGROV-1, but also the FO-1 melanoma, the JA3 leukemia, the Daudi Burkitt lymphoma and even normal PHA-induced lymphoblasts, involved both NCR and NKG2D. Notably, NK cell clones expressing low surface densities of NCR (NCRdull) could lyse these tumors in an exclusively NKG2D-dependent fashion. Remarkably, notall of these targets expressed MICA/B, thus implying the existence of additional ligands recognized by NKG2D, possibly represented by GPI-linked molecules. Finally, we show that the engagement of different HLA class I-specific inhibitory receptors by either specific antibodies or the appropriate HLA class I ligand led to inhibition of NKG2D-mediated NK cell triggering.
Corticosteroids have previously been reported to partially inhibit the natural cytotoxic activity of peripheral blood lymphocytes. However, since only a few percent of peripheral lymphocytes are natural killer (NK) cells, it has not been possible to determine whether corticosteroids directly inhibit NK cells or mediate this effect via other cell types. This report documents direct functional inactivation, but unimpeded proliferation, of cloned human NK cells by subphysiologic levels of cortisol. In contrast, high concentrations of testosterone, progesterone or estradiol had no significant effect on proliferation or cytotoxic activity of the cloned NK cells. The kinetics of inhibition of NK function by cortisol are consistent with a transcription-dependent mechanism.
The in vitro effect of prednisolone (PRD) on NK and ADCC activities of human lymphocytes was investigated. PRD at concentrations ranging from 7.5 X 10(-3) to 1 X 10(-5) M significantly inhibited NK activity, while concentrations of 7.5 X 10(-3) to 1 X 10(-4) M inhibited ADCC activities of PBL when added directly to the mixture of effector and target cells. Lymphocytes pre-cultured for 24 hr with PRD at concentrations ranging from 1 X 10(-4) M to 1 X 10(-6) M showed significant suppression of their NK activity. Inhibition was proportional to the concentration of the drug, and was observed at as early as 1 hr of incubation at various effector to target cell ratios with several targets. PRD also inhibited NK and ADCC activities of purified T cells, non-T cells, and NK-enriched effector cells. In target-binding assays, PRD decreased the target-binding capacity of effector lymphocytes in a dose-dependent manner. PRD-induced inhibition could be reversed by incubating lymphocytes for 1 hr with interferon or IL 2. Pretreatment of targets with PRD for 4 hr did not affect cytotoxic activity. Inhibition of cytotoxicity was not due to direct toxicity to effector cells because lymphocytes treated with PRD showed normal spontaneous 51Cr release, and their viability after 24 hr of pre-culture with PRD was comparable to that of untreated control cells. These results demonstrate that PRD has significant immunomodulatory effects on human NK and ADCC activities that may be of clinical relevance.
Glucocorticoid hormones are effective in controlling inflammation, but the mechanisms that confer this action are largely unknown. Recent advances in this field have shown that both positive and negative regulation of gene expression are necessary for this process. The genes whose activity are modulated in the anti-inflammatory process code for several cytokines, adhesion molecules and enzymes. Most of them do not carry a classical binding site for regulation by a glucocorticoid receptor, but have instead regulatory sequences for transcription factors such as AP-1 or NF-kappa B. This makes them unusual targets for glucocorticoid action and emphasizes the need for novel regulatory mechanisms. Recent studies describe an important contribution by protein-protein interactions, in which several domains of the receptor participate; these studies provide a better understanding of the action of the receptor and offer opportunities for the design of steroidal compounds that could function more effectively as anti-inflammatory drugs.
Natural killer cells are likely to play an important role in the host defenses because they kill virally infected or tumor cells but spare normal self-cells. The molecular mechanism that explains why NK cells do not kill indiscriminately has recently been elucidated. It is due to several specialized receptors that recognize major histocompatibility complex (MHC) class I molecules expressed on normal cells. The lack of expression of one or more class I alleles leads to NK-mediated target cell lysis. During NK cell development, the class I-specific receptors have adapted to self-class I molecules on which they recognize epitopes shared by groups of class I alleles. As such, they may fail to recognize either self-molecules that bound unusual peptides or allogeneic class I molecules unrelated to self-alleles. Different types of receptors specific for groups of HLA-C or HLA-B alleles have been identified. While in most instances, they function as inhibiting receptors, an activating form of the HLA-C-specific receptors has been identified in some donors. Molecular cloning of HLA-C- and HLA-B-specific receptors has revealed new members of the immunoglobulin superfamily with two or three Ig-like domains, respectively, in their extracellular portion. While the inhibiting form is characterized by a long cytoplasmic tail associated with a nonpolar transmembrane portion, the activating one has a short tail associated with a Lys-containing transmembrane portion. Thus, these human NK receptors are different from the murine Ly49 that is a type II transmembrane protein characterized by a C type lectin domain. A subset of cytolytic T lymphocytes expresses NK-type class I-specific receptors. These receptors exert an inhibiting activity on T cell receptor-mediated functions and offer a valuable model to analyze the regulatory mechanisms involved in receptor-mediated cell activation and inactivation.
1. Glucocorticoids are widely used for the suppression of inflammation in chronic inflammatory diseases such as asthma, rheumatoid arthritis, inflammatory bowel disease and autoimmune diseases, all of which are associated with increased expression of inflammatory genes. The molecular mechanisms involved in this anti-inflammatory action of glucocorticoids is discussed, particularly in asthma, which accounts for the highest clinical use of these agents. 2. Glucocorticoids bind to glucocorticoid receptors in the cytoplasm which then dimerize and translocate to the nucleus, where they bind to glucocorticoid response elements (GRE) on glucocorticoid-responsive genes, resulting in increased transcription. Glucocorticoids may increase the transcription of genes coding for anti-inflammatory proteins, including lipocortin-1, interleukin-10, interleukin-1 receptor antagonist and neutral endopeptidase, but this is unlikely to account for all of the widespread anti-inflammatory actions of glucocorticoids. 3. The most striking effect of glucocorticoids is to inhibit the expression of multiple inflammatory genes (cytokines, enzymes, receptors and adhesion molecules). This cannot be due to a direct interaction between glucocorticoid receptors and GRE, as these binding sites are absent from the promoter regions of most inflammatory genes. It is more likely to be due to a direct inhibitory interaction between activated glucocorticoid receptors and activated transcription factors, such as nuclear factor-kappa B and activator protein-1, which regulate the inflammatory gene expression. 4. It is increasingly recognized that glucocorticoids change the chromatin structure. Glucocorticoid receptors also interact with CREB-binding protein (CBP), which acts as a co-activator of transcription, binding several other transcription factors that compete for binding sites on this molecule. Increased transcription is associated with uncoiling of DNA wound around histone and this is secondary to acetylation of the histone residues by the enzymic action of CBP. Glucocorticoids may lead to deacetylation of histone, resulting in tighter coiling of DNA and reduced access of transcription factors to their binding sites, thereby suppressing gene expression. 5. Rarely patients with chronic inflammatory diseases fail to respond to glucocorticoids, although endocrine function of steroids is preserved. This may be due to excessive formation of activator protein-1 at the inflammatory site, which consumes activated glucocorticoid receptors so that they are not available for suppressing inflammatory genes. 6. This new understanding of glucocorticoid mechanisms may lead to the development of novel steroids with less risk of side effects (which are due to the endocrine and metabolic actions of steroids). 'Dissociated' steroids which are more active in transrepression (interaction with transcription factors) than transactivation (GRE binding) have now been developed. Some of the transcription factors that are inhibited by glucocorticoid, such as nuclear factor-kappa B, are also targets for novel anti-inflammatory therapies.
Biological responses, including those occurring during immune responses, must be controlled by signals that turn them on, as well as signals that turn them off. In addition, fine-tuning can be achieved through signals that amplify or downmodulate responses. The complexity in signal transduction pathways that regulate these responses is daunting. Even terminal responses, such as degranulation by effector cells in the immune system, do not follow a simple one-way signaling pathway. The required initial protein tyrosine phosphorylation events are rapidly reversed by dephosphorylation, and the depleted calcium stores are replenished. Unless the effector cell undergoes programmed cell death after activation, it returns to a basal state of activation from where it can be activated again. Negative regulation of effector cell function is sometimes very tight, such that cellular activation is prevented altogether. This type of inhibitory control is an essential feature of natural killer (NK) cells. Unlike cytotoxic T cells that kill target cells upon a recognition event mediated by their antigen-specific receptors, NK cells are triggered to kill target cells by interactions between activating NK receptors and ligands that are expressed fairly ubiquitously on target cells. The specificity in target cell recognition is achieved primarily by inhibitory receptors specific for MHC class I molecules.
NKp46 is a novel triggering receptor expressed by all human NK cells that is involved in natural cytotoxicity. In this study we show that the surface density of NKp46 may vary in different NK cells and that a precise correlation exists between the NKp46 phenotype of NK clones and their natural cytotoxicity against HLA-class I-unprotected allogeneic or xenogeneic cells. Thus, NKp46bright clones efficiently lysed human and murine tumor cells while NKp46dull clones were poorly cytolytic against both types of target cells. We also show that the NKp46 phenotype of NK clones correlates with their ability to lyse HLA-class I-unprotected autologous cells. Finally, NKp46 was found to be deeply involved in the natural cytotoxicity mediated by freshly derived NK cells. This was indicated both by the inhibition of cytolysis after monoclonal antibody-mediated masking of NKp46 and by the correlation existing between the natural cytotoxicity of fresh NK cells derived from different donors and their NKp46 phenotype. In conclusion, these studies strongly support the concept that NKp46 plays a central role in the physiological triggering of NK cells and, as a consequence (in concert with killer inhibitory receptors), in the NK-mediated clearance of abnormal cells expressing inadequate amounts of HLA-class I molecules.
Natural cytotoxicity receptors (NKp46, NKp44 and NKp300) play a predominant role in human NK cell triggering during natural cytotoxicity. Human 2B4 also induced NK cell activation in redirected killing assays using anti-2B4 monoclonal antibodies (mAb) and murine targets. Since this effect was confined to a fraction of NK cells, this suggested a functional heterogeneity of 2B4 molecules. Here we show that activation via 2B4 in redirected killing against murine targets is strictly dependent upon the engagement of NKp46 by murine ligand (s) on target cells. Thus, NK cell clones expressing high surface density of NKp46 (NKp46bright) were triggered by anti-2B4 mAb, whereas NKp46dull clones were not although they expressed a comparable surface density of 2B4. mAb-mediated modulation of NKp46 molecules in NKp46bright clones had no effect on the expression of 2B4 while it rendered cells unresponsive to anti-2B4 mAb. Finally, anti-2B4 mAb could induce NK cell triggering in NKp46dull clones provided that suboptimal doses of anti-NKp44 or anti-CD16 mAb were added to the redirected killing assay. These results indicate that differences in responses do not reflect a functional heterogeneity of 2B4 but rather depend on the co-engagement of triggering receptors.
Natural killer (NK) cells can detect whether cells have undergone tumour transformation or viral infection. The discovery of specific inhibitory receptors for major histocompatibility complex class I molecules clarified the basis of this discrimination. However, the receptors responsible for NK-cell triggering in the process of natural cytotoxicity remained elusive until recently. Here, Allessandro Moretta and colleagues describe the identification and characterization of several such receptors.
Natural killer cells can discriminate between normal cells and cells that do not express adequate amounts of major histocompatibility complex (MHC) class I molecules. The discovery, both in mouse and in human, of MHC-specific inhibitory receptors clarified the molecular basis of this important NK cell function. However, the triggering receptors responsible for positive NK cell stimulation remained elusive until recently. Some of these receptors have now been identified in humans, thus shedding some light on the molecular mechanisms involved in NK cell activation during the process of natural cytotoxicity. Three novel, NK-specific, triggering surface molecules (NKp46, NKp30, and NKp44) have been identified. They represent the first members of a novel emerging group of receptors collectively termed natural cytotoxicity receptors (NCR). Monoclonal antibodies (mAbs) to NCR block to differing extents the NK-mediated lysis of various tumors. Moreover, lysis of certain tumors can be virtually abrogated by the simultaneous masking of the three NCRs. There is a coordinated surface expression of the three NCRs, their surface density varying in different individuals and also in the NK cells isolated from a given individual. A direct correlation exists between the surface density of NCR and the ability of NK cells to kill various tumors. NKp46 is the only NCR involved in human NK-mediated killing of murine target cells. Accordingly, a homologue of NKp46 has been detected in mouse. Molecular cloning of NCR revealed novel members of the Ig superfamily displaying a low degree of similarity to each other and to known human molecules. NCRs are coupled to different signal transducing adaptor proteins, including CD3 zeta, Fc epsilon RI gamma, and KARAP/DAP12. Another triggering NK receptor is NKG2D. It appears to play either a complementary or a synergistic role with NCRs. Thus, the triggering of NK cells in the process of tumor cell lysis may often depend on the concerted action of NCR and NKG2D. In some instances, however, it may uniquely depend upon the activity of NCR or NKG2D only. Strict NKG2D-dependency can be appreciated using clones that, in spite of their NCR(dull) phenotype, efficiently lyse certain epithelial tumors or leukemic cell lines. Other triggering surface molecules including 2B4 and the novel NKp80 appear to function as coreceptors rather than as true receptors. Indeed, they can induce natural cytotoxicity only when co-engaged with a triggering receptor. While an altered expression or function of NCR or NKG2D is being explored as a possible cause of immunological disorders, 2B4 dysfunction has already been associated with a severe form of immunodeficiency. Indeed, in patients with the X-linked lymphoproliferative disease, the inability to control Epstein-Barr virus infections may be consequent to a major dysfunction of 2B4 that exerts inhibitory instead of activating functions.
The ability of NK cells to kill a wide range of tumor or virally infected target cells as well as normal allogeneic T cell blasts appears to depend upon the concerted action of multiple triggering NK receptors. In this study, using two specific monoclonal antibodies [(mAb) MA152 and LAP171], we identified a triggering NK receptor expressed at the cell surface as a dimer of approximately 80 kDa (NKp80). NKp80 is expressed by virtually all fresh or activated NK cells and by a minor subset of T cells characterized by the CD56 surface antigen. NKp80 surface expression was also detected in all CD3- and in 6 / 10 CD3+ large granular lymphocyte expansions derived from patients with lymphoproliferative disease of granular lymphocytes. In polyclonal NK cells, mAb-mediated cross-linking of NKp80 resulted in induction of cytolytic activity and Ca2+ mobilization. A marked heterogeneity existed in the magnitude of the cytolytic responses of different NK cell clones to anti-NKp80 mAb. This heterogeneity correlated with the surface density of NKp46 molecules expressed by different NK clones. The mAb-mediated masking of NKp80 led to a partial inhibition of the NK-mediated lysis of appropriate allogeneic phytohemagglutinin-induced T cell blasts, while it had no effect on the lysis of different tumor target cells, including T cell leukemia cells. These data suggest that NKp80 recognizes a ligand on normal T cells that may be down-regulated during tumor transformation. Molecular cloning of the cDNA coding for NKp80 revealed a type II transmembrane molecule of 231 amino acids identical to the putative protein encoded by a recently identified cDNA termed KLRF1.
It is not known to what extent glucocorticoid hormones cause their anti-inflammatory actions and their undesirable side effects by the same or different molecular mechanisms. Glucocorticoids combine with a cytoplasmic receptor that alters gene expression in two ways. One way is dependent on the receptor's binding directly to DNA and acting (positively or negatively) as a transcription factor. The other is dependent on its binding to and interfering with other transcription factors. Both mechanisms could underlie suppression of inflammation. The liganded receptor binds and inhibits the inflammatory transcription factors activator protein-1 and NF-kappaB. It also directly induces anti-inflammatory genes such as that encoding the protein inhibitor of NF-kappaB. Recent work has shown that glucocorticoids inhibit signalling in the mitogen-activated protein kinase pathways that mediate the expression of inflammatory genes. This inhibition is dependent on de novo gene expression. It is important to establish the significance of these different mechanisms for the various physiological effects of glucocorticoids, because it may be possible to produce steroid-related drugs that selectively target the inflammatory process.
Perforin/granzyme-induced apoptosis is the main pathway used by cytotoxic lymphocytes to eliminate virus-infected or transformed cells. Studies in gene-disrupted mice indicate that perforin is vital for cytotoxic effector function; it has an indispensable, but undefined, role in granzyme-mediated apoptosis. Despite its vital importance, the molecular and cellular functions of perforin and the basis of perforin and granzyme synergy remain poorly understood. The purpose of this review is to evaluate critically recent findings on cytotoxic granule-mediated cell death and to assess the functional significance of postulated cell-death pathways in appropriate pathophysiological contexts, including virus infection and susceptibility to experimental or spontaneous tumorigenesis.
Natural killer (NK) cells and dendritic cells (DCs) are two types of specialized cell of the innate immune system, the reciprocal interaction of which results in a potent, activating cross-talk. For example, DCs can prime resting NK cells, which, in turn, after activation, might induce DC maturation. However, NK cells negatively regulate the function of DCs also by killing immature DCs in peripheral tissues. Moreover, a subset of NK cells, after migration to secondary lymphoid tissues, might have a role in the editing of mature DCs based on the selective killing of mature DCs that do not express optimal surface densities of MHC class I molecules. So, cognate interactions between NK cells and DCs provide a coordinated mechanism that is involved not only in the regulation of innate immunity, but also in the promotion of appropriate downstream adaptive responses for defence against pathogens.
Signals leading to NK cell triggering are primarily mediated by natural cytotoxicity receptors (NCR) upon binding to as-yet-undefined cell surface ligand(s) on normal hematopoietic cells, pathogen-infected cells or tumor cells. In this study we tried to determine whether the decreased NK cell cytolytic function that is observed in HIV-1-infected patients may be related to a decreased expression of NCR. In HIV-1-infected patients, freshly drawn, purified NK cells expressed significantly decreased surface densities of NKp46 and NKp30 NCR. The low surface density of NKp46, NKp30 and NKp44 was also confirmed in in-vitro-activated NK cell populations and NK cell clones derived from HIV-1 patients compared with uninfected donors. This defective NCR expression in HIV-1 patients was associated with a parallel decrease of NCR-mediated killing of different tumor target cells. Thus, the present study indicates that the defective expression of NCR represents at least one of the possible mechanisms leading to the impaired NK cell function in HIV-1 infection and it can contribute to explain the relatively high frequency of opportunistic tumors reported in cohorts of untreated patients before the occurrence of profound immunosuppression (<200 CD4(+) cells/mm(3)).
Patients undergoing allogeneic bone marrow transplantation offer a unique system to analyze NK cell development in vivo. We analyzed NK cells from 23 such patients to assess the acquisition of activating receptors. Four patients displayed an immature NK cell surface phenotype at engraftment, as their cells were CD16(-)KIR(-) and NKG2D(-) but expressed low levels of NKp46, NKp30, 2B4 and NKG2A. These NK cells had particularly low cytolytic activity against the HLA-class-I(-) melanoma F01 cell line and the 721-221 EBV-infected B cell line. Moreover, cytoxicity was inhibited upon mAb-mediated crosslinking of 2B4. Analysis of NK cells at day 30 after bone marrow transplantation revealed the occurrence of both phenotypic and functional maturation. These data are in agreement with a previous in vitro study showing that immature NK cell precursors express CD16, NKG2D and KIR only at a late stage of differentiation and also express inhibitory 2B4. Our present study allows a better understanding of the NK cell differentiation in vivo.
34: 3028–3038 Corticosteroid-induced inhibitory effect on activating NK receptors NKp44, a novel triggering surface molecule specifically ex-pressed by activated natural killer cells is involved in non-MHC restricted tumor cell lysis
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- A Moretta
Eur. J. Immunol. 2004. 34: 3028–3038 Corticosteroid-induced inhibitory effect on activating NK receptors R., Marcenaro, E., Augugliaro, R., Moretta, L. and Moretta, A., NKp44, a novel triggering surface molecule specifically ex-pressed by activated natural killer cells is involved in non-MHC restricted tumor cell lysis. J. Exp. Med. 1998. 187: 2065–2072.
Glucocorticoids inhibit MAP kinase via increased expression and decreased degradation of MKP-1
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Kassel, O., Sancono, A., Kratzschmar, J., Kreft, B., Stassen,
M. and Cato, A. C. B., Glucocorticoids inhibit MAP kinase via
increased expression and decreased degradation of MKP-1.
34: 3028-3038 Corticosteroid-induced inhibitory effect on activating NK receptors 20 Anti-inflammatory action of glucocorticoids: molecular mechanisms
- J Eur
- P J Barnes
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molecular mechanisms. Clin. Sci 1998. 94: 557-572.
Molecular mechanisms of antiinflammatory action of glucocorticoids
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