In contrast to mice, CD25 deficiency in humans is accompanied by severe immunodeficiency that is characterized by susceptibility to opportunistic pathogens and a normal Treg frequency [9, 14, 15, 21-24]. In addition, IL-2-deficient mice are fully capable of rejecting allografts, whereas CD25-deficient humans are not [24, 49, 50]. Therefore, CD25 may be more important for effector function in humans and more

important for tolerance in mice since only Treg cells constitutively express CD25 in mice. This may explain why blocking CD25 during tumor immuno-therapy has not translated well from mice to humans [51]. Discrepancies between mouse and human immunology selleckchem have been described elsewhere and is not unexpected since the species diverged 65–75 million years ago [52]. Therefore, studies conducted in mice on the role of IL-2 Wnt signaling in T-cell function may not exactly translate to humans, and this study may offer one possible explanation for these differences.

We believe that the discovery of this CD4+CD25INT population is particularly important for therapies that target CD25/IL-2 and that hopefully by studying the response of this population we can better understand the mechanism of these therapies and improve their clinical efficacy. We evaluated the response of the CD4+CD25INTFOXP3− population to IL-2 immunotherapy. Over the course of IL-2 immunotherapy in cancer patients, the percentage

of CD4+ T cells that were CD25INT population decreased, while the CD25NEG increased and Treg populations stayed relatively stable, Erastin order suggesting these populations were differentially affected by the therapy. From these studies, it was clear that the CD25INT population was affected by the IL-2 therapy, however, it is currently not known exactly how the CD25INT population responded to the therapy. One possibility is that the CD25INT cells may have downregulated or shed CD25 [53]. However, we did not see diminution of CD25 on the Treg cells, and we demonstrated that not all of the CD25INT population downregulated expression of CD25 in response to rhIL-2 in vitro and that some even increased CD25 expression. In addition, in vitro stimulation with rhIL-2 also suggested that the CD25INT cells are differentially responsive to rhIL-2, as shown by Ki67 staining, and could therefore be act-ivated to a greater degree than the CD25NEG and Treg populations. Therefore, we believe that the disappearance of the CD25INT population observed in IL-2 cancer patients is most likely a combination of events, including decreased surface expression of CD25 and increased activation, which might have led to AICD and/or egress from the blood to tissue. Nevertheless, it is clear that the CD25INT population is greatly affected by IL-2 immunotherapy and may be integral to the antitumor immune response.

As the common clinical features of XLP are FIM, EBV-associated HLH and lymphoproliferative disorder [2, 3], we completed SH2D1A and XIAP gene sequencing in the patients with one or more of these symptoms in this study. Most XLP patients appear healthy prior to contracting EBV [16]. However, following infection, patients often develop T and B cell lymphoproliferation and secondary HLH [16, 17]. Using gene sequencing, we diagnosed five patients with XLP of the 21 male patients in our study with FIM, EBV-associated HLH or persistent EBV

viremia. The overall clinical phenotypes of the affected persons matched those previously reported. All of the five patients had symptoms of HLH and four tested positive for EBV-DNA. This finding indicated that EBV infection triggers HLH in patients with SH2D1A or XIAP deficiency. Although Patient 2 was EBV-DNA negative, we still consider HLH as triggered

click here by EBV infection based on the elevated atypical lymphocyte counts. Previous study reported that about 13 XLP patients showed hypogammaglobulinemia [18]. In our study, 1 patient with SH2D1A deficiency had lower IgG, IgA and IgM levels, especially IgG. The results indicate that the patient had hypogammaglobulinemia. All four patients evaluated for immunological function showed a low CD4/CD8 ratio, which may be associated with EBV infection. JAK inhibitor In patients with XLP, disease onset is usually NADPH-cytochrome-c2 reductase at 2–5 years of age and is often triggered by EBV infection [16, 19]. Among the five patients in the study, the youngest one was only 1 month old at time of onset. It is different with the western world, maybe due to early encountering of the EBV infection. Although there is no precise epidemiological data of EBV infection, the age of onset is thought to vary widely, with developed countries having

higher ages at primary infection, most likely due to better hygienic conditions and other socioeconomic and demographic factors including household size and population density [20]. The result indicates that patients with SH2D1A or XIAP deficiency can show XLP associated symptoms at a very young age. Prior reports indicate that the prognosis for XLP is poor, with 70% of patients dying before the age of 10 and mortality nearing 96% for those with a history of EBV infection [2, 4, 5]. In our study, three patients had rapid disease progression and died. Only one patient received HSCT and is well. The prognosis observed in our study is therefore similar to previous studies. In summary, we report the clinical and genetic features of five Chinese patients with SH2D1A/XIAP deficiency in this study. For patients with severe EBV-associated HLH, our results indicate the need to consider the possibility of XLP. This work was supported by the National Natural Science Foundation of China (81172877, 81000260) and Shanghai Rising-Star Program (11QA1400700). All authors declare no conflict of interest.

Quantitative PCR was performed in a 20-μl reaction mixture containing 0·2 μl cDNA, 0·5 μm forward and reverse primers and 2× Power SYBR Green PCR Master Mix (Applied Biosystems, Foster city, CA) using an ABI PRISM 7300 real-time cycler (Applied Biosystems). The transcript levels of target genes were normalized to β-actin. The primers used for quantitative PCR are listed in Table 1. Macrophages were lysed using RIPA lysis buffer (Applygen Technologies Inc., Beijing, China). Equal amounts of proteins

www.selleckchem.com/products/torin-1.html were separated on 10% SDS–PAGE gel and subsequently electrotransferred onto PVDF membranes (Millipore, Bedford, MA). The membranes were blocked for 1 hr in Tris-buffered saline (TBS) containing 5% non-fat dried milk and incubated overnight with the primary antibodies at 4°. The membranes were then washed with TBS containing 0·1% Tween-20 (TBST) and Selleckchem ONO-4538 incubated with the appropriate horseradish peroxidase-conjugated secondary antibodies (Zhongshan, Beijing, China) at room temperature for 1 hr. Peroxidase colour

visualization was achieved using an enhanced chemiluminescence detection kit (Pierce Biotechnology, Rockford, IL). Macrophages were cultured in 24-well plates at 37° at a density of 1 × 106 cells/well, and stimulated with TLR ligands. The concentration of cytokines in the culture medium was measured using ELISA kits: those for IL-1β (MLB00B), IL-6 (M6000B), TNF-α (MTA00B) and Gas6 (DY986) were purchased from R&D Systems (Minneapolis, MN); and the kit for ProS (E0735h) was purchased from Wuhan EIAab Science Co. Ltd (Wuhan, China). ELISAs were performed according to the manufacturer’s instructions. Data are presented as mean ± standard error of mean (SEM). These data were analysed using

the Student’s t-test or analysis of variance test. All calculations were performed with spss version 11.0 statistical software package (SPSS, Chicago, IL). Values of P < 0·05 and < 0·01 were considered significant and very significant, respectively. Peritoneal macrophages from 10-week-old C57BL/6 mice were used for Gas6/ProS-TAM expression analysis. Cell purity and viability were higher than 95%, based on immunofluorescence staining for F4/80 (Fig. 1a) and flow cytometry after double staining with PE-conjugated antibodies against F4/80 and FITC-conjugated annexin V (Fig. 1b). Axl and Mer were clearly detected, as well as very weak BCKDHB Tyro3 in wild-type (WT) macrophages, using quantitative PCR (Fig. 1c). In contrast, the mRNA of all three TAM receptors were absent in TAM knock-out (TAM−/−) macrophages. Gas6 and ProS mRNA were expressed in both WT and TAM−/− macrophages, with significantly high levels of ProS compared with Gas6 mRNA. Axl and Mer proteins, but not Tyro3, were detected in the WT cells by Western blotting (Fig. 1d), which is consistent with mRNAs. The TAM proteins were not detected in the TAM−/− macrophages. However, secreted Gas6 and ProS were detected in the culture media of both WT and TAM−/− macrophages.

Only select initial trials used DC derived from CD34+ cells, perhaps a result of a more difficult production process 36. All the various trials are difficult to compare due to a range of differences, but the immunogenicity of mature DC was obvious, and hints for clinical efficacy have been observed. The first vaccine with proven clinical benefit (Dendreon’s Sipuleucel-T, Provenge™) is, however, not based on highly enriched ex vivo isolated or in vitro generated DC, but is a cellular

vaccine prepared by isolating via gradient centrifugation a DC-precursor-enriched fraction from apheresis products, which is exposed for 2 days to a fusion protein consisting of GM-CSF and prostatic acid phosphatase antigen. This results in targeting to the GM-CSF receptor-expressing cells, including DC precursors, which then undergo ACP-196 mw activation/maturation in vitro (leading to CD54 upregulation which serves as a potency marker 37). Dendreon has

recently confirmed in its pivotal phase III study (IMPACT trial, n=512) that its first-generation cellular vaccine product Sipuleucel-T (Provenge™) significantly improved overall survival (by a median of 4.1 months, reducing the risk of death by 22.5% compared to the control group) in asymptomatic or minimally symptomatic metastatic, hormone-refractory prostate cancer even though classical regressions did not occur and time to progression was not prolonged 38. The result of the IMPACT MI-503 manufacturer trial led to the approval by the FDA of the first therapeutic cancer vaccine ever on April 29th, 2010. The positive outcome has already fostered interest in the development of cancer vaccines in general and DC in particular. The Dendreon story is interesting in several aspects. It reflects,

for example, that in the cancer research community until recently the classical acute response criteria developed for chemotherapy were considered indispensable in judging vaccine effectiveness, even though it diglyceride had been pointed out early in the 1990s that vaccines require time but not necessarily classical regressions to produce clinical benefit 39. In 2007, the Cancer Vaccine Clinical Trial Working Group came up with an important position paper proposing a new clinical development paradigm for cancer vaccines 40, and phase II trials employing anti-CTLA-4 antibodies also unraveled distinct response patterns associated with favorable survival 41. The Sipuleucel-T product development – a nerve-wracking roller coaster – also shows that one may succeed with a product that for practical reasons has not been extensively optimized (e.g. to better enrich DC) as long as it can be reproducibly manufactured, a potency marker is available, and one has chosen a tumor amenable to the cancer vaccine.

In addition, CD69 might act specifically on the Treg cell subset, directly suppressing the activity of effector T cells [56]. After MSC/CD4+CD25– co-cultures, we observed that SSc cells were able

to induce normally functioning Tregs from the T lymphocytes of HC and SSc patients. As selleck compound CD69 expression by Tregs has been associated with the production of TGF-β [55], we analysed the surface expression of this molecule in induced Tregs. Interestingly, although the CD69 surface expression was decreased in circulating SSc Tregs, an increased expression of this molecule was observed in induced cells without differences between patients and controls. Consistent with this evidence, this website induced SSc Tregs showed a normal ability to inhibit immunoproliferation of CD4+ T cells. We observed an increase of TGF-β production in the supernatants of SSc–MSC co-cultures, and this

production was associated with an increase of TGF-β gene expression in the SSc–MSCs. During SSc, IL-6 and TGF-β are involved not only in immunoregulatory mechanisms but also in the pathogenesis of the fibrotic process, which is the main feature of the disease. Further experiments are ongoing in our laboratory in order to evaluate the role of these cytokines, produced by MSCs, on collagen production as well as on modulation of the myofibroblast phenotype. These Ponatinib nmr findings might suggest that, during SSc, an adaptive cytokine profile with an increase in both TGF-β and IL-6 expression avoids senescence interfering with MSC activity, thus maintaining their role in inducing fully functional Tregs. In this work we did not investigate the immunosuppressive role of senescent SSc–MSCs on dendritic cell functions, already shown in other conditions. It is well known that these cells produce higher levels of IL-10 and

might contribute to the specific cytokine milieu in the disease [57]. Furthermore, recent reports showed that dendritic cells might express TGF-β and support fibrogenesis [58]. In this setting, the possible modulation of dendritic cells might offer a new future target for MSC therapeutic application. The in-vitro immunosuppressive activity of MSCs is mediated by direct interaction with lymphocytes at a MSC : PBMC ratio of 1:1 [59]. This raises a question: are these MSC : PBMC ratios achieved normally in vivo, when MSC are utilized clinically in the clinical setting? Indeed, according to the immunosuppression observed in vivo [60], relatively high numbers of MSC should be injected to obtain this effect. This may be of great relevance in planning the dose of MSC to administer. However, some difficulties in obtaining a sufficient number of MSCs for clinical purposes have been described previously [61].

Here, we identified allograft inflammatory factor 1 (AIF1, Iba1) and sialic acid binding Ig-like lectin 1 (SIGLEC1) as putative NHD-specific biomarkers by bioinformatics analysis of microarray

data of NHD DC. We studied three NHD and eight control brains by immunohistochemistry with a panel of 16 antibodies, including those against Iba1 and SIGLEC1. We verified the absence of DAP12 expression in NHD brains and the expression of DAP12 immunoreactivity MG-132 in vitro on ramified microglia in control brains. Unexpectedly, TREM2 was not expressed on microglia but expressed on a small subset of intravascular monocytes/macrophages in control and NHD brains. In the cortex of NHD brains, we identified accumulation of numerous Iba1-positive microglia to an extent similar to control brains, while SIGLEC1 was undetectable on microglia in all the brains examined. These observations indicate that human

microglia in brain tissues EPZ 6438 do not express TREM2 and DAP12-deficient microglia are preserved in NHD brains, suggesting that the loss of DAP2/TREM2 function in microglia might not be primarily responsible for the neuropathological phenotype of NHD. “
“Glucose transporter-1 (GLUT-1) is one of the major isoforms of the family of glucose transporter proteins that facilitates the import of glucose in human cells to fuel anaerobic metabolism. The present study was meant to determine the extent of the anaerobic/hypoxic state of the intratumoral microenvironment by staining for GLUT-1 in intracranial non-embolized typical (WHO grade I; n = 40), brain invasive and atypical (each WHO grade II; n = 38) and anaplastic meningiomas (WHO grade III, n = 6). In addition, GLUT-1 staining levels were compared

with the various histological criteria used for diagnosing WHO grade II and III meningiomas, namely, brain invasion, increased mitotic activity and atypical cytoarchitectural change, defined by the presence of at least three out of hypercellularity, sheet-like growth, prominent Y-27632 2HCl nucleoli, small cell change and “spontaneous” necrosis. The level of tumor hypoxia was assessed by converting the extent and intensity of the stainings by multiplication in an immunoreactive score (IRS) and statistically evaluated. The results were as follows. (1) While GLUT-1 expression was found to be mainly weak in WHO grade I meningiomas (IRS = 1–4) and to be consistently strong in WHO grade III meningiomas (IRS = 6–12), in WHO grade II meningiomas GLUT-1 expression was variable (IRS = 1–9). (2) Histologically typical, but brain invasive meningiomas (WHO grade II) showed no or similarly low levels of GLUT-1 expression as observed in WHO grade I meningiomas (IRS = 0–4).

1b). The lungs were washed by cannulating the buy LEE011 trachea and gently injecting/recovering (3×) 1·0 ml of PBS. The bronchoalveolar lavage fluid (BAL) was centrifuged at 300 g at 4°C for 5 min and the supernatants were stored at −20°C for cytokine analysis. The cell pellet was resuspended in 0·1 ml of 3% bovine serum albumin (BSA) and cells counted using a haemocytometer. The cells were then cytocentrifuged and stained with haematoxylin and eosin (H&E) for differential

counting based on cell morphology and staining patterns. The means of three independent counts of 100 cells in a randomized field were shown. Following bronchoalveolar lavage, the lungs were fixed with formalin. Serial sagittal sections of whole lung (3–4 µm Selleckchem Talazoparib thick) were cut and stained with Gomori trichome for light microscopy. At least 10 fields were selected randomly and examined. The severity of the inflammatory process in the lungs was scored by two pathologists who were blinded to group identity. The scale varied from 0 to 5 as follows: 0, no inflammation, 1, minimal; 2, mild; 3,

medium; 4, moderate; and 5, marked [35,36]. The EPO assay was performed as described previously [37]. Briefly, a 100-mg sample of tissue from each lung was homogenized in 1·9 ml of PBS and centrifuged at 12 000 g for 10 min. The supernatant was discarded and the erythrocytes were lysed. The samples were centrifuged, the supernatant discarded and the pellet resuspended in 1·9 ml of 0·5% hexadecyltrimethyl ammonium bromide in PBS saline. The samples were frozen in liquid nitrogen and centrifuged at 4°C at 12 000 g for 10 min. The supernatant was used for the enzymatic assay. Briefly, o-phenylenediamine (OPD) (10 mg) triclocarban was dissolved in 5·5 ml distilled water, and then 1·5 ml of OPD solution was added to 8·5 ml of Tris buffer (pH 8·0), followed by addition of 7·5 µl H2O2. In a 96-well plate, 100 µl of substrate solution was added to 50 µl of each sample. After 30 min, the reaction was stopped with 50 µl of 1 M H2SO4 and the absorbance was read at 492 nm. Levels of IL-4, IL-5,

IL-10, TNF-α and IFN-γ were determined by bronchoalveolar lavage (BAL) of the different groups of mice with an enzyme-linked immunosorbent assay (ELISA) sandwich technique using commercially available kits (OptEIA; BD Bioscience, San Jose, CA, USA), according to the manufacturer’s protocol. The optical density (OD) values were read at 450 nm. The results were expressed as picograms per millilitre, compared to a standard curve. The levels of OVA-specific IgE in serum were determined by ELISA, as described previously [38,39]. Briefly, Maxisorp 96-well microtitre plates (nunc, Roskilde, Denmark) were coated with rat anti-mouse unlabelled IgE (1 : 250; Southern Biotechnology, AL, USA) in pH 9·6 carbonate-bicarbonate buffer for 12–16 h at 4°C and then blocked for 1 h at room temperature with 200 µl/well of 0·25% PBS-casein.

The only other study to examine Tregs within canine tumours found similar results to

the many other Vemurafenib studies of human tumours and experimental cancer models. They reported that the percentage of FoxP3+ CD4+ cells in dogs with malignant melanoma was significantly increased in the blood compared with healthy control dogs, and the percentage of FoxP3+ CD4+ cells within tumours compared to blood was also significantly increased (31). Therefore, this study clearly demonstrates that the developing dogma that FoxP3+ T cells are highly prevalent in tumour-associated inflammation is not universally true and emphasizes that malignant transformation can still occur in the absence of immunosuppressive FoxP3+ T cells. It is in agreement with the canine literature learn more on sarcoma (16), especially osteosarcoma (32). Interestingly, in humans with Ewing’s sarcoma, there was also no infiltration of FoxP3+ cells into the tumours, whereas in patients with metastases, the number of FoxP3+ cells only increased in the bone marrow (33). The fact that a large number of positive cells were observed in a few cases, as well as in lymph nodes, but not in the iso- or tissue controls,

excludes technical error. Moreover, all samples were fixed by the same method (formalin-fixed and paraffin-embedded), and the nine positive controls (lymph nodes) originate from nine of the study cases. Therefore, it seems feasible that there is a real difference in the immune response to sarcomas (especially in dogs), compared to other tumours, especially melanomas. The possible role of Tregs in the pathogenesis of spirocercosis-induced sarcoma is especially intriguing, because of the well-documented role of Tregs in helminth infection. In chronic helminth infection (and spirocercosis-induced inflammation is, indeed, chronic) Tregs reduce the intensity of the infection (8). There

is evidence that the increased Tregs response facilitates long-lasting chronic PAK5 inflammation that reduces auto-immunity and allergy in infected subjects (34). This notion is part of the proposed mechanism of what is known as the ‘hygiene hypothesis’ that describes the association between of helminth infection and low incidence of autoimmunity (35). The Tregs-induced increased ‘self-tolerance’ may reduce anti-tumour immunity, and this could potentially be the link between spirocercosis and tumour formation. It appears, however, that although FoxP3+ cells were circulating in lymphatics around S. lupi nodules, ‘homing’ into the nodules did not take place. The low number of FoxP3+ cells does not entirely preclude their potential role in local or systemic immune inhibition in spirocercosis, but functional assays are required.

TDP-43-immunoreactive inclusions affected more of the cortical profile in longer duration cases; their distribution varied with disease subtype, but was unrelated to Braak tangle score. Different TDP-43-immunoreactive

inclusions were not spatially correlated. Conclusions: Laminar distribution of pathological features in 10 sporadic cases of FTLD-TDP is heterogeneous and may be accounted for, in part, by disease subtype and disease duration. In addition, the feedforward and feedback cortico-cortical connections may be compromised in FTLD-TDP. “
“Angiocentric glioma (AG) is an epileptogenic benign cerebral tumor primarily affecting children and young adults, and characterized histopathologically ubiquitin-Proteasome system by an angiocentric pattern of growth of monomorphous bipolar cells with features of ependymal

differentiation (WHO grade I). We report an unusual cerebral glial tumor in a 66-year-old woman with generalized tonic-clonic seizure; the patient also had a 6-year history of headache. On MRI, the tumor appeared as a large T2-hyperintense lesion involving the right insular gyri-anterior temporal lobe, with post-contrast enhancement in the BMN 673 ic50 insula region. Histopathologically, the tumor involving the insular cortex-subcortical white matter was composed of GFAP-positive glial cells showing two different morphologies: one type had monomorphous bipolar cytoplasm and was angiocentric with circumferential alignment to the blood vessels, with dot-like structures positive for epithelial membrane antigen and a Ki-67 labeling index of <1%, and the other was apparently astrocytic, being diffusely and more widely distributed in the parenchyma, showing mitoses and a Ki-67 labeling index of >5%. In the anterior temporal lobe, a diffuse increase in the number of astrocytic cells was evident in part of the cortex and subcortical white matter. On the basis of these findings, we considered whether the present

Tobramycin tumor may represent an unusual example of AG with infiltrating astrocytic cells showing primary anaplastic features (AG with anaplastic features), or anaplastic astrocytoma showing primary vascular-associated ependymal differentiation (anaplastic astrocytoma with angiocentric ependymal differentiation). At present, the latter appears to be the more appropriate interpretation. “
“Malignant peripheral nerve sheath tumor (MPNST) is an uncommon type of sarcoma that arises from peripheral nerve sheaths and rarely involves the spinal roots. The origin of this tumor is thought to be Schwann cells or pluripotent cells of the neural crest. The subgroup of tumors in which malignant Schwann cells coexist with malignant rhabdomyoblasts is termed malignant triton tumor (MTT). MPNSTs can show different degrees of malignancy, but overall spinal MTTs are high-grade lesions.

In agreement with previous studies, we found higher expression of NKG2C in seropositive donors. However, co-expression of NKG2C

with activating KIR2DS1 and KIR3DS1 was not different in CMV-seropositive or -seronegative donors (data not shown). Collectively, these data show that the resting NK-cell KIR repertoire is not modulated by previous CMV infection. We next assessed how NK-cell subsets respond to in vitro exposure to CMV using a co-culture model using the fibroblast line MRC-5 (which supports CMV replication in vitro and carries all relevant ligands to inhibitory KIRs, that is, HLA groups C1, C2, and AZD9668 chemical structure Bw4) in the presence or absence of CMV. In both CMV-seropositive and CMV-seronegative donors, the frequency of NK cells

within the PBMC population increased during CMV co-culture (day 0: 8 and 6%, day 21: 17 and 20%, respectively, for seropositive and seronegative donors). Compared with noninfected MRC-5, co-culture with CMV-infected MRC-5 induced specific changes in the KIR repertoire (Fig. 1). KIR repertoire changes on the total NK-cell population were exclusively detected in CMV-seropositive Regorafenib donors. The frequency of NK cells expressing the inhibitory receptors KIR2DL1, KIR2DL2/3, and natural killer cell group antigen 2A (NKG2A) increased significantly in PBMCs co-cultured with CMV-infected MRC-5 cells (Fig. 1A, B, and D), if NK cells were derived from a donor carrying anti-CMV-IgG antibodies. No expansion of KIR3DL1 was observed (Fig. 1C). Strikingly, no expansion of KIR2DL1 and KIR2DL2/3 expressing NK cells occurred in CMV-seronegative

donors upon co-culture on CMV-infected MRC-5. Of the activating receptors studied, we found no significant change in the expression of KIR2DS1 (Fig. 1E), whereas the frequency of KIR3DS1-expressing NK 3-mercaptopyruvate sulfurtransferase cells increased significantly after co-culture with CMV-infected MRC-5 (Fig. 1F). This was exclusively observed if the donor had previously undergone CMV infection. Importantly, both in CMV-seropositive and CMV-seronegative donors, NK cells were polyclonal after co-culture, as evidenced by a variegated pattern of KIR and NKG2A expression. In CMV-seronegative donors, the only alteration induced by CMV infection was an increase in the expression of NKG2A by day 21. As NKG2C expression has previously been shown to be up-regulated in patients during and after CMV replication [13, 15, 16], we assessed total NKG2C expression and KIR expression on NKG2C+ cells before and after 14-day culture, as a more sensitive assay directly investigating putative CMV-specific NK cells. NKG2C expression was nonsignificantly elevated in CMV-seropositive donors compared with that in seronegative donors at baseline.