During the past few years, a number of studies reported that different melanoma cell lines could be extensively lysed in vitro by IL-2-activated NK cells at appropriate Effector/Target ratios. Here we show, by histological evaluation of different melanoma lesions, that NK/target cell ratios compatible with those allowing efficient melanoma cell killing in vitro are hardly reached at the tumor site. We then investigated the outcome of co-cultures established at low NK/melanoma cell ratios. After initial NK-mediated lysis, residual melanoma cells acquired resistance to IL-2-activated NK cells. This reflected primarily an increased expression, on melanoma cells, of classical and non-classical HLA-class I molecules, accompanied by a partial down-regulation of NKG2D-ligands, and was dependent on NK-mediated IFN-γ release. Consistently, melanoma lesions showed a higher HLA-class I expression on tumor cells that were proximal to infiltrating NK cells. In long-term co-cultures, the "protective phenotype" acquired by melanoma cells was lost over time. However this phenomenon was counteracted by down-regulation of relevant activating receptors in co-cultured NK cells. Analysis of different NK-cell-activating cytokines indicated that IL-15 can partially overcome this novel tumor escape mechanism suggesting that IL-15, rather than IL-2, may be eligible for NK cell-based immunotherapy.

Natural killer (NK) cells play a key role in tumor immune surveillance. However, adoptive immunotherapy protocols using NK cells have shown limited clinical efficacy to date, possibly due to tumor escape mechanisms that inhibit NK cell function. In this study, we analyzed the effect of coculturing melanoma cells and NK cells on their phenotype and function. We found that melanoma cells inhibited the expression of major NK receptors that trigger their immune function, including NKp30, NKp44, and NKG2D, with consequent impairment of NK cell-mediated cytolytic activity against various melanoma cell lines. This inhibitory effect was primarily mediated by indoleamine 2,3-dioxygenase (IDO) and prostaglandin E2 (PGE2). Together, our findings suggest that immunosuppressive barriers erected by tumors greatly hamper the antitumor activity of human NK cells, thereby favoring tumor outgrowth and progression. Cancer Res; 72(6); 1407-15. ©2012 AACR.

Eosinophilic fasciitis (EF) is a rare disease characterized by symmetrical thickness and hardening of the skin, especially localized to forearms and thorax, with eosinophilia. Corticosteroids represent the first-line therapy, even if some patients are scarcely responsive and/or may develop important side effects due to long-term treatment. Here, we describe three cases of EF, two of them refractory to previous steroid therapy, successfully treated with D-penicillamine. The present clinical observations together with the updated review of the literature suggest usefulness of D-penicillamine in EF patients, as well as its potential steroid-sparing value.

Most patients with chronic musculoskeletal pain report poor-quality sleep. The impact of chronic pain on sleep can be described as a vicious circle with mutual deleterious influences between pain and sleep-associated symptoms. It is difficult, however, to extract quantitative or consistent and specific sleep variables (eg, total sleep time, slow-wave sleep, sleep stage duration) that characterize the pain-related disruption of sleep. Comorbidity (eg, fatigue; depression; anxiety, sleep, movement, or breathing disorders) often confounds the reading and interpretation of sleep traces. Furthermore, many other methodologic issues complicate our ability to generalize findings (low external validity) to first-line medicine. Because sleep alterations in common musculoskeletal pain are neither specific nor pathognomonic, the aim is to provide a critical overview of the current understanding of pain and sleep interaction, discussing evidence-based and empiric knowledge that should be considered in further research and clinical applications.

Clinicians and investigators need a simple and reliable recording device to diagnose or monitor sleep bruxism (SB). The aim of this study was to compare recordings made with an ambulatory electromyographic telemetry recorder (TEL-EMG) with those made with standard sleep laboratory polysomnography with synchronised audio-visual recording (PSG-AV). Eight volunteer subjects without current history of tooth grinding spent one night in a sleep laboratory. Simultaneous bilateral masseter EMG recordings were made with a TEL-EMG and standard PSG. All types of oromotor activity and rhythmic masseter muscle activity (RMMA), typical of SB, were independently scored by two individuals. Correlation and intra-class coefficient (ICC) were estimated for scores on each system. The TEL-EMG was highly sensitive to detect RMMA (0·988), but with low positive predictive value (0·231) because of a high rate of oromotor activity detection (e.g. swallowing and scratching). Almost 72% of false-positive oromotor activity scored with the TEL-EMG occurred during the transient wake period of sleep. A non-significant correlation between recording systems was found (r = 0·49). Because of the high frequency of wake periods during sleep, ICC was low (0·47), and the removal of the influence of wake periods improved the detection reliability of the TEL-EMG (ICC = 0·88). The TEL-EMG is sensitive to detect RMMA in normal subjects. However, it obtained a high rate of false-positive detections because of the presence of frequent oromotor activities and transient wake periods of sleep. New algorithms are needed to improve the validity of TEL-EMG recordings.

Human leukocyte antigen (HLA)-E is a nonclassical major histocompatibility complex (MHC) class I molecule of limited sequence variability that is expressed by most tissues albeit at low levels. HLA-E has been first described as the ligand of CD94/NKG2 receptors expressed mainly by natural killer (NK) cells, thus confining its role to the regulation of NK-cell function. However, recent evidences obtained by our and other groups indicate that HLA-E complexed with peptides can interact with alphabeta T-cell receptor (TCR) expressed on CD8(+) T cells. Although, HLA-E displays a selective preference for nonameric peptides, derived from the leader sequence of various HLA class I alleles, several reports indicate that it can present also "noncanonical" peptides derived from both stress-related and pathogen-associated proteins. Because HLA-E displays binding specificity for innate CD94/NKG2 receptors, as well as all the features of an antigen-presenting molecule, its role in both natural and acquired immune responses has recently been re-evaluated.

Although the role of the tumor microenvironment in the process of cancer progression has been extensively investigated, the contribution of different stromal components to tumor growth and/or evasion from immune surveillance is still only partially defined. In this study we analyzed fibroblasts derived from metastatic melanomas and provide evidence for their strong immunosuppressive activity. In coculture experiments, melanoma-derived fibroblasts sharply interfered with NK cell functions including cytotoxicity and cytokine production. Thus, both the IL-2-induced up-regulation of the surface expression of NKp44, NKp30, and DNAM-1 triggering receptors and the acquisition of cytolytic granules were inhibited in NK cells. This resulted in an impairment of the NK cell-mediated killing of melanoma target cells. Transwell cocultures and the use of specific inhibitors suggested that cell-to-cell contact was required for inducing DNAM-1 modulation. In contrast, modulation of NKp44 and NKp30 was due to PGE(2) released by fibroblasts during coculture. Normal skin fibroblasts could also partially affect NK cell phenotype and function. However, the inhibitory effect of tumor-derived fibroblasts was far stronger and directly correlated with their ability to produce PGE(2) either constitutively or upon induction by NK cells.

Experimental and clinical data suggest that tumours harbour a cell population retaining stem cell characteristics that can drive tumorigenesis. CD133 is considered an important cancer stem cells (CSC)-associated marker. In a large variety of human malignancies, including melanoma, CD133(+) cells have been reported to comprise CSC. In this study, we show that melanoma cell lines are highly heterogeneous for the expression of several stem cell-associated markers including CD133, c-kit/CD117 and p75 neurotrophin receptor/CD271. Since no information is available on the ability of NK cells to recognize and lyse melanoma stem cells, we assessed whether melanoma cell lines, characterized by stem cell-like features, were susceptible to lysis by IL-2-activated NK cells. We show that activated NK cells efficiently kill malignant melanoma cell lines that were enriched in putative CSC by the use of different selection methods (i.e. CD133 expression, radioresistance or the ability to form melanospheres in stem cell-supportive medium). NK cell-mediated recognition and lysis of melanoma cells involved different combinations of activating NK receptors. Since CSC have been reported to be both drug resistant and radioresistant, our present data suggest that NK-based adoptive immunotherapy could represent a novel therapeutic approach to possibly eradicate metastatic melanoma.

Bronchoalveolar lavage (BAL) is a minimally invasive method possibly representing a diagnostic tool in the evaluation of interstitial lung diseases (ILDs) of different causes. We first describe herein the morphologic, histochemical, and immunohistochemical features of previously unreported eosinophilic globular deposits of acellular amorphous material of uncertain nature in a relatively large series of 227 BAL samples obtained from patients with various ILDs. Overall, eosinophilic globules were detected in 18 cases (7.9%), 16 of which were in patients with systemic sclerosis (SSc)-related ILD (16/50 [32%]) and in 2 cases of apparently idiopathic usual interstitial pneumonia. Apart from the possible diagnostic information of this finding, in patients with SSc, the globules were significantly related to BAL neutrophilia or eosinophilia and extensive ILD in high-resolution computed tomography (P <.0001). Differential diagnosis with other types of acellular globular materials observed in BAL samples is also discussed.