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Anti-PD-1 antibodies revolutionized the treatment of advanced melanoma patients. However, one out of three do not respond to this therapy, with an overall poor prognosis. Identification of predictive biomarkers in patients receiving immune-based therapies is necessary for minimizing risk of toxicity and optimizing patient benefit and is still an important unmet clinical need. Recently, many studies have evaluated peripheral blood markers as potential biomarkers, but none so far have been validated. We collected at baseline peripheral blood samples from 18 consecutive advanced melanoma patients treated with anti-PD-1 therapy. Main pro- and anti-inflammatory cytokines were studied in PBMCs from baseline blood samples both evaluating mRNA expression by qRT-PCR and identifying PBMCs subpopulations by FACS analysis. We found that IFN-γ mRNA expression levels were significantly higher in responder patients compared to non-responder ones. Moreover, to better validate its role, we evaluated the IFN-γ/IL-10 ratio. This value was higher in responder patients. FACS analysis confirmed that CD4 + IFN-γ + PBMCs percentage was higher in responders. Our data suggest an interesting correlation between IFN-γ/IL-10 ratio and response to anti-PD-1 therapy in advanced melanoma patients, suggesting a new biomarker that could be easily incorporated in clinical practice.

Immunotherapy takes advantage of the immune system to prevent, control, and eliminate neoplastic cells. The research in the field has already led to major breakthroughs to treat cancer. In this work, we describe a platform that integrates in vitro bioassays to test the immune response and direct antitumor effects for the preclinical discovery of anticancer candidates. The platform relies on the use of dendritic cells that are professional antigen-presenting cells (APC) able to activate T cells and trigger a primary adaptive immune response. The experimental procedure is based on two phenotypic assays for the selection of chemical leads by both a panel of nine tumor cell lines and growth factor-dependent immature mouse dendritic cells (D1). The positive hits are then validated by a secondary test on human monocyte-derived dendritic cells (MoDCs). The aim of this approach is the selection of potential immunotherapeutic small molecules from natural extracts or chemical libraries.

We recently demonstrated that human T helper (Th) 17 cells, unlike Th1 cells, do not proliferate in response to T-cell receptor stimulation, mainly because of their reduced capacity to produce and respond to IL-2. In this study we show that their lower responsiveness to IL-2 is due to the selective expression of Musculin (MSC), a member of the basic helix-loop-helix transcription factors. We show that MSC expression in human Th17 cells is retinoic acid orphan receptor (ROR)γt-dependent, and allows the up-regulation of PPP2R2B, a regulatory member of the protein phosphatase 2A (PP2A) enzyme. High PPP2R2B levels in human Th17 cells were responsible for the reduced STAT5B Ser-193 phosphorylation upon IL-2 signalling and, therefore, impaired STAT5B DNA binding and transcriptional activity on IL-2 target genes. PP2A, observed in Th17 cells, controls also STAT3, dephosphorylating Ser727, thus increasing its activity which plays a crucial role in Th17 development and/or maintenance. Thus, our findings identify an additional mechanism responsible for the limited expansion of human Th17 cells, and could provide a further explanation for the rarity of these cells in inflamed tissues. This article is protected by copyright. All rights reserved

Background: Systemic sclerosis (SSc) T cells can induce apoptosis of autologous skin fibroblasts in vitro. Th17 cells have been reported to increase in SSc patients, and interleukin-17A (IL-17A) has a profibrotic function. We used a system based on T-cell-autologous fibroblast co-cultures to further investigate a possible role of IL-17A in SSc. Methods: T cells from diffuse SSc patients were co-cultured with autologous skin fibroblasts. IL17A mRNA was assessed by real-time PCR in co-cultured and control T cells, while IL17RA, CXCL1, CCL2, CCL3, COL1A1, COL3A1, CTGF, TGFBR2, and SMAD3 mRNAs were assessed in co-cultured and control fibroblasts. In subset experiments, co-cultures and control cells were treated with either IL-17A or IL-17A plus anti-IL17 receptor monoclonal antibody (α-IL-17RA mAb). Chemokine and procollagen type I (PCI) production was further investigated at the protein level in cell culture supernatants by multiple suspension immunoassay and sandwich ELISA, respectively. Co-cultured and control fibroblasts were also stained with Annexin V and analyzed by flow cytometry. Results: T cell-fibroblast co-cultures overexpressed IL17A and IL17RA. Furthermore, co-cultured fibroblasts upregulated IL-17A targets CXCL1, CCL2, and CCL3, while COL1A1, COL3A1, CTGF, and two key effectors of the TGF-β signaling, TGFBR2 and SMAD3, were found downregulated. Consistently, chemokine concentrations were increased in co-culture supernatants, while PCI levels were reduced, especially after stimulation with ectopic IL-17A. Finally, simultaneous α-IL-17RA mAb treatment restored PCI levels and reduced fibroblast apoptosis in IL-17A-stimulated co-cultures. Conclusion: These data suggest that IL-17A upregulation might play a role in modulating T cell-mediated antifibrotic and proapoptotic effects in co-cultured autologous skin fibroblasts.

Aims T-lymphocytes plays an important role in the pathophysiology of acute coronary syndromes. T-cell activation in vitro by pro-inflammatory cytokines may lead to functional tissue factor (TF) expression, indicating a possible contribution of immunity to thrombosis. Oxidized low-density lipoproteins (oxLDLs) are found abundantly in atherosclerotic plaques. We aimed at evaluating the effects of oxLDLs on TF expression in T cells and the role of the lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1). Methods and results CD3+ cells were isolated from healthy volunteers. Gene, protein, and surface expression of TF, as well as of LOX-1, were assessed at different time-points after oxLDL stimulation. To determine whether oxLDL-induced TF was LOX-1 dependent, T cells were pre-incubated with an LOX-1 inhibiting peptide (L-RBP) or with an anti-LOX-1 blocking antibody. To exclude that TF expression was mediated by reactive oxygen species (ROS) generation, oxLDL-stimulated T cells were pre-incubated with superoxide dismutase + catalase or with 4-Hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl (Tempol), an intracellular free radical scavenger. Finally, to determine if the observed findings in vitro may have a biological relevance, the presence of CD3+/TF+/LOX-1+ cells was evaluated by immunofluorescence in human carotid atherosclerotic lesions. oxLDLs induced functionally active TF expression in T cells in a dose- and time-dependent manner, independently on ROS generation. No effect was observed in native LDL-treated T cells. LOX-1 expression was also induced by oxLDLs in a time- and dose-dependent manner. Pre-incubation with L-RBP or anti-LOX-1 antibody almost completely inhibited oxLDL-mediated TF expression. Interestingly, human carotid plaques showed significant infiltration of CD3+ cells (mainly CD8+ cells), some of which were positive for both TF and LOX-1. Conclusion oxLDLs induce functional TF expression in T-lymphocytes in vitro via interaction of oxLDLs with LOX-1. Human carotid atherosclerotic plaques contain CD3+/CD8+cells that express both TF and LOX-1, indicating that also in patients these mechanisms may play an important role.

Tissue-resident memory (Trm) cells are specialized components of both CD4+ and CD8+ T cell subsets that persist in peripheral nonlymphoid tissues following infections and provide fast response in case of a secondary invasion by the same pathogen. Trm cells express the surface markers CD69, CD103, and the immune checkpoint molecule PD-1. Trm cells develop not only in the context of infections but also in tumors, where they can provide a line of defense as suggested by the positive correlation between the frequency of tumor-infiltrating Trm cells and patients' survival. Trm cells persistence in peripheral tissues depends on their adaptation to the local microenvironment and the presence of survival factors, mainly IL-7, IL-15, and Notch ligands. However, the cell sources of these factors are largely unknown, especially in the context of tumors. Here, we show that head-neck squamous cell carcinoma (HNSCC) is enriched in CD4+ and CD8+ T cells with a Trm phenotype. Moreover, we show that mesenchymal stromal cells that accumulate in HNSCC are a source of survival factors and allow proper expression of Trm-typical markers in a VCAM1-dependent manner.

// Giuseppina Conteduca 1 , Daniela Fenoglio 1,2,3 , Alessia Parodi 1 , Florinda Battaglia 1 , Francesca Kalli 1 , Simone Negrini 1 , Samuele Tardito 1 , Francesca Ferrera 1 , Annalisa Salis 1 , Enrico Millo 1 , Giuseppe Pasquale 4 , Giusi Barra 4 , Gianluca Damonte 1 , Francesco Indiveri 1,2 , Soldano Ferrone 5,* and Gilberto Filaci 1,2,3,* 1 Centre of Excellence for Biomedical Research, University of Genoa, Genoa, Italy 2 Department of Internal Medicine, University of Genoa, Genoa, Italy 3 IRCCS AOU San Martino – IST, Genoa, Italy 4 Department of Clinical and Experimental Medicine, Second University of Naples, Naples, Italy 5 Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA * These authors have contributed equally to the manuscript Correspondence to: Gilberto Filaci, email: // Keywords : AIRE, MAGE, medullary thymic epithelial cells, single nucleotide polymorphism, tolerance, Immunology and Microbiology Section, Immune response, Immunity Received : January 27, 2016 Accepted : August 08, 2016 Published : August 22, 2016 Abstract AIRE is involved in susceptibility to melanoma perhaps regulating T cell immunity against melanoma antigens (MA). To address this issue, AIRE and MAGEB2 expressions were measured by real time PCR in medullary thymic epithelial cells (mTECs) from two strains of C57BL/6 mice bearing either T or C allelic variant of the rs1800522 AIRE SNP. Moreover, the extent of apoptosis induced by mTECs in MAGEB2-specific T cells and the susceptibility to in vivo melanoma B16F10 cell challenge were compared in the two mouse strains. The C allelic variant, protective in humans against melanoma, induced lower AIRE and MAGEB2 expression in C57BL/6 mouse mTECs than the T allele. Moreover, mTECs expressing the C allelic variant induced lower extent of apoptosis in MAGEB2-specific syngeneic T cells than mTECs bearing the T allelic variant ( p < 0.05). Vaccination against MAGEB2 induced higher frequency of MAGEB2-specific CTL and exerted higher protective effect against melanoma development in mice bearing the CC AIRE genotype than in those bearing the TT one ( p < 0.05). These findings show that allelic variants of one AIRE SNP may differentially shape the MA-specific T cell repertoire potentially influencing susceptibility to melanoma.

Background Anti-PD-1/PD-L1 drugs are effective as monotherapy in a proportion of NSCLC patients and there is a strong rationale for combining them with targeted therapy. Inhibition of MAPK pathway may have pleiotropic effects on the microenvironment. This work investigates the efficacy of combining MEK and PD-L1 inhibition in pre-clinical and ex-vivo NSCLC models. Methods We studied the effects of MEK inhibitors (MEK-I) on PD-L1 and MCH-I protein expression and cytokine production in vitro in NSCLC cell lines and in PBMCs from healthy donors and NSCLC patients, the efficacy of combining MEK-I with anti-PD-L1 antibody in ex-vivo human spheroid cultures obtained from fresh biopsies from NSCLC patients in terms of cell growth arrest, cytokine production and T-cell activation by flow cytometry. Results MEK-I modulates in–vitro the immune micro-environment through a transcriptionally decrease of PD-L1 expression, enhance of MHC-I expression on tumor cells, increase of the production of several cytokines, like IFNγ, IL-6, IL-1β and TNFα. These effects trigger a more permissive anti-tumor immune reaction, recruiting immune cells to the tumor sites. We confirmed these data on ex-vivo human spheroids, showing a synergism of MEK and PD-L1 inhibition as result of both direct cancer cell toxicity of MEK-I and its immune-stimulatory effect on cytokine secretion profile of cancer cells and PBMCs with the induction of the ones that sustain an immune-reactive and inflammatory micro-environment. Conclusions Our work shows the biological rationale for combining immunotherapy with MEK-I in a reproducible ex-vivo 3D-culture model, useful to predict sensitivity of patients to such therapies. Electronic supplementary material The online version of this article (10.1186/s13046-019-1257-1) contains supplementary material, which is available to authorized users.

Colorectal cancer; Epidermal growth factor receptor (EGFR); MAPK pathway Càncer colorectal; Receptor epidèrmic de factor del creixement (EGFR); Via MAPK Cáncer colorrectal; Receptor epidérmico de factor del crecimiento (EGFR); Vía MAPK BACKGROUND: Previous studies showed that the combination of an anti-Epidermal growth factor (EGFR) and a MEK-inhibitor is able to prevent the onset of resistance to anti-EGFR monoclonal antibodies in KRAS-wild type colorectal cancer (CRC), while the same combination reverts anti-EGFR primary resistance in KRAS mutated CRC cell lines. However, rapid onset of resistance is a limit to combination therapies in KRAS mutated CRC. METHODS: We generated four different KRAS mutated CRC cell lines resistant to a combination of cetuximab (an anti-EGFR antibody) and refametinib (a selective MEK-inhibitor) after continuous exposure to increasing concentration of the drugs. We characterized these resistant cell lines by evaluating the expression and activation status of a panel of receptor tyrosine kinases (RTKs) and intracellular transducers by immunoblot and qRT-PCR. Oncomine comprehensive assay and microarray analysis were carried out to investigate new acquired mutations or transcriptomic adaptation, respectively, in the resistant cell lines. Immunofluorescence assay was used to show the localization of RTKs in resistant and parental clones. RESULTS: We found that PI3K-AKT pathway activation acts as an escape mechanism in cell lines with acquired resistance to combined inhibition of EGFR and MEK. AKT pathway activation is coupled to the activation of multiple RTKs such as HER2, HER3 and IGF1R, though its pharmacological inhibition is not sufficient to revert the resistant phenotype. PI3K pathway activation is mediated by autocrine loops and by heterodimerization of multiple receptors. CONCLUSIONS: PI3K activation plays a central role in the acquired resistance to the combination of anti-EGFR and MEK-inhibitor in KRAS mutated colorectal cancer cell lines. PI3K activation is cooperatively achieved through the activation of multiple RTKs such as HER2, HER3 and IGF1R.

Background Previous studies have demonstrated that IgG1 mAbs as cetuximab, stimulate Antibody-Dependent Cell Cytotoxicity (ADCC). Among immune checkpoint inhibitors, avelumab is the only fully human IgG1 anti-PD-L1 mAb with ADCC properties. Anti-PD-L1 and anti-EGFR mediated NK cytotoxicity is evaluated. Methods LDH release was analyzed to study NK-mediated cytotoxicity by LDH Cytotoxicity Assay Kit and results was correlated to the level of PD-L1, EGFR and MHC-I cell surface expression analyzed by flow cytometry. NK-mediated cytotoxicity of the combination of anti-PD-L1 and anti-EGFR mAbs was studied in a panel of NSCLC cells lines encompassing different tumor types, using as effector NK cells isolated from healthy donors or NSCLC patients. Results PD-L1/EGFR/MHC-I expression levels correlated with enhanced ADCC lysis by the combination of avelumab and cetuximab as demonstrated by LDH assay, CD16 and CD107a mRNA. No significant difference in avelumab plus cetuximab-mediated ADCC between NK cells from healthy donors or from NSCLC patients was observed with a trend in favor of cancer patients, indicating that NK from cancer patients maintain lytic activity. ADCC capability of NK cells isolated from patients enrolled in the phase II study CAVE (Cetuximab-AVElumab)-lung, a single arm phase II clinical study of the combination of avelumab plus cetuximab in the second line treatment of metastatic non small cell lung cancer (NSCLC) patients (EUDRACT 2017- 004195-58) study resulted significantly enhanced after the experimental treatment compared to untreated baseline and healthy donors samples. Conclusions The combination of anti-EGFR and anti PD-L1 IgG1 antibodies is synergistic in terms of ADCC, where each antibody complements each other by promoting a more permissive immune reaction against the tumor, active also in otherwise immune-resistant cancers. Legal entity responsible for the study Universita degli studi della Campania Luigi Vanvitelli. Funding Merck KGaA. Disclosure F. Ciardiello: Advisory / Consultancy, Research grant / Funding (institution): Roche; Advisory / Consultancy, Research grant / Funding (institution): Bayer; Advisory / Consultancy, Research grant / Funding (institution): Amgen; Advisory / Consultancy: Merck; Advisory / Consultancy: Pfizer; Advisory / Consultancy: Sanofi; Advisory / Consultancy: Servier; Advisory / Consultancy: BMS; Advisory / Consultancy: Cellgene; Advisory / Consultancy: Lilly; Research grant / Funding (institution): AstraZeneca; Research grant / Funding (institution): Ipsen. F. Morgillo: Advisory / Consultancy: MSD; Advisory / Consultancy: Lilly; Research grant / Funding (institution): AstraZeneca. All other authors have declared no conflicts of interest.

Sphingosine 1-phosphate (S1P) has a role in many cellular processes. S1P is involved in cell growth and apoptosis, regulation of cell trafficking, production of cytokines and chemokines. The kinases SphK1 and SphK2 (SphKs) phosphorilate Sphingosine (Sph) to S1P and several phosphatases revert S1P to sphingosine, thus assuring a balanced pool that can be depleted by a Sphingosine lyase in hexadecenal compounds and aldehydes. There are evidences that SphK1 and 2 may per se control cellular processes. Here, we report that Sph kinases regulate IL-17 expression in human T cells. SphKs inhibition impairs the production of IL-17, while their overexpression up-regulates expression of the cytokine through acetylation of IL-17 promoter. SphKs were up-regulated also in PBMCs of patients affected by IL-17 related diseases. Thus, S1P/S1P kinases axis is a mechanism likely to promote IL-17 expression in human T cells, representing a possible therapeutic target in human inflammatory diseases.

Background Molecular mechanisms driving acquired resistance to anti-EGFR therapies in metastatic Colorectal Cancer (mCRC) are complex but generally involve the activation of the downstream RAS-RAF-MEK-MAPK pathway. Methods In order to understand the mechanism underlying MEK inhibitor (MEKi) resistance, we generated three different MEKi resistant models, two all RAS WT (SW48-MR and LIM1215-MR) and one mutated in RAS (HCT116-MR). Results These models showed features related to the gene signature of Colorectal Cancer CMS4 with upregulation of immune pathway as confirmed by Microarray and Western blot analysis. In particular, moving forward the MEKi phenotype we assisted to the loss of epithelial features and acquisition of mesenchymal markers and morphology. Moreover, this change in the morphology is accompanied by up-regulation of PD-L1 expression and activation of EGFR and its downstream pathway, independently of cell line status mutation. To extend these in vitro findings, we performed an in vivo study using MC38 and CT26 MEKi resistant syngeneic models that we have previously generated. Combined treatment of MEKi, EGFR inhibitor (EGFRi) and PD-L1inhibitor (PD-L1i) resulted in a marked inhibition of tumor growth in both MC38–MR and CT26-MR xenograft model. Conclusions These results suggest a strategy to potentially overcome immunotherapy resistance in CMS4-like tumors and to improve the efficacy of MEK inhibition by co-treatment with other agents providing an additional therapeutic strategy via modulation of host immune responses. Legal entity responsible for the study The authors. Funding Has not received any funding. Disclosure All authors have declared no conflicts of interest.

Objective T-lymphocyte activation plays an important role in the pathophysiology of acute coronary syndromes (ACS). Plaques from ACS patients show a selective oligoclonal expansion of T-cells, indicating a specific, antigen-driven recruitment of T-lymphocytes within the unstable lesions. At present, however, it is not known whether T-cells may contribute directly to thrombosis by expressing functional tissue factor (TF). Accordingly, the aim of the present study was to investigate whether T-cells are able to express functional TF in their activated status. Methods In vitro , CD3 + -cells, isolated from buffy coats, were stimulated with anti-CD3/CD28 beads, IL-6, TNF-α, IL-17, INF-γ or PMA/ionomycin. Following stimulation, TF expression on cell-surface, at gene and protein levels, as well as its procoagulant activity in whole cells and microparticles was measured. In vivo , TF expression was evaluated in CD3 + -cells isolated from the aorta and the coronary sinus of ACS-NSTEMI and stable coronary artery disease (SCAD) patients. The presence of CD3 + -TF + cells was also evaluated by immunohistochemistry in thrombi aspirated from ACS-STEMI patients. Results PMA/ionomycin and IL-17 plus INF-γ stimulation resulted in a significant TF increase at gene and protein levels as well as at cell-surface expression. This was accompanied by a parallel increase in FXa generation, both in whole cells and in microparticles, indicating that the induced membrane-bound TF was active. Furthermore, transcardiac TF gradient was significantly higher in CD3 + -cells obtained from ACS-patients compared to SCAD-patients. Interestingly, thrombi from ACS-STEMI patients resulted enriched in CD3 + -cells, most of them expressing TF. Conclusions Our data demonstrate that activated T-lymphocytes in vitro express functional TF on their membranes, suggesting a direct pathophysiological role of these cells in the thrombotic process; this hypothesis is further supported by the observations in vivo that CD3 + -cells from coronary circulation of ACS-NSTEMI patients show increased TF levels and that coronary thrombi from ACS-STEMI patients are enriched in CD3 + -cells expressing TF.

Flavonoids are nutrients usually included in human diet with several significant biological activities. Nobiletin is a flavonoid that, besides having anti-inflammatory and anti-tumoral activity, seems to exert protective effects on cardiovascular system. Several studies investigated nobiletin as a natural drug to antagonize the atherosclerotic disease. On the contrary, literature about its potential role in modulating the main acute complication of atherosclerosis, thrombosis, is still scanty. Several studies have indicated that Tissue Factor (TF) plays a pivotal role in the pathophysiology of cardiovascular thrombotic events by triggering the formation of intracoronary thrombi. Oxidized-LDL have an important role in promoting athero-thrombotic events. This study investigates whether nobiletin might exert protective cardiovascular effects by preventing the oxidized-LDL mediated expression of TF in human endothelial cells in vitro. Moreover, we have studied whether the nobiletin effects might be modulated by the inhibition of the NF-kB pathway. Introduction: Flavonoids are nutrients usually included in human diet with several significant biological activities. Nobiletin is a flavonoid that, besides having anti-inflammatory and anti-tumoral activity, seems to exert protective effects on cardiovascular system. Several studies investigated nobiletin as a natural drug to antagonize the atherosclerotic disease. On the contrary, literature about its potential role in modulating the main acute complication of atherosclerosis, thrombosis, is still scanty.Several studies have indicated that Tissue Factor (TF) plays a pivotal role in the pathophysiology of cardiovascular thrombotic events by triggering the formation of intracoronary thrombi. Oxidized-LDL have an important role in promoting athero-thrombotic events. This study investigates whether nobiletin might exert protective cardiovascular effects by preventing the oxidized-LDL mediated expression of TF in human endothelial cells in vitro. Moreover, we have studied whether the nobiletin effects might be modulated by the inhibition of the NF-kappa B pathway.Methods and results: In HUVEC, ox-LDL induced TF-mRNA transcription as demonstrated by real time PCR and expression of functionally active TF as demonstrated by Western-blot, FACS analysis and pro coagulant activity assay. Nobiletin prevented these ox-LDL-mediated effects by exerting antioxidant effects, finally leading to inhibition of the transcription factor NF-kappa B.Conclusions: These data suggest that nobiletin might be a potential antithrombotic agent of dietary origin. This flavonoid, through its antioxidant proprieties, might potentially exert an antithrombotic activity by inhibiting TF expression/activity in a cell population never investigated before in this context and that is normally represented in vessel wall such as endothelial cells. (C) 2016 Elsevier Inc. All rights reserved.

pURPOSE: To assess the serum profile of factors involved in endothelial, T-cell, and fibroblast interplay in patients with Raynaud's phenomenon (RP) associated with nailfold vodeocapillaroscopy (NVC) scleroderma findings and/or systemic sclerosis (SSc) marker autoantibodies, recently labeled as early SSc patients. METHODS: Serum levels of soluble intercellular adhesion molecule-1 (sICAM-1), soluble vascular adhesion molecule-1 (sVCAM-1), CCL2, CXCL8, IL-13, IL-33, and transforming growth factor-β (TGF-β) were measured in 24 early SSc patients, 48 definite SSc patients, and 24 osteoarthritis/fibromyalgia controls by multiplex suspension immunoassay. All SSc patients were investigated for the presence/absence of preclinical and clinical organ involvement, SSc marker autoantibodies, and NVC abnormalities. RESULTS: Serum sICAM-1, CCL2, CXCL8, and IL-13 were increased in all SSc patients as compared to controls, and paralleled the severity of the disease subset (early SSc < limited cutaneous SSc < diffuse cutaneous SSc; p

Objective To investigate the phenotype and function of CD4+ T cells in synovial fluid (SF) from the affected joints of children with oligoarticular-onset juvenile idiopathic arthritis (JIA), and to establish a possible link with disease activity. Methods CD4+ T cells were obtained from the peripheral blood (PB) and SF of 23 children with oligoarticular-onset JIA, as well as from the PB of 15 healthy children. The cells were analyzed for the expression of CXCR3, CCR6, and CD161 and for the production of interferon-Î and interleukin-17A (IL-17A). Spectratyping and clonotype analyses were performed to assess different T cell subsets. Results The numbers of CD4+CD161+ cells showing either the Th1 or the Th17/Th1 phenotype were higher in the SF than in the PB of children with JIA. The few Th17 cells from JIA SF underwent a spontaneous shift to the Th1 phenotype in vitro, whereas Th17 cells from the PB of healthy children shifted only in the presence of JIA SF; this effect was neutralized by antibody blockade of IL-12 activity. Spectratyping and clonotype analyses showed a similar skewing of the T cell receptor V β repertoire in both CD161+ Th17 cells and CD161+ Th1 cells derived from the SF of the same JIA patient. The frequencies of CD4+CD161+ cells, particularly the Th17/Th1 cells, in the JIA SF positively correlated with the erythrocyte sedimentation rate and levels of C-reactive protein. Conclusion These findings suggest that a shifting of CD4+CD161+ T cells from Th17 to the Th17/Th1 or Th1 phenotype can occur in the SF of children with oligoarticular-onset JIA, and indicate that the accumulation of these cells is correlated with parameters of inflammation. Thus, the results support the hypothesis that these cells may play a role in JIA disease activity. Copyright © 2011 by the American College of Rheumatology.

Obesity is increasingly being recognized as a risk factor for a number of benign and malignant gastrointestinal conditions. However, literature on the underlying pathophysiological mechanisms is sparse and ambiguous. There is compelling evidence that both overnutrition and undernutrition negatively interfere with the immune system. Overnutrition has been found to increase susceptibility to the development of inflammatory diseases, autoimmune diseases and cancer. In the regulation of immune and inflammatory processes, white adipose tissue plays a critical role, not only as an energy store but also as an important endocrine organ. The obese state is characterised by a low-grade systemic inflammation, mainly as a result of increased adipocytes as well as fat resident- and recruited-macrophage activity. In the past few years, various products of adipose tissue including adipokines and cytokines have been characterised and a number of pathways linking adipose tissue metabolism with the immune system have been identified. Activation of the innate immune system plays a major role in hepatic steatosis. Non-alcoholic fatty liver disease includes a wide spectrum of diseases, from pure steatosis to non-alcoholic steatohepatitis in the absence of significant alcohol consumption. Although steatosis is considered a non-progressive disease, non-alcoholic steatohepatitis may deteriorate in advanced chronic liver diseases, cirrhosis, and hepatocellular carcinoma. An important parallel between obesity-related pathology of adipose tissue and liver pertains to the emerging role of macrophages, and growing evidence suggests that Kupffer cells critically contribute to progression of non-alcoholic fatty liver disease. Moreover, a close link between specific immune activation and atherosclerosis has been well established, suggesting that fat can directly trigger immune responses. This review discusses the role of fat as "a matter of disturbance for the immune system" with a focus on hepatic steatosis.