OBJECTIVE

To determine the vitreous levels of <em>27</em> types of cytokines in eyes with retinopathy of prematurity (ROP).

METHODS

Retrospective case-control study.

METHODS

Twenty-seven eyes of 19 infants with stage 4 ROP were studied. Six eyes of 5 patients with congenital cataract who underwent lensectomy were used as controls.

METHODS

The ROP eyes were divided into 2 groups according to vascular activity: 12 eyes with vascularly active ROP and 15 eyes with vascularly inactive ROP. Undiluted vitreous samples were collected, and the vitreous concentrations of <em>27</em> types of cytokines were determined by a multiplex bead analysis system: interleukin (IL)-1b, IL-1ra, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12, IL-13, IL-15, IL-17, Eotaxin, fibroblast growth factor (FGF) basic, granulocyte-colony stimulating factor (G-CSF), granulocyte macrophage colony-stimulating factor (GM-CSF), interferon-r, interferon-gamma-inducible protein (IP)-10, monocyte chemoattractant protein 1, macrophage inflammatory protein (MIP)-1a, MIP-1b, platelet-derived growth factor bb, regulated on activation, normal T cell expressed and secreted (RANTES), tumor necrosis factor alpha, and vascular endothelial growth factor (VEGF).

METHODS

The vitreous levels of the <em>27</em> types of cytokines and a comparison of the levels in the 3 groups.

RESULTS

The postmenstrual age at vitrectomy was significantly younger in the vascularly active ROP eyes than in vascularly inactive ROP eyes. The cytokines that had significantly different vitreous levels among the 3 groups were: IL-6, IL-7, IL-10, IL-15, Eotaxin, FGF basic, G-CSF, GM-CSF, IP-10, RANTES, and VEGF (P<0.05). The vitreous levels of IL-6, IL-7, IL-15, Eotaxin, G-CSF, IP-10, and RANTES were significantly higher (P<0.05) in both vascularly active and inactive ROP eyes than in control eyes, whereas the vitreous level of VEGF was significantly higher (P<0.05) only in vascularly active ROP eyes than in control eyes. There was a significantly negative correlation (r = -0.382; P = 0.0495) between the VEGF level and the postmenstrual age at vitrectomy.

CONCLUSIONS

These results indicate that, although cytokines other than VEGF may be involved in the pathologic changes in eyes with ROP, VEGF is likely to have the strongest correlation with the vascular activity in ROP eyes among these cytokines.

A large body of data indicate that antibody class switching is directed by cytokines by inducing or repressing transcription from unrearranged, or germline, CH genes. <em>Interleukin</em> 4 (IL-4) induces transcription of the germline C epsilon genes in activated B cells and subsequently, cells in this population will undergo switch recombination to immunoglobulin E. Furthermore, the data suggest that transcription of germline C epsilon genes is required for class switching. In this paper we define DNA elements required for induction of transcription of the germline C epsilon genes by IL-4. To do this, segments of DNA from the 5' flank of the initiation sites for germline epsilon RNA were ligated to a luciferase reporter gene and transfected into two mouse B cell lines, one of which can be induced to switch to IgE. By analysis of a series of 5' deletion constructs and linker-scanning mutations, we demonstrate that a 46-bp segment (residing at -126/-79 relative to the first RNA initiation site) contains an IL-4 responsive region. By electrophoretic mobility shift assays, we find that this segment binds three transcription factors: the recently described NF-IL4, one or more members of the C/EBP family of transcription factors, and NF-kappa B/p50. Mutation of any of the binding sites for these three factors abolishes or reduces IL-4 inducibility of the epsilon promoter. A <em>27</em>-bp segment within this IL-4 response region containing binding sites for NF-IL4 and a C/EBP factor is sufficient to transfer IL-4 inducibility to a minimal c-fos promoter.

OBJECTIVE

Despite the high metabolic demands of the neural retina, its detachment from the retinal pigment epithelium does not lead to immediate death for most of the cells. This study was undertaken to test the hypothesis that intrinsic protective mechanisms are activated in the neural retina during early stages of retinal detachment.

METHODS

Retinal detachments were created in Brown Norway rats by injection of 1% hyaluronic acid into the subretinal space. Gene expression profiles of retinas detached for 24 hours were generated with a gene microarray (rat U34 GeneChips; Affymetrix, Santa Clara, CA) and compared to the profiles from control attached retinas in a robust multiarray protocol and false-discovery-rate analysis. Changes in individual, differentially expressed genes were validated by quantitative real-time polymerase chain reaction (qRT-PCR) analysis. Additional qRT-PCR and immunoblot analyses were performed for additional selected genes.

RESULTS

Genome-wide expression profiling revealed <em>27</em> genes that are differentially expressed in retinas detached for 24 hours. In silico analysis and functional clustering suggested that most genes belonged to three signaling pathways: <em>interleukin</em>-6/STAT, transforming growth factor-beta/Smad, and aryl hydrocarbon receptor oxidative stress response. Additional analyses of selected genes from these pathways demonstrated a time-dependent increase in their expression in detached retinas.

CONCLUSIONS

Retinal detachment results in the early activation of stress-response genes and specific signaling pathways. This adaptive response may enable the photoreceptor cells to survive the acute phase of a retinal detachment, and it is the breakdown of these protective mechanisms in chronic disease that leads to the ultimate death of the cell.

BACKGROUND

Melanoma antigen (MA)-specific vaccination strongly enhances antitumor reactivity in vivo and is capable of producing strong cytotoxic T lymphocyte responses in vitro. Furthermore, specific human leukocyte antigen-restricted T cell activation is hypothesized to occur in response to peptide-based immunotherapy, which may lead to the preferential killing of tumor cells bearing the relevant MA. The development of melanoma antigen-loss variants may subsequently occur in vivo.

METHODS

Analysis of 532 melanoma lesions from 204 patients was performed on fine-needle aspiration biopsy specimens. Lesions were graded for the expression of the MAs gp100 and MART-1 with use of immunocytochemistry. A total of 351 melanoma lesions were divided into cohorts on the basis of the treatment received. The pretreatment group (n = 175) consisted of lesions obtained before any form of gp100 immunotherapy, with the posttreatment group (n = 176) consisting of lesions obtained after vaccination with a modified gp100 epitope, gp209-2M +/- interleukin 2 (IL-2).

RESULTS

The percentage of lesions not expressing the gp100 antigen is greater than the percentage not expressing MART-1 (26% vs 14%). The frequency of lesions with high expression >> 75%) of gp100 significantly decreased with therapy (47% vs 34%) and conversely negative lesions increased (18% vs 29%). Treatment of lesions with peptide alone (no IL-2) revealed a significant decrease in gp100 expression (47% vs 32%), enhanced with the addition of IL-2 to therapy (47% vs 35%). The expression of MART-1 remained essentially unchanged unless IL-2 was added (54% vs 54%, MART-1 peptide alone, 54% vs 43%, MART-1 peptide + IL-2). Of 94 patients (181 lesions) assessed for gp100 expression before treatment, 10 patients responded to therapy. Pretreatment lesions in responding patients expressed some level of gp100 in all cases compared with 27% of nonresponding lesions, which were negative for gp100 expression. CONCLUSIONS. This study indirectly demonstrates that vaccination with an MA-derived peptide can result in immune selection in vivo. Furthermore, it provides strong immunologic evidence for the specificity of MA vaccines and to the relevance of MA expression in predicting the response to vaccination.

The purpose of this study was to investigate whether an age-associated impaired acute-phase response exists. Nine healthy elderly volunteers (median, 66 years; range, 61 to 69 years) and eight young controls (median, 24 years; range, 20 to <em>27</em> years) were given an intravenous bolus of endotoxin (2 ng/kg). The rectal temperature was monitored continuously, and blood samples for cytokine measurements were obtained before endotoxin administration as well as 0.5, 1, 1.5, 2, 3, 4, 8, 12, and 24 h after the injection. The elderly subjects showed a more prolonged fever response compared to the young controls. Levels of tumor necrosis factor alpha (TNF-alpha), soluble TNF receptors (sTNFR-I), <em>interleukin</em>-6 (IL-6), IL-8, IL-10, and IL-1 receptor antagonist (IL-1ra) in plasma increased markedly following endotoxin administration in both groups. The elderly group showed larger initial increases in TNF-alpha and sTNFR-I levels and prolonged increased levels of sTNFR-I. Monocyte concentrations decreased in both groups, with the elderly group showing a more rapid decrease and a slower subsequent increase than did the young group. Furthermore, the elderly group had a more rapid increase in C-reactive protein levels than did the young group. In conclusion, ageing is associated with an altered acute-phase response including initial hyperreactivity, prolonged inflammatory activity, and prolonged fever response.

OBJECTIVE

Interleukin (IL) 34 is a new cytokine implicated in macrophage differentiation and osteoclastogenesis. This study assessed IL-34 expression in the tissue of patients with rheumatoid arthritis (RA).

METHODS

Immunohistochemistry was performed in synovial biopsies from patients with RA (n=20), osteoarthritis (n=3) or other inflammatory arthritis (n=4). IL-34 was detected in the synovial fluid by ELISA and its messenger RNA expression was studied by quantitative PCR in rheumatoid synovial fibroblasts after stimulation by tumour necrosis factor α (TNFα) and IL-1β. Wild-type, jnk1(-/-)-jnk2(-/-) and nemo(-/-) murine fibroblasts and pharmacological inhibition were used to determine the involvement of nuclear factor kappa B (NF-κB) and JNK in that effect.

RESULTS

IL-34 was expressed in 24/27 biopsies, with three samples from RA patients being negative. A significant association was found between IL-34 expression and synovitis severity. Levels of IL-34 and the total leucocyte count in synovial fluid were correlated. TNFα and IL-1β stimulated IL-34 expression by synovial fibroblasts in a dose/time-dependent manner through the NF-κB and JNK pathway.

CONCLUSIONS

This work for the first time identifies IL-34 expression in the synovial tissue of patients with arthritis. This cytokine, as a downstream effector of TNFα and IL-1β, may contribute to inflammation and bone erosions in RA.

Despite recent advances, median survival for patients with resectable glioblastoma multiforme (GBM) is only 12 to 15 months. We previously observed minimal toxicity and a 9.0-month median survival after treatment with intralesional autologous lymphokine-activated killer (LAK) cells in 40 patients with recurrent GBM. In this study, GBM patients were treated with adjuvant intralesional LAK cells. Eligible patients had completed primary therapy for GBM without disease progression. LAK cells were produced by incubating autologous peripheral blood mononuclear cells with <em>interleukin</em>-2 for 3 to 7 days and then placed into the surgically exposed tumor cavity by a neurosurgeon. The 19 men and 14 women had a median age of 57 years. Prior therapy included surgical resection (97%), partial brain irradiation (97%), gamma knife radiosurgery (97%), and temozolomide chemotherapy (70%). Median time from diagnosis to LAK cell therapy was 5.3 months (range: 3.0 to 11.1 mo). LAK cell treatment was well tolerated; average length of hospitalization was 3 days. At the time of this analysis, <em>27</em> patients have died; the median survival from the date of original diagnosis is 20.5 months with a 1-year survival rate of 75%. In subset analyses, superior survival was observed for patients who received higher numbers of CD3+/CD16+/CD56+ (T-LAK) cells in the cell products, which was associated with not taking corticosteroids in the month before leukopheresis. Intralesional LAK cell therapy is safe and the survival sufficiently encouraging to warrant further evaluation in a randomized phase 2 trial of intralesional therapies with LAK or carmustine-impregnated wafers.

OBJECTIVE

Interleukin (IL)-8, a pro-inflammatory cytokine, is a potent chemoattractant factor and an activator of neutrophils produced by many cell types after stimulation by IL-1, tumor necrosis factor (TNF), or microbial products such as endotoxins. We investigated whether the presence of measurable IL-8 in plasma was associated with the clinical status of severely ill septic or nonseptic patients susceptible to the development of multiple organ failure.

METHODS

Cohort study.

METHODS

A collaborative study between an intensive care unit and a research laboratory.

METHODS

Circulating IL-8 concentrations were measured in the plasma of 27 patients with sepsis syndrome and in 16 patients with noninfectious shock because these two conditions put patients at risk for the development of multiple organ failure. Sixteen of 27 patients with severe infection and 13 of 16 patients with noninfectious pathologies developed multiple organ failure.

RESULTS

A specific enzyme-linked immunosorbent assay (ELISA) for IL-8 was set up with a monoclonal and a rabbit polyclonal antihuman IL-8 using a sandwich technique. High concentrations of circulating IL-8 were found in the plasma of patients with sepsis syndrome. Among septic patients, a significant difference was observed between concentrations of IL-8 in survivors (n = 16) and nonsurvivors (n = 11) (81 +/- 13 pg/mL vs. 3326 +/- 1219 pg/mL, respectively; p = .001). A correlation was noticed between plasma IL-8 and IL-6 concentrations (r2 = .42; p = .001), while no correlation was observed between IL-8 and TNF-alpha values, or between IL-8 and IL-1 beta. Although the mortality rate of nonseptic, multiple organ failure patients was 92%, low plasma concentrations of IL-8 were found (78 +/- 34 pg/mL), while high plasma concentrations were measured in septic, multiple organ failure patients (mortality rate 69%) who were sampled at a similar stage. By contrast, increased IL-6 values were observed in both septic and nonseptic, multiple organ failure patients.

CONCLUSIONS

In septic patients, high amounts of circulating IL-8 concentrations correlate with fatal outcome, whereas only low plasma concentrations of IL-8 are present in patients with nonseptic, multiple organ failure. This finding suggests that the signals involved in the exacerbation of IL-8 production are different, depending on infectious or noninfectious etiology.

<em>Interleukin</em>-<em>27</em> (IL-<em>27</em>) is a new member of the IL-12 family. It is produced by activated antigen-presenting cells and plays an important role in the regulation of CD4+ T cell differentiation and immune response. IL-<em>27</em> activates multiple signaling cascades, including the JAK-STAT and p38 MAPK pathways. Several studies have revealed that IL-<em>27</em> promotes the differentiation of Th1 and Tr1, but inhibits Th2, Th17, and Treg cells. However, a few studies have shown an opposite effect on certain T cell subsets, such as Treg. IL-<em>27</em> displays both pro- and anti- inflammatory activities in different autoimmune diseases. Here, we have discussed the role of IL-<em>27</em> in rheumatoid arthritis, multiple sclerosis, colitis, lupus, psoriasis, type 1 diabetes, and uveitis. Most of this information is derived from experimental models of these autoimmune diseases. The mechanistic basis of the dual role of IL-<em>27</em> in inflammation and autoimmunity is still not fully defined. In general, the pro-/anti-inflammatory activity of IL-<em>27</em> is influenced by the underlying immune effector pathways, the phase of the disease, the presence or absence of counter-regulatory cytokines/T cell subsets, and the tissue/cell type under study. Despite a spectrum of outcomes in various autoimmune diseases, mostly anti-inflammatory and immunomodulatory effects of IL-<em>27</em> have been observed in this category of diseases. Accordingly, IL-<em>27</em> represents a novel, promising target/agent for the treatment of autoimmune diseases.

Smac, second mitochondria-derived activator of caspases, promotes apoptosis via activation of caspases. Heat shock protein <em>27</em> (Hsp<em>27</em>) negatively regulates another mitochondrial protein, cytochrome c, during apoptosis; however, the role of Hsp<em>27</em> in modulating Smac release is unknown. Here we show that Hsp<em>27</em> is overexpressed in both dexamethasone (Dex)-resistant multiple myeloma (MM) cell lines (MM.1R, U266, RPMI-8226) and primary patient cells. Blocking Hsp<em>27</em> by an antisense (AS) strategy restores the apoptotic response to Dex in Dex-resistant MM cells by triggering the release of mitochondrial protein Smac, followed by activation of caspase-9 and caspase-3. Moreover, AS-Hsp<em>27</em> overcomes <em>interleukin</em>-6 (IL-6)-mediated protection against Dex-induced apoptosis. These data demonstrate that Hsp<em>27</em> inhibits the release of Smac, and thereby confers Dex resistance in MM cells.

Severe asthma is a complex heterogeneous disease associated with older age and obesity. The presence of eosinophilic (type 2) inflammation in some but not all patients with severe asthma predicts responsiveness to current treatments, but new treatment approaches will require a better understanding of non-type 2 mechanisms of severe asthma. We considered the possibility that systemic inflammation, which arises in subgroups of obese and older patients, increases the severity of asthma. Interleukin-6 (IL-6) is a biomarker of systemic inflammation and metabolic dysfunction, and we aimed to explore the association between IL-6 concentrations, metabolic dysfunction, and asthma severity.

In this cross-sectional analysis, patients were recruited from two cohorts: mainly non-severe asthmatics from the University of California San Francisco (UCSF) and mainly severe asthmatics from the Severe Asthma Research Program (SARP). We generated a reference range for plasma IL-6 in a cohort of healthy control patients. We compared the clinical characteristics of asthmatics with plasma IL-6 concentrations above (IL-6 high) and below (IL-6 low) the upper 95% centile value for plasma IL-6 concentration in the healthy cohort. We also compared how pulmonary function, frequency of asthma exacerbations, and frequency of severe asthma differed between IL-6 low and IL-6 high asthma populations in the two asthma cohorts.

Between Jan 1, 2005, and Dec 31, 2014, we recruited 249 patients from UCSF and between Nov 1, 2012, and Oct 1, 2014, we recruited 387 patients from SARP. The upper 95th centile value for plasma IL-6 concentration in the healthy cohort (n=93) was 3·1 pg/mL, and 14% (36/249) of UCSF cohort and 26% (102/387) of the SARP cohort had plasma IL-6 concentrations above this upper limit. The IL-6 high patients in both asthma cohorts had a significantly higher average BMI (p<0·0001) and a higher prevalence of hypertension (p<0·0001) and diabetes (p=0·04) than the IL-6 low patients. IL-6 high patients also had significantly worse lung function and more frequent asthma exacerbations than IL-6 low patients (all p values <0·0001). Although 80% (111/138) of IL-6 high asthmatic patients were obese, 62% (178/289) of obese asthmatic patients were IL-6 low. Among obese patients, the forced expiratory volume in 1 s (FEV1) was significantly lower in IL-6 high than in IL-6 low patients (mean percent predicted FEV1=70·8% [SD 19·5] vs 78·3% [19·7]; p=0·002), and the percentage of patients reporting an asthma exacerbation in the past 1-2 years was higher in IL-6 high than in IL-6 low patients (66% [73/111] vs 48% [85/178]; p=0·003). Among non-obese asthmatics, FEV1 values and the frequency of asthma exacerbations within the past 1-2 years were also significantly worse in IL-6 high than in IL-6 low patients (mean FEV1 66·4% [SD 23·1] vs 83·2% [20·4] predicted; p<0·0001; 59% [16/27] vs 34% [108/320]; p=0·01).

Systemic IL-6 inflammation and clinical features of metabolic dysfunction, which occur most commonly in a subset of obese asthma patients but also in a small subset of non-obese patients, are associated with more severe asthma. These data provide strong rationale to undertake clinical trials of IL-6 inhibitors or treatments that reduce metabolic dysfunction in a subset of patients with severe asthma. Plasma IL-6 is a biomarker that could guide patient stratification in these trials.

NIH and the Parker B Francis Foundation.

Asthma is a heterogeneous inflammatory airways disorder where <em>interleukin</em> (IL)-1β is thought to be a key mediator, especially in the neutrophilic subtype of asthma. The generation of active IL-1β requires proteolytic cleavage typically mediated through the formation of a caspase-1-containing inflammasome. This study hypothesised that an IL-1β endotype associated with the nucleotide-binding domain, leucine-rich repeat-containing family protein (NLRP)3/apoptosis-associated speck-like protein containing a caspase-recruitment domain (ASC)/caspase-1 inflammasome is characteristic of patients with the neutrophilic subtype of asthma. Participants with asthma (n=85) and healthy controls (n=<em>27</em>) underwent clinical assessment, spirometry and sputum induction. Sputum was processed for differential cell count, gene expression and protein mediators. NLRP3 and caspase-1 expression was also determined by immunocytochemistry. Sputum macrophages were isolated (n=8) and gene expression of NLRP3 and IL-1β determined. There was significantly elevated gene expression of NLRP3, caspase-1, caspase-4, caspase-5 and IL-1β in participants with neutrophilic asthma. Protein levels of IL-1β were significantly higher in those with neutrophilic asthma and correlated with sputum IL-8 levels. Sputum macrophages, as well as sputum neutrophils in neutrophilic asthma, expressed NLRP3 and caspase-1 protein. NLRP3 inflammasome is upregulated in neutrophilic asthma and may regulate the inflammation process observed in this asthma phenotype through production of IL-1β.

Chronic HIV infection is characterized by chronic immune activation and dysfunctional T cells with elevated intracellular cyclic AMP (cAMP), which inhibits the T cell activation capability. cAMP may be induced by prostaglandin E(2) following lipopolysaccharide (LPS)-induced upregulation of cyclooxygenase type 2 (COX-2) in monocytes due to the elevated LPS levels in patients with chronic HIV infection. This hypothesis was tested using celecoxib, a COX-2 inhibitor, for 12 weeks in HIV-infected patients without antiretroviral treatment in a prospective, open, randomized exploratory trial. Thirty-one patients were randomized in the trial; <em>27</em> completed the study, including 13 patients on celecoxib. Celecoxib reduced chronic immune activation in terms of CD38 density on CD8(+) T cells (-24%; P = 0.04), IgA levels (P = 0.04), and a combined score for inflammatory markers (P < 0.05). Celecoxib further reduced the inhibitory surface receptor programmed death 1 (PD-1) on CD8(+) T cells (P = 0.01), including PD-1 on the HIV Gag-specific subset (P = 0.02), enhanced the number of CD3(+) CD4(+) CD25(+) CD1<em>27</em>(lo/-) Treg or activated cells (P = 0.02), and improved humoral memory recall responses to a T cell-dependent vaccine (P = 0.04). HIV RNA (P = 0.06) and D dimers (P = 0.07) tended to increase in the controls, whereas <em>interleukin</em>-6 (IL-6) possibly decreased in the treatment arm (P = 0.10). In conclusion, celecoxib downmodulated the immune activation related to clinical progression of chronic HIV infection and improved T cell-dependent functions in vivo.

The <em>interleukin</em>-12 (IL-12) family of cytokines which includes IL-12, IL-23, and IL-<em>27</em> play critical roles in T cell differentiation and are important modulators of multiple sclerosis and experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis. Previously, we demonstrated that peroxisome proliferator-activated receptor (PPAR) -alpha agonists suppress the development of EAE. The present studies demonstrated that the PPAR-alpha agonist fenofibrate inhibited the secretion of IL-12p40, IL-12p70 (p35/p40), IL-23 (p19/p40), and IL-<em>27</em>p28 by lipopolysaccharide-stimulated microglia. The cytokines interferon-gamma and tumor necrosis factor-alpha also stimulated IL-12 p40 and IL-<em>27</em> p28 expression by microglia, which was suppressed by fenofibrate. Furthermore, fenofibrate inhibited microglial expression of CD14 which plays a critical role in TLR signaling, suggesting a mechanism by which this PPAR-alpha agonist regulates the production of these pro-inflammatory molecules. In addition, fenofibrate suppressed the secretion of IL-12p40, IL-23, and IL-<em>27</em>p28 by lipopolysaccharide-stimulated astrocytes. Importantly, fenofibrate suppression of EAE was associated with decreased expression of IL-12 family cytokine mRNAs as well as mRNAs encoding TLR4, CD14, and MyD88 known to play critical roles in MyD88-dependent TLR signaling. These novel observations suggest that PPAR-alpha agonists including fenofibrate may modulate the development of EAE, at least in part, by suppressing the production of IL-12 family cytokines and MyD88-dependent signaling.

BACKGROUND

Differentiating between sterile inflammation and bacterial infection in critically ill patients with fever and other signs of the systemic inflammatory response syndrome (SIRS) remains a clinical challenge. The objective of our study was to mine an existing genome-wide expression database for the discovery of candidate diagnostic biomarkers to predict the presence of bacterial infection in critically ill children.

METHODS

Genome-wide expression data were compared between patients with SIRS having negative bacterial cultures (n = 21) and patients with sepsis having positive bacterial cultures (n = 60). Differentially expressed genes were subjected to a leave-one-out cross-validation (LOOCV) procedure to predict SIRS or sepsis classes. Serum concentrations of <em>interleukin</em>-<em>27</em> (IL-<em>27</em>) and procalcitonin (PCT) were compared between 101 patients with SIRS and 130 patients with sepsis. All data represent the first 24 hours of meeting criteria for either SIRS or sepsis.

RESULTS

Two hundred twenty one gene probes were differentially regulated between patients with SIRS and patients with sepsis. The LOOCV procedure correctly predicted 86% of the SIRS and sepsis classes, and Epstein-Barr virus-induced gene 3 (EBI3) had the highest predictive strength. Computer-assisted image analyses of gene-expression mosaics were able to predict infection with a specificity of 90% and a positive predictive value of 94%. Because EBI3 is a subunit of the heterodimeric cytokine, IL-<em>27</em>, we tested the ability of serum IL-<em>27</em> protein concentrations to predict infection. At a cut-point value of ≥5 ng/ml, serum IL-<em>27</em> protein concentrations predicted infection with a specificity and a positive predictive value of >90%, and the overall performance of IL-<em>27</em> was generally better than that of PCT. A decision tree combining IL-<em>27</em> and PCT improved overall predictive capacity compared with that of either biomarker alone.

CONCLUSIONS

Genome-wide expression analysis has provided the foundation for the identification of IL-<em>27</em> as a novel candidate diagnostic biomarker for predicting bacterial infection in critically ill children. Additional studies will be required to test further the diagnostic performance of IL-<em>27</em>. The microarray data reported in this article have been deposited in the Gene Expression Omnibus under accession number GSE4607.

BACKGROUND

Supplementation with 1 g/d omega-3-polyunsaturated fatty acids (n3-PUFAs) demonstrated a small survival advantage in patients with chronic heart failure (CHF) in the GISSI-HF trial. However, a dose-efficacy relationship was postulated for the beneficial effects of n3-PUFA before. Therefore, we evaluated dose-dependent effects of n3-PUFA in patients with severe CHF.

METHODS

In a double-blind, randomized, controlled pilot trial, 43 patients with severe, nonischemic heart failure received 1 g/d n3-PUFA (n = 14), 4 g/d n3-PUFA (n = 13), or placebo (n = 16) for 3 months. Changes in left ventricular ejection fraction (LVEF), flow-mediated vasodilation, plasma high-sensitive interleukin 6 and high-sensitive tumor necrosis factor α, and exercise peak oxygen consumption were assessed.

RESULTS

Left ventricular ejection fraction increased in a dose-dependent manner (P = .01 for linear trend) in the 4 (baseline vs 3 months [mean ± SD]: 24% ± 7% vs 29% ± 8%, P = .005) and 1 g/d treatment groups (24% ± 8% vs 27% ± 8%, P = .02). Flow-mediated vasodilation increased significantly with high-dose 4 g/d n3-PUFA (8.4% ± 4.8% vs 11.6% ± 7.0%, P = .01) but only trendwise with low-dose 1 g/d (8.3% ± 5.3% vs 10.2% ± 4.3%, P = .07). Interleukin 6 significantly decreased with 4 g/d n3-PUFA (3.0 ± 2.9 pg/mL vs 0.7 ± 0.8 pg/mL, P = .03) but only trendwise with 1 g/d (4.5 ± 6.6 pg/mL to 1.6 ± 2.1 pg/mL, P = .1). High-sensitive tumor necrosis factor α decreased trendwise with 4 g/d n3-PUFA but remained unchanged with 1 g/d. In patients with maximal exercise effort, only 4 g/d increased the peak oxygen consumption. No changes in any investigated parameters were noted with placebo.

CONCLUSIONS

Treatment with n3-PUFA for 3 months exerts a dose-dependent increase of LVEF in patients with CHF. In parallel, a significant improvement of endothelial function and decrease of interleukin 6 is found with high-dose n3-PUFA intervention.

MUC1 over-expression in renal clear-cell carcinoma (RCC) is associated with poor prognosis. This phase II study determined the efficacy and tolerability of TG4010, a cancer vaccine based on a modified vaccinia virus expressing MUC1 and <em>interleukin</em>-2, in combination with cytokines, as first-line therapy in metastatic RCC. Thirty-seven patients with progressive, MUC1-positive RCC received TG4010 10(8) pfu/inj weekly for 6 weeks, then every 3 weeks until progression, when TG4010 was continued in combination with interferon-α2a and <em>interleukin</em>-2. Assessments included clinical response (primary endpoint), safety, time to treatment failure (TTF), overall survival (OS), and immune response. No objective clinical responses occurred. Five of the <em>27</em> evaluable patients (18%) had stable disease for >6 months with TG4010 alone and six of 20 patients (30%) had stable disease for >6 months with TG4010 plus cytokines. Median TTF was 4.1, 3.6, and 9.3 months for monotherapy, combination therapy, and overall, respectively. Median OS was 19.3 months for all patients and 22.4 months combination therapy recipients. The most frequent TG4010-related adverse events were minor-to-moderate injection-site reactions, fatigue, and flu-like symptoms. Six of 28 patients showed a MUC1 CD4+ T cell proliferative response during therapy. Anti-MUC1 CD8+ T cells were detected before and after therapy in 3 and 4 patients, respectively. MUC1-specific CD8+ T cell responses were associated with longer survival. Therapy with TG4010 plus cytokines appears to be feasible and well tolerated in patients with metastatic RCC. However, these data should be interpreted with caution, as additional prospective studies are necessary to clarify the clinical efficacy of this therapy.

OBJECTIVE

To compare the changes in the levels of <em>27</em> aqueous humor cytokines between nondiabetic controls and patients with type 2 diabetes and to ascertain the association of these cytokines with diabetic retinopathy (DR).

METHODS

Undiluted aqueous humor samples were obtained from 102 nondiabetic patients (102 eyes) and 136 consecutive diabetic patients (136 eyes) who were divided into nine groups according to the Early Treatment of Diabetic Retinopathy Study severity scale. The concentrations of <em>27</em> cytokines in the aqueous humor samples were measured using a multiplex bead immunoassay.

RESULTS

Compared with the nondiabetic controls, the diabetic patients had significantly higher concentrations of interleukin-1β (IL-1β; p<0.001), IL-6 (p<0.001), IL-8 (p<0.001), monocyte chemoattractant protein-1 (p<0.001), interferon gamma-induced protein-10 (p<0.001), and vascular endothelial growth factor (p<0.001) in the aqueous humor. However, the IL-10 (p=0.002) and IL-12 (p=0.013) concentrations were significantly lower for the diabetic patients. There were no significant differences in the concentrations of other cytokines between the diabetic patients and the controls. The IL-1β, IL-6, IL-8, monocyte chemoattractant protein-1, and interferon gamma-induced protein-10 levels in the aqueous humor increased as the severity of DR increased. The correlation was significant. However, the vascular endothelial growth factor concentration was not correlated with the severity of DR. In addition, the IL-10 and IL-12 levels in the aqueous humor decreased as the severity of DR increased, and this negative correlation was significant.

CONCLUSIONS

Various cytokines associated with inflammation and angiogenesis may contribute to the pathogenesis of DR, and chemokines may be more closely related to the development of this disease.

BACKGROUND

Interleukin-1 (IL-1) plays an important role in the pathophysiology and progression of rheumatoid arthritis (RA) by contributing to destruction of cartilage, bone, and periarticular tissues. Inhibiting IL-1 synthesis or activity with the use of recombinant human IL-1 receptor antagonist (anakinra) may prove to be an effective approach to the treatment of RA.

OBJECTIVE

The purpose of this article is to review the effects of anakinra in the treatment of RA.

METHODS

A MEDLINE search from 1982 to 2003 was used to identify animal studies and randomized clinical trials of anakinra and other therapies that target IL-1.

RESULTS

Clinical trials of anakinra have shown that it reduces the signs and symptoms of active disease and slows the rate of radiographic destruction in adults with RA. With anakinra 150 mg/d, 43% of patients achieved an American College of Rheumatology (ACR) 20% response, compared with 27% with placebo (P = 0.014). The ACR20 score indicates at least 20% improvement in the ACR composite score, which includes assessment of tender and swollen joint count, and other clinical end points such as pain and disability assessment. Patients treated with anakinra also experienced a 59% reduction in new bony erosion compared with controls (P < 0.001) and a 65% reduction in joint space narrowing as measured by the modified Sharp score (P = 0.020). Injection-site reactions were the most commonly reported adverse event, occurring in 50%, 73%, and 81% of patients receiving anakinra 30, 75, and 150 mg/d, respectively, compared with 25% of patients receiving placebo. Few serious adverse events were reported, and they typically occurred in patients receiving the highest daily dosage.

CONCLUSIONS

IL-1 is an important cytokine in promoting the damage associated with RA. Anakinra is mildly to moderately effective and well tolerated in patients with active RA when used as monotherapy or in combination with methotrexate.

Purpose <em>Interleukin</em>-10 (IL-10) stimulates the expansion and cytotoxicity of tumor-infiltrating CD8+ T cells and inhibits inflammatory CD4+ T cells. Pegylation prolongs the serum concentration of IL-10 without changing the immunologic profile. This phase I study sought to determine the safety and antitumor activity of AM0010. Patients and Methods Patients with selected advanced solid tumors were treated with AM0010 in a dose-escalation study, which was followed by a renal cell cancer (RCC) dose-expansion cohort. AM0010 was self-administered subcutaneously at doses of 1 to 40 μg/kg once per day. Primary end points were safety and tolerability; clinical activity and immune activation were secondary end points. Results In the dose-escalation and -expansion cohorts, 33 and 18 patients, respectively, were treated with daily subcutaneous injection of AM0010. AM0010 was tolerated in a heavily pretreated patient population. Treatment-related adverse events (AEs) included anemia, fatigue, thrombocytopenia, fever, and injection site reactions. Grade 3 to 4 nonhematopoietic treatment-related AEs, including rash (n = 2) and transaminitis (n = 1), were observed in five of 33 patients. Grade 3 to 4 anemia or thrombocytopenia was observed in five patients. Most treatment-related AEs were transient or reversible. AM0010 led to systemic immune activation with elevated immune-stimulatory cytokines and reduced transforming growth factor beta in the serum. Partial responses were observed in one patient with uveal melanoma and four of 15 evaluable patients with RCC treated at 20 μg/kg (overall response rate, <em>27</em>%). Prolonged stable disease of at least 4 months was observed in four patients, including one with colorectal cancer with disease stabilization for 20 months. Conclusion AM0010 has an acceptable toxicity profile with early evidence of antitumor activity, particularly in RCC. These data support the further evaluation of AM0010 both alone and in combination with other immune therapies and chemotherapies.

The genetic contribution to common forms of osteoarthritis (OA) is well established but poorly understood. We performed a genome scan, using 302 markers for loci predisposing to distal interphalangeal joint (DIP) OA. To minimize genetic heterogeneity in our study sample, we identified siblings with a severe, radiologically defined phenotype from the nationwide registers of Finland. In the initial genome scan, linkage analysis in <em>27</em> sibships gave a pairwise LOD score (Z) >1.00 with nine of the screening markers. In the second stage, additional markers and family members were genotyped in these chromosomal regions. On 2q12-q13, IL1R1 resulted in Z=2.34 at recombination fraction (theta) 0, allowing a dominant mode of inheritance. Association analysis of markers D2S2264, IL1R1, D2S373, and D2S1789 jointly provided some evidence for a shared haplotype among the affected individuals (P value of.012). Also, multipoint nonparametric linkage analysis yielded a P value of.0001 near the locus IL1R1 and P=.0007 approximately 20 cM telomeric near marker D2S1399, which, in two-point analysis, gave Z=1.48 (straight theta=. 02). This chromosomal region on 2q harbors the <em>interleukin</em> 1 gene cluster and, thus, represents a good candidate region for inflammatory and autoimmune disorders. Three additional chromosomal regions-4q26-q<em>27</em>, 7p15-p21, and Xcen-also provided some evidence for linkage, and further analyses would be justified to clarify their potential involvement in the genetic predisposition to DIP OA.

Histone methylation is crucial for transcriptional regulation and chromatin remodeling. It has been suggested that the SET domain containing protein RE-IIBP (<em>interleukin</em>-5 [IL-5] response element II binding protein) may perform a function in the carcinogenesis of certain tumor types, including myeloma. However, the pathogenic role of RE-IIBP in these diseases remains to be clearly elucidated. In this study, we have conducted an investigation into the relationship between the histone-methylating activity of RE-IIBP and transcriptional regulation. Here, we report that RE-IIBP is up-regulated in the blood cells of leukemia patients, and we characterized the histone H3 lysine <em>27</em> (H3-K<em>27</em>) methyltransferase activity of RE-IIBP. Point mutant analysis revealed that SET domain cysteine 483 and arginine 477 are critical residues for the histone methyltransferase (HMTase) activity of RE-IIBP. RE-IIBP also represses basal transcription via histone deacetylase (HDAC) recruitment, which may be mediated by H3-K<em>27</em> methylation. In the chromatin immunoprecipitation assays, we showed that RE-IIBP overexpression induces histone H3-K<em>27</em> methylation, HDAC recruitment, and histone H3 hypoacetylation on the IL-5 promoter and represses expression. Conversely, short hairpin RNA-mediated knockdown of RE-IIBP reduces histone H3-K<em>27</em> methylation and HDAC occupancy around the IL-5 promoter. These data illustrate the important regulatory role of RE-IIBP in transcriptional regulation, thereby pointing to the important role of HMTase activity in carcinogenesis.

Deficient production of <em>interleukin</em>-2 has been reported in Type I diabetes, but its cause has not been elucidated. We therefore measured <em>interleukin</em>-2 production in <em>27</em> patients with Type I diabetes, 20 patients with Type II diabetes (6 requiring insulin), 5 monozygotic twin pairs discordant for Type I diabetes, and 10 nondiabetic persons with islet-cell antibodies. <em>Interleukin</em>-2 production was decreased in patients with Type I diabetes as compared with controls (35.8 +/- 2.5 vs. 61.6 +/- 4.6 percent, P less than 0.001). <em>Interleukin</em>-2 production did not differ between patients with Type II diabetes and controls, regardless of whether the patients used insulin. Twins with Type I diabetes had decreased <em>interleukin</em>-2 production as compared with normal controls (33.2 +/- 5.4 vs. 61.6 +/- 4.6 percent, P less than 0.001) and with their nondiabetic twins (33.2 +/- 5.4 vs. 54.5 +/- 3.4 percent, P less than 0.005). <em>Interleukin</em>-2 production in nondiabetic twins and in nondiabetic persons with islet-cell antibodies was normal. There was no correlation between glycosylated hemoglobin levels and <em>interleukin</em>-2 production in any diabetic group. We conclude that patients with Type I diabetes have an acquired defect in <em>interleukin</em>-2 production, whereas patients with Type II diabetes do not, and that this defect is not correlated with an ongoing autoimmune process, with hyperglycemia, or with insulin administration or oral hypoglycemic therapy. Thus, the defect appears to be related to marked beta-cell destruction, although not to the metabolic consequences thereof or the responsible autoimmune process.

Angiogenesis has an important role in the progression of solid tumors. Therefore, we measured the blood levels (ELISA) of angiogenic factors [basic fibroblast growth factor (bFGF), hepatocyte growth factor/scatter factor, and vascular endothelial growth factor (VEGF)] and soluble adhesion molecules [E-selectin, intercellular adhesion molecule (ICAM-1), platelet endothelial cell adhesion molecule-1, and vascular cell adhesion molecule-1] in 76 consecutive patients with untreated renal cell carcinoma and 41 healthy controls to evaluate their prognostic value. The serum levels of bFGF, hepatocyte growth factor, and VEGF were significantly higher in patients with renal cancer than they were in healthy subjects. bFGF and VEGF values were significantly higher in patients with disseminated cancer (N+ and/or M+) than they were in those with undisseminated (M-N-) cancer: median = <em>27</em> pg/ml, range = 5-118, n = 15 versus median = 8 pg/ml, range = 1-149, n = 61 (P = 10(-4)) for bFGF; and median = 883 pg/ml, range = 200-2317, n = 15 versus median = <em>27</em>8 pg/ml, range = 0-1704, n = 61 (P = 0.006) for VEGF. The blood levels of ICAM-1 and vascular cell adhesion molecule-1 were significantly higher, and the levels of E-selectin and platelet endothelial cell adhesion molecule-1 were significantly lower in patients with renal cancer than they were in controls. Plasma ICAM-1 was higher in metastatic patients (M+) than they were in nonmetastatic (M-) patients: median = 687 ng/ml, range = 294-1091, n = 12 versus median = 408 ng/ml, range = 217-1375, n = 64 (P = 10(-4)). ICAM-1 and bFGF blood values were correlated with the size of the primary tumor. The <em>interleukin</em> 6 and tumor necrosis factor-alpha (TNF-alpha) values of these patients have been previously published and are included in the survival analysis. Univariate analysis showed that bFGF, ICAM-1, <em>interleukin</em> 6, and TNF-alpha, before treatment, were prognostic factors. In multivariate analysis for proportional hazard regression, only TNF-alpha was an independent prognostic indicator, with a normal plasma TNF-alpha being highly predictive for a good prognosis in patients with untreated renal cell carcinoma.