OBJECTIVE

To conduct in vitro studies as well as a phase 2 clinical trial in patients with smoldering or indolent multiple myeloma to determine if interleukin 1 (IL-1) inhibitors can delay or prevent active myeloma.

METHODS

Stromal cells were cocultured with IL-1beta-expressing myeloma cells in the presence of dexamethasone, IL-1 receptor antagonist (IL-1Ra), or both. Levels of interleukin 6 (IL-6) and of apoptosis were also quantified. Between November 19, 2002, and May 24, 2007, 47 patients were enrolled in the study and subsequently treated with IL-1Ra. In 25 (53%) of the 47 study patients, low-dose dexamethasone (20 mg/wk) was added. The primary end point was progression-free survival (PFS).

RESULTS

In vitro, IL-1Ra was superior to dexamethasone at inhibiting IL-6 production; maximal IL-6 inhibition and apoptosis induction were achieved by addition of both IL-1Ra and dexamethasone. In the clinical trial, 3 patients achieved a minor response to IL-1Ra alone; 5 patients achieved a partial response and 4 patients a minor response after addition of dexamethasone. Seven patients showed a decrease in the plasma cell labeling index that paralleled a decrease in high-sensitivity C-reactive protein (hs-CRP) levels. The median overall PFS was 37.5 months. The median PFS for patients without (n=12) or with (n=35) a greater than 15% decrease in 6-month vs baseline hs-CRP levels was 6 months and more than 3 years, respectively (P=.002). Disease stability was maintained in 8 patients who received therapy for more than 4 years.

CONCLUSIONS

In patients with smoldering or indolent multiple myeloma who were at risk of progression to active myeloma, treatment with IL-1 inhibitors decreased the myeloma proliferative rate and hs-CRP levels in those who responded, leading to a chronic disease state and an improved PFS.

BACKGROUND

clinicaltrials.gov identifier: NCT00635154.

OBJECTIVE

<em>Interleukin</em>-<em>15</em> (IL-<em>15</em>) is a proinflammatory, innate response cytokine that mediates pleiotropic effector function in rheumatoid arthritis (RA) inflammatory synovitis. Our objective was to study the ability of HuMax-IL<em>15</em>, a human IgG1 anti-IL-<em>15</em> monoclonal antibody, to neutralize exogenous and endogenous IL-<em>15</em> activity in vitro and to perform a phase I-II dose-escalation trial with HuMax-IL<em>15</em> in patients with active RA.

METHODS

Mononuclear cells from blood and synovial fluid (SF) of RA patients were isolated and cultured in vitro under experimental conditions involving the addition of HuMax-IL<em>15</em>. HuMax-IL<em>15</em> was administered to 30 RA patients who received no other disease-modifying antirheumatic drugs in a 12-week, dose-ascending, placebo-controlled, double-blind, phase I-II proof-of-concept study.

RESULTS

In vitro studies showed that HuMax-IL<em>15</em> suppressed proliferation and induced apoptosis in an IL-<em>15</em>-dependent cell line, BDB2, and was capable of suppressing the release of interferon-gamma by synovial fluid mononuclear cell (SFMC) cultures induced by exogenous IL-<em>15</em>. Furthermore, HuMax-IL<em>15</em> F(ab')2 fragments suppressed exogenous IL-<em>15</em>-induced CD69 expression in RA peripheral blood mononuclear cells and SFMCs, which indicates that HuMax-IL<em>15</em> can specifically neutralize several biologic effects of IL-<em>15</em> in synovial tissue in vitro. In a phase I-II clinical trial, HuMax-IL<em>15</em> was well tolerated clinically, with no significant effects on T lymphocyte subset and natural killer cell numbers. Substantial improvements in disease activity were observed according to the American College of Rheumatology criteria for 20% improvement (63% of patients), 50% improvement (38%), and 70% improvement (25%).

CONCLUSIONS

These clinical data suggest for the first time that IL-<em>15</em> could represent a novel therapeutic target in RA.

BACKGROUND

Macrophages are believed to play a central role in emphysema based largely on data from mouse models. However, the relevance of these models to smoking-related lung disease in humans is uncertain.

OBJECTIVE

We sought to comprehensively characterize the effects of smoking on gene expression in human alveolar macrophages and to compare these with effects seen in transgenic mouse models of emphysema.

METHODS

We used DNA microarrays with genomewide coverage to analyze alveolar macrophages from <em>15</em> smokers, <em>15</em> nonsmokers, and <em>15</em> subjects with asthma (disease control). Selected gene expression changes were validated by polymerase chain reaction and ELISA. Expression changes were compared with those identified by microarray analysis of <em>interleukin</em>-13-overexpressing and integrin-beta6-deficient mice, which both develop emphysema.

RESULTS

All <em>15</em> smokers shared a common pattern of macrophage gene expression that distinguished them from nonsmokers, a finding not observed in subjects with asthma. We identified 110 genes as differentially expressed in smokers despite using conservative statistical methods. Matrix metalloproteinase 12, a proteinase that plays a critical role in mouse models, was the third most highly induced gene in smokers (ninefold, p < 0.0001). However, most changes in smokers were not reflected in mouse models. One such finding was increased osteopontin expression in smokers (fourfold, p = 0.006), which was confirmed at the protein level and correlated with the degree of airway obstruction.

CONCLUSIONS

Smoking induces a remarkably consistent and distinctive pattern of alveolar macrophage activation. These studies identify aspects of mouse models that are directly relevant to human smokers and also reveal novel potential mediators of smoking-related diseases.

Chimeric antigen receptors (CARs) have been used to redirect the specificity of autologous T cells against leukemia and lymphoma with promising clinical results. Extending this approach to allogeneic T cells is problematic as they carry a significant risk of graft-versus-host disease (GVHD). Natural killer (NK) cells are highly cytotoxic effectors, killing their targets in a non-antigen-specific manner without causing GVHD. Cord blood (CB) offers an attractive, allogeneic, off-the-self source of NK cells for immunotherapy. We transduced CB-derived NK cells with a retroviral vector incorporating the genes for CAR-CD19, IL-<em>15</em> and inducible caspase-9-based suicide gene (iC9), and demonstrated efficient killing of CD19-expressing cell lines and primary leukemia cells in vitro, with marked prolongation of survival in a xenograft Raji lymphoma murine model. <em>Interleukin</em>-<em>15</em> (IL-<em>15</em>) production by the transduced CB-NK cells critically improved their function. Moreover, iC9/CAR.19/IL-<em>15</em> CB-NK cells were readily eliminated upon pharmacologic activation of the iC9 suicide gene. In conclusion, we have developed a novel approach to immunotherapy using engineered CB-derived NK cells, which are easy to produce, exhibit striking efficacy and incorporate safety measures to limit toxicity. This approach should greatly improve the logistics of delivering this therapy to large numbers of patients, a major limitation to current CAR-T-cell therapies.

CONCLUSIONS

<em>Interleukin</em>-21 (IL-21) belongs to a family of cytokines that includes IL-2, IL-4, IL-7, IL-9, and IL-<em>15</em>, all of which bind to private (or shared) receptors as well as the common cytokine receptor gamma-chain as a component. Most cytokines in this family are critically important for both the maintenance and function of T cells and B cells. The receptor for IL-21 is widely distributed on lymphohematopoietic cells, and IL-21 plays many biologic roles, including maintenance and function of CD8(+) memory T cells and natural killer cells, as well as promoting the generation of Th17 cells in the mouse. One principal non-redundant role of IL-21 is the promotion of B-cell activation, differentiation or death during humoral immune responses. Furthermore, increased IL-21 production is characteristic of certain autoimmune diseases and is likely to contribute to autoantibody production as well as pathologic features of autoimmune disease. In contrast, IL-21 may function as a co-adjuvant to enhance antibody responses and thereby facilitate host defense to malignances and infectious diseases. The critical role of IL-21 in promoting humoral immune responses makes it an important focus of potential therapeutic interventions in conditions characterized by either overproduction of pathogenic autoantibodies or under production of protective antibodies.

OBJECTIVE

Cerium oxide (CeO2) nanoparticles have been shown to protect cells in culture from lethal stress, but no protection in vivo has been reported. Cardiac-specific expression of monocyte chemoattractant protein (MCP)-1 in mice causes ischemic cardiomyopathy associated with activation of endoplasmic reticulum (ER) stress. The aim of this study was to assess the effects of CeO2 nanoparticles on cardiac function and remodeling as well as ER stress response in this murine model of cardiomyopathy.

METHODS

MCP-1 transgenic mice (MCP mice) and wild-type controls were administered intravenously <em>15</em> nmol of CeO2 nanoparticles or vehicle only twice a week for 2 weeks. Cardiac function, myocardial histology, nitrotyrosine formation, expression of cytokines, and ER stress-associated genes were evaluated.

RESULTS

Treatment with CeO2 nanoparticles markedly inhibited progressive left ventricular dysfunction and dilatation in MCP mice and caused a significant decrease in serum levels of MCP-1, C-reactive protein, and total nitrated proteins. The infiltration of monocytes/macrophages, accumulation of 3-nitrotyrosine, apoptotic cell death, and expression of proinflammatory cytokines, tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, and IL-6 in the myocardium were markedly inhibited by CeO2 nanoparticles. Expression of the key ER stress-associated genes, including glucose-regulated protein 78 (Grp78), protein disulfide isomerase (PDI), and heat shock proteins (HSP25, HSP40, HSP70), were also suppressed by CeO2 nanoparticles.

CONCLUSIONS

CeO2 nanoparticles protect against the progression of cardiac dysfunction and remodeling by attenuation of myocardial oxidative stress, ER stress, and inflammatory processes probably through their autoregenerative antioxidant properties.

Severe and fatal viral infections remain common after hematopoietic stem cell transplantation. Adoptive transfer of cytotoxic T lymphocytes (CTLs) specific for Epstein-Barr virus (EBV), cytomegalovirus (CMV), and adenoviral antigens can treat infections that are impervious to conventional therapies, but broader implementation and extension to additional viruses is limited by competition between virus-derived antigens and time-consuming and laborious manufacturing procedures. We now describe a system that rapidly generates a single preparation of polyclonal (CD4(+) and CD8(+)) CTLs that is consistently specific for <em>15</em> immunodominant and subdominant antigens derived from 7 viruses (EBV, CMV, Adenovirus (Adv), BK, human herpes virus (HHV)-6, respiratory syncytial virus (RSV), and Influenza) that commonly cause post-transplant morbidity and mortality. CTLs can be rapidly produced (10 days) by a single stimulation of donor peripheral blood mononuclear cells (PBMCs) with a peptide mixture spanning the target antigens in the presence of the potent prosurvival cytokines <em>interleukin</em>-4 (IL4) and IL7. This approach reduces the impact of antigenic competition with a consequent increase in the antigenic repertoire and frequency of virus-specific T cells. Our approach can be readily introduced into clinical practice and should be a cost-effective alternative to common antiviral prophylactic agents for allogeneic hematopoietic stem cell transplant (HSCT) recipients.

Bcl family members Bcl-2, Bcl-x(L), and Mcl-1, are frequently expressed and implicated in the survival of myeloma cells. Obatoclax (GX0<em>15</em>-070) is a novel, small-molecule antagonist of the BH3-binding groove of the Bcl family of proteins. We show that GX0<em>15</em>-070 inhibits the binding of Bak to Mcl-1, up-regulates Bim, induces cytochrome c release, and activates capase-3 in human myeloma cell lines (HMCLs), confirming the predicted mechanism of action. Consequently, GX0<em>15</em>-070 potently inhibited the viability of <em>15</em> of 16 HMCLs (mean IC(50) of 246 nM), including those resistant to melphalan and dexamethasone. In combination studies, GX0<em>15</em>-070 enhanced the antimyeloma activity induced by melphalan, dexamethasone, or bortezomib. Sensitivity to GX0<em>15</em>-070 correlated with the absence or near absence of Bcl-x(L). Coculture with <em>interleukin</em>-6 or adherence to bone marrow stroma conferred modest resistance; however, it did not overcome GX0<em>15</em>-070-induced cytotoxicity. Of importance, GX0<em>15</em>-070 as a single agent induced potent cytotoxic responses against patient-derived tumor cells. GX0<em>15</em>-070 inhibited normal bone marrow-derived colony formation; however, cytotoxicity to human blood lymphocytes was not observed. Taken together, these studies describe a novel BH3 mimic with selectivity for Mcl-1, and support the therapeutic application of GX0<em>15</em>-070 for diverse neoplasias including multiple myeloma.

Resting autoreactive T cells are present in the circulation of normal individuals without pathologic consequences. In autoimmune animal models, stimulation of these self-reactive T cells in the presence of costimulatory molecules B7-1 results in T cell-mediated autoimmune disease, whereas B7-2 stimulation generates regulatory autoreactive T cells that abrogate disease severity. Thus, reactivation in the brain of myelin-autoreactive T cells by antigen with costimulatory molecules may be a critical event in the pathophysiology of multiple sclerosis (MS), a putative autoimmune disease of central nervous system (CNS) myelin. We investigated the expression of cytokines and costimulatory molecules in a panel of 41 histologically characterized CNS specimens from <em>15</em> MS and 10 control cases using semiquantitative reverse transcriptase-polymerase chain reaction and immunocytochemistry. In four cases, vascular CNS infarcts with inflammation were compared with MS plaques from the same brain. We observed increased expression of B7-1 and <em>interleukin</em> (IL) 12p40 in acute MS plaques, particularly from early disease cases but not in inflammatory infarcts. B7-1 staining was localized predominantly to the lymphocytes in perivenular inflammatory cuffs but not the parenchyma. In contrast, B7-2 was expressed predominantly on macrophages both in MS lesions of varied time duration and in inflammatory infarcts. These findings indicate that an early event in the initiation of MS involves upregulation of B7-1 and IL-12, resulting in conditions that maximally stimulate T cell activation and induction of T helper 1-type immune responses.

Plants and animals mediate early steps of the innate immune response through pathogen recognition receptors (PRRs). PRRs commonly associate with or contain members of a monophyletic group of kinases called the <em>interleukin</em>-1 receptor-associated kinase (IRAK) family that include Drosophila Pelle, human IRAKs, rice XA21 and Arabidopsis FLS2. In mammals, PRRs can also associate with members of the receptor-interacting protein (RIP) kinase family, distant relatives to the IRAK family. Some IRAK and RIP family kinases fall into a small functional class of kinases termed non-RD, many of which do not autophosphorylate the activation loop. We surveyed the yeast, fly, worm, human, Arabidopsis, and rice kinomes (3,723 kinases) and found that despite the small number of non-RD kinases in these genomes (9%-29%), 12 of <em>15</em> kinases known or predicted to function in PRR signaling fall into the non-RD class. These data indicate that kinases associated with PRRs can largely be predicted by the lack of a single conserved residue and reveal new potential plant PRR subfamilies.

<b>Objective:</b> To analyze the clinical characteristics of 2019 novel coronavirus (2019-nCoV) pneumonia and to investigate the correlation between serum inflammatory cytokines and severity of the disease. <b>Methods:</b> 29 patients with 2019-ncov admitted to the isolation ward of Tongji hospital affiliated to Tongji medical college of Huazhong University of Science and Technology in January 2020 were selected as the study subjects. Clinical data were collected and the general information, clinical symptoms, blood test and CT imaging characteristics were analyzed. According to the relevant diagnostic criteria, the patients were divided into three groups: mild (<em>15</em> cases), severe (9 cases) and critical (5 cases). The expression levels of inflammatory cytokines and other markers in the serum of each group were detected, and the changes of these indicators of the three groups were compared and analyzed, as well as their relationship with the clinical classification of the disease. <b>Results:</b> (1) The main symptoms of 2019-nCoV pneumonia was fever (28/29) with or without respiratory and other systemic symptoms. Two patients died with underlying disease and co-bacterial infection, respectively. (2) The blood test of the patients showed normal or decreased white blood cell count (23/29), decreased lymphocyte count (20/29), increased hypersensitive C reactive protein (hs-CRP) (27/29), and normal procalcitonin. In most patients,serum lactate dehydrogenase (LDH) was significantly increased (20/29), while albumin was decreased(<em>15</em>/29). Alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (Tbil), serum creatinine (Scr) and other items showed no significant changes. (3) CT findings of typical cases were single or multiple patchy ground glass shadows accompanied by septal thickening. When the disease progresses, the lesion increases and the scope expands, and the ground glass shadow coexists with the solid shadow or the stripe shadow. (4) There were statistically significant differences in the expression levels of <em>interleukin</em>-2 receptor (IL-2R) and IL-6 in the serum of the three groups (P<0.05), among which the critical group was higher than the severe group and the severe group was higher than the mildgroup. However, there were no statistically significant differences in serum levels of tumor necrosis factor-alpha (TNF-α), IL-1, IL-8, IL-10, hs-CRP, lymphocyte count and LDH among the three groups (P>0.05). <b>Conclusion:</b> The clinical characteristics of 2019-nCoV pneumonia are similar to those of common viral pneumonia. High resolution CT is of great value in the differential diagnosis of this disease. The increased expression of IL-2R and IL-6 in serum is expected to predict the severity of the 2019-nCoV pneumonia and the prognosis of patients.

OBJECTIVE

To evaluate levels of biomarkers in preclinical rheumatoid arthritis (RA) and to use elevated biomarkers to develop a model for the prediction of time to future diagnosis of seropositive RA.

METHODS

Stored samples obtained from 73 military cases with seropositive RA prior to RA diagnosis and from controls (mean 2.9 samples per case; samples collected a mean of 6.6 years prior to diagnosis) were tested for rheumatoid factor (RF) isotypes, anti-cyclic citrullinated peptide (anti-CCP) antibodies, 14 cytokines and chemokines (by bead-based assay), and C-reactive protein (CRP).

RESULTS

Preclinical positivity for anti-CCP and/or ≥2 RF isotypes was >96% specific for future RA. In preclinical RA, levels of the following were positive in a significantly greater proportion of RA cases versus controls: <em>interleukin</em>-1α (IL-1α), IL-1β, IL-6, IL-10, IL-12p40, IL-12p70, IL-<em>15</em>, fibroblast growth factor 2, flt-3 ligand, tumor necrosis factor α, interferon-γ-inducible 10-kd protein, granulocyte-macrophage colony-stimulating factor, and CRP. Also, increasing numbers of elevated cytokines/chemokines were present in cases nearer to the time of diagnosis. RA patients who were ≥40 years old at diagnosis had a higher proportion of samples positive for cytokines/chemokines 5-10 years prior to diagnosis than did patients who were <40 years old at diagnosis (P < 0.01). In regression modeling using only case samples positive for autoantibodies highly specific for future RA, increasing numbers of cytokines/chemokines were predictive of decreased time to diagnosis, and the predicted time to diagnosis based on cytokines/chemokines was longer in older compared with younger cases.

CONCLUSIONS

Levels of autoantibodies, cytokines/chemokines, and CRP are elevated in the preclinical period of RA development. In preclinical autoantibody-positive cases, the number of elevated cytokines/chemokines is predictive of the time of diagnosis of future RA in an age-dependent manner.

There is substantial evidence that host genetic factors are important in determining susceptibility to mycobacteria. Several different techniques have been used to identify the genes involved. Studies of an inbred strain of mice with increased susceptibility to mycobacteria, salmonella and leishmania infections led to the identification of the natural resistance-associated macrophage protein gene (Nramp1). Case-control studies have confirmed the importance of the human equivalent of this gene, NRAMP1, and have also suggested that the major histocompatibility complex and vitamin-D receptor genes may be involved in determining human susceptibility to mycobacteria. Studies of individuals with the rare condition of increased susceptibility to disseminated bacille Calmette-Guerin and other atypical mycobacterial infections have identified several abnormalities in the genes encoding the interferon gamma receptor (IFNgammaR) ligand binding chain, IFNgammaR signal transduction chain, IFNgamma signal transduction and activation of transcription-1, <em>interleukin</em> 12 receptor beta1 subunit and <em>interleukin</em> 12 p40 subunit. A genome-wide linkage study has been performed to identify genes exerting a major effect on tuberculosis susceptibility in the general population. Linkages were found to markers on chromosomes <em>15</em> and X. Studies to identify the genes responsible are in progress.

The antitumor effect of adoptively transferred tumor-specific cytotoxic T lymphocytes (CTLs) is impaired by the limited capacity of these cells to expand within the tumor microenvironment. Administration of <em>interleukin</em> 2 (IL-2) has been used to overcome this limitation, but the systemic toxicity and the expansion of unwanted cells, including regulatory T cells, limit the clinical value of this strategy. To discover whether transgenic expression of lymphokines by the CTLs themselves might overcome these limitations, we evaluated the effects of transgenic expression of IL-2 and IL-<em>15</em> in our model of Epstein Barr Virus-specific CTLs (EBV-CTLs). We found that transgenic expression of IL-2 or IL-<em>15</em> increased the expansion of EBV-CTLs both in vitro and in vivo in a severe combined immunodeficiency disease (SCID) mouse model and enhanced antitumor activity. Although the proliferation of these cytokine genes transduced CTLs remained strictly antigen dependent, clinical application of this approach likely requires the inclusion of a suicide gene to deal with the potential development of T-cell mutants with autonomous growth. We found that the incorporation of an inducible caspase-9 suicide gene allowed efficient elimination of transgenic CTLs after exposure to a chemical inducer of dimerization, thereby increasing the safety and feasibility of the approach.

It has been more than <em>15</em> years since the identification of individual <em>interleukin</em>-27 (IL-27) and IL-27 receptor components. The last decade has seen the description of the signaling pathways engaged by IL-27, and an appreciation has emerged that this cytokine can modulate the intensity and duration of many classes of T cell responses. Here we provide an overview of the immunobiology of IL-27 and review advances in understanding the functions of individual IL-27 and IL-27 receptor subunits and the role of IL-27 in dictating the balance between protective and pathological immunity. Additionally, this cytokine has been proposed as a therapy to modify inflammatory conditions or to promote antitumor responses, and situations where experimental and clinical data sets implicate IL-27 in the outcome of disease are highlighted.

Enriched populations of human microglial cells were isolated from mixed cell cultures prepared from embryonic human telencephalon tissues. Human microglial cells exhibited cell type-specific antigens for macrophage-microglia lineage cells including CD11b (Mac-1), CD68, B7-2 (CD86), HLA-ABC, HLA-DR and ricinus communis aggulutinin lectin-1 (RCA-1), and actively phagocytosed latex beads. Gene expression and protein production of cytokines, chemokines and cytokine/chemokine receptors were investigated in the purified populations of human microglia. Normal unstimulated human microglia expressed constitutively mRNA transcripts for <em>interleukin</em>- 1beta (IL-1beta) -6, -8, -10, -12, -<em>15</em>, tumor necrosis factor-alpha (TNF-alpha), macrophage inflammatory protein-1alpha (MIP-1alpha), MIP-1beta, and monocyte chemoattractant protein-1 (MCP-1), while treatment with lipopolysaccharide (LPS) or amyloid beta peptides (Abeta) led to increased expression of mRNA levels of IL-8, IL-10, IL-12, TNF-alpha, MIP-1alpha, MIP-1beta, and MCP-1. Human microglia, in addition, expressed mRNA transcripts for IL-1RI, IL-1RII, IL-5R, IL-6R, IL-8R, IL-9R, IL-10R, IL-12R, IL-13R, and IL-<em>15</em>R. Enzyme-linked immunosorbent assays (ELISA) showed increased protein levels in culture media of IL-1beta, IL-8, TNF-alpha, and MIP-1alpha in human microglia following treatment with LPS or Abeta. Increased TNF-alpha release from human microglia following LPS treatment was completely inhibited with IL-10 pretreatment, but not with IL-6, IL-9, IL-12, IL-13, or transforming growth factor-beta (TGF-beta). Present results should help in understanding the basic microglial biology, but also the pathophysiology of activated microglia in neurological diseases such as Alzheimer disease, Parkinson disease, Huntington disease, amyotrophic lateral sclerosis, stroke, and neurotrauma.

<em>Interleukin</em>-6 (IL-6) is a multifunctional cytokine expressed by angiotensin II (Ang II)-stimulated vascular smooth muscle cells (VSMCs) that functions as an autocrine growth factor. In this study, we analyze the mechanism for Ang II-inducible IL-6 expression in quiescent rat VSMCs. Stimulation with the Ang II agonist Sar1 Ang II (100 nmol/L) induced transcriptional expression of IL-6 mRNA transcripts of 1.8 and 2.4 kb. In transient transfection assays of IL-6 promoter/luciferase reporter plasmids, Sar1 Ang II treatment induced IL-6 transcription in a manner completely dependent on the nuclear factor-kappaB (NF-kappaB) motif. Sar1 Ang II induced cytoplasmic-to-nuclear translocation of the NF-kappaB subunits Rel A and NF-kappaB1 with parallel changes in DNA-binding activity in a biphasic manner, which produced an early peak at <em>15</em> minutes followed by a nadir 1 to 6 hours later and a later peak at 24 hours. The early phase of NF-kappaB translocation was dependent on weak simultaneous proteolysis of the IkappaBalpha and beta inhibitors, whereas later translocation was associated with enhanced processing of the p105 precursor into the mature 50-kDa NF-kappaB1 form. Pretreatment with a potent inhibitor of IkappaBalpha proteolysis, TPCK, completely blocked Sar1 Ang IIAng II-induced NF-kappaB activation and induction of endogenous IL-6 gene expression, which indicated the essential role of NF-kappaB in mediating IL-6 expression. We conclude that Ang II is a pleiotropic regulator of the NF-kappaB transcription factor family and may be responsible for activating the expression of cytokine gene networks in VSMCs.

In the maternal decidua, natural killer (NK) cells, characterized by lack of CD16, are found in direct contact with the fetal extravillous trophoblasts (EVTs). It is yet unknown which factors contribute to the specific homing of this unique NK subset to the decidua. In this study we analyze the chemokine receptor repertoire on various NK populations derived from the peripheral blood and decidua. We show that CXCR4 and CXCR3 receptors are preferentially expressed on CD16- NK subsets derived either from the peripheral blood or the decidua and that these receptors are involved in migration of all NK subsets to their ligands. We further demonstrate in vivo that invading EVTs that eventually perform endovascular invasion express CXCL12, the ligand for CXCR4, but not ligands for CXCR3. Indeed, specific accumulation of the CD16- NK cells at the expense of CD16+ cells was observed only when in vitro migration was performed with ligands for CXCR4. Finally, incubation of the peripheral blood CD16- NK cells with cytokines present in the decidua, especially <em>interleukin</em> <em>15</em> (IL-<em>15</em>), resulted in the expression of chemokine receptor repertoire similar to that observed on decidual NK cells, suggesting an additional important regulatory effect of local decidual cytokines.

Like their normal hematopoietic stem cell counterparts, leukemia stem cells (LSCs) in chronic myelogenous leukemia (CML) and acute myeloid leukemia (AML) are presumed to reside in specific niches in the bone marrow microenvironment (BMM) and may be the cause of relapse following chemotherapy. Targeting the niche is a new strategy to eliminate persistent and drug-resistant LSCs. CD44 (refs. 3,4) and <em>interleukin</em>-6 (ref. 5) have been implicated previously in the LSC niche. Transforming growth factor-β1 (TGF-β1) is released during bone remodeling and plays a part in maintenance of CML LSCs, but a role for TGF-β1 from the BMM has not been defined. Here, we show that alteration of the BMM by osteoblastic cell-specific activation of the parathyroid hormone (PTH) receptor attenuates BCR-ABL1 oncogene-induced CML-like myeloproliferative neoplasia (MPN) but enhances MLL-AF9 oncogene-induced AML in mouse transplantation models, possibly through opposing effects of increased TGF-β1 on the respective LSCs. PTH treatment caused a <em>15</em>-fold decrease in LSCs in wild-type mice with CML-like MPN and reduced engraftment of immune-deficient mice with primary human CML cells. These results demonstrate that LSC niches in CML and AML are distinct and suggest that modulation of the BMM by PTH may be a feasible strategy to reduce LSCs, a prerequisite for the cure of CML.

OBJECTIVE

The proinflammatory cytokines interleukin (IL)-1beta and IL-6 are supposed to be involved in various cardiovascular diseases including reperfusion injury and cardiac hypertrophy.

RESULTS

In the present study, we have examined the cytokine expression from 3 h up to 12 weeks after permanent coronary artery occlusion in rats. In the first 3-12 h, there was a strong induction in IL-1beta and IL-6 mRNA expression in the infarct area (up to 50-fold) as well as in the non-infarcted myocardium (up to 15-fold). From day 3 onwards the cytokine expression was not significantly altered compared to sham-operated controls. In addition, the expression of C/AATT-enhancer binding protein-beta was about fourfold elevated in the first hours after myocardial infarction, but not thereafter. Furthermore, the expression of gp130 and IL-6 receptor increased significantly in the infarct area. The elevation in cytokine expression preceded the increase in matrix-metalloproteinase-9 in the infarct area as well as the increase in ANP and collagen expression in the non-infarcted myocardium.

CONCLUSIONS

We suggest that IL-6 and IL-1beta act synergistically in promoting resorption of the necrotic tissue, matrix remodeling and wound healing. Furthermore, they may be involved in the early induction of fibrosis and compensatory cardiac hypertrophy of the non-infarcted myocardium, but seem not to play a key role in long-term cardiac remodeling in chronic heart failure after myocardial infarction.

BACKGROUND

Fruits and vegetables, foods rich in flavonoids and antioxidants, have been associated with lower risk of stroke, coronary heart disease, and markers of inflammation and oxidative stress in adults. Markers of inflammation and oxidative stress are predictors of coronary heart disease risk; however, it is unknown whether these markers are related to dietary flavonoid and antioxidant intake in youth.

OBJECTIVE

To determine whether greater intakes of fruit and vegetables, antioxidants, folate, and total flavonoids were inversely associated with markers of inflammation and oxidative stress in 285 adolescent boys and girls aged 13 to 17 years.

METHODS

In this cross-sectional study conducted between February 1996 and January 2000, diet was assessed by a 127-item food frequency questionnaire. Height and weight measurements were obtained and a fasting blood sample drawn. Spearman partial correlation analyses evaluated the relation of intakes of fruit and vegetables, antioxidants, folate, and flavonoids with markers of inflammation (C-reactive protein, <em>interleukin</em>-6, tumor necrosis factor-alpha, and <em>15</em>-keto-dihydro-PGF(2alpha) metabolite and oxidative stress (urinary 8-iso prostaglandin F(2alpha), an F(2)-isoprostane), adjusting for age, sex, race, Tanner stage, energy intake, and body mass index.

RESULTS

Urinary F(2)-isoprostane was inversely correlated with intakes of total fruit and vegetables, vitamin C, beta carotene, and flavonoids. Serum C-reactive protein was significantly inversely associated with intakes of fruit (r=-0.19; P=0.004), vitamin C (r=-0.13, P=0.03), and folate (r=-0.18; P=0.004). Serum <em>interleukin</em>-6 was inversely associated with intakes of legumes, vegetables, beta carotene, and vitamin C. Serum tumor necrosis factor-alpha was inversely associated with beta carotene (r=-0.14, P=0.02) and luteolin (r=-0.<em>15</em>, P=0.02).

CONCLUSIONS

Study results show that the beneficial effects of fruit and vegetable intake on markers of inflammation and oxidative stress are already present by early adolescence and provide support for the Dietary Guidelines for Americans "to consume five or more servings per day" of fruits and vegetables to promote beneficial cardiovascular health.

Tissue-resident memory T (Trm) cells contribute to local immune protection in non-lymphoid tissues such as skin and mucosa, but little is known about their transcriptional regulation. Here we showed that CD8(+)CD103(+) Trm cells, independent of circulating memory T cells, were sufficient for protection against infection and described molecular elements that were crucial for their development in skin and lung. We demonstrated that the T-box transcription factors (TFs) Eomes and T-bet combined to control CD8(+)CD103(+) Trm cell formation, such that their coordinate downregulation was crucial for TGF-β cytokine signaling. TGF-β signaling, in turn, resulted in reciprocal T-box TF downregulation. However, whereas extinguishment of Eomes was necessary for CD8(+)CD103(+) Trm cell development, residual T-bet expression maintained cell surface <em>interleukin</em>-<em>15</em> (IL-<em>15</em>) receptor β-chain (CD122) expression and thus IL-<em>15</em> responsiveness. These findings indicate that the T-box TFs control the two cytokines, TGF-β and IL-<em>15</em>, which are pivotal for CD8(+)CD103(+) Trm cell development and survival.

<em>Interleukin</em> 21 (IL-21) is a newly described cytokine with homology to IL-4 and IL-<em>15</em>. They belong to a cytokine family that uses the common gamma chain for signaling but also have their private high-affinity receptors. Since it is well known that IL-4 modulates differentiation and activation of dendritic cells (DCs), we analyzed effects of IL-21 compared with IL-<em>15</em> on DC differentiation, maturation, and function. Here we show that DCs generated with granulocyte-macrophage colony-stimulating factor (GMCSF) in the presence of IL-21 (IL-21DCs) differentiated into phenotypically and functionally altered DCs characterized by reduced major histocompatibility complex class II (MHCII) expression, high antigen uptake, and low stimulatory capacity for T-cell activation in vitro. Additionally, IL-21DCs completely failed to induce antigen (Ag)-specific T-cell mediated contact hypersensitivity. Furthermore, IL-21 blocked lipopolysaccharide (LPS)-induced activation and maturation of DCs, which was not mediated by release of the anti-inflammatory cytokine IL-10. In contrast, when supplementing GMCSF with IL-<em>15</em>, DCs differentiated into mature antigen-presenting cells (APCs) with low antigen uptake and highly significant increased capacities to stimulate T cells in vitro and in vivo. Taken together, these results identify a dichotomous action of these structurally related cytokines on DCs, establishing IL-21 as inhibitory cytokine on DC activation and IL-<em>15</em> as potent stimulator of DC function, making both cytokines interesting targets for therapeutic manipulation of DC-induced immune reactions.

OBJECTIVE

To determine the safety, dose-limiting toxicity, maximum tolerated dose, and pharmacokinetic and pharmacodynamic profiles of the novel hydroxamate histone deacetylase inhibitor belinostat (previously named PXD101) in patients with advanced refractory solid tumors.

METHODS

Sequential dose-escalating cohorts of three to six patients received belinostat administered as a 30-min i.v. infusion on days 1 to 5 of a 21-day cycle. Pharmacokinetic variables were evaluated at all dose levels. Pharmacodynamic measurements included acetylation of histones extracted from peripheral blood mononuclear cells, caspase-dependent cleavage of cytokeratin-18, and interleukin-6 levels.

RESULTS

Forty-six patients received belinostat at one of six dose levels (150-1,200 mg/m(2)/d). Dose-limiting toxicities were grade 3 fatigue (one patient at 600 mg/m(2); one patient at 1,200 mg/m(2)), grade 3 diarrhea combined with fatigue (one patient at 1,200 mg/m(2)), grade 3 atrial fibrillation (one patient at 1,200 mg/m(2); one patient at 1,000 mg/m(2)), and grade 2 nausea/vomiting leading to inability to complete a full 5-day cycle (two patients at 1,000 mg/m(2)). The maximum tolerated dose was 1,000 mg/m(2)/d. I.v. belinostat displayed linear pharmacokinetics with respect to C(max) and AUC. The intermediate elimination half-life was 0.3 to 1.3 h and was independent of dose. Histone H4 hyperacetylation was observed after each infusion and was sustained for 4 to 24 h in a dose-dependent manner. Increases in interleukin-6 levels were detected following belinostat treatment. Stable disease was observed in a total of 18 (39%) patients, including 15 treated for>> or =4 cycles, and this was associated with caspase-dependent cleavage of cytokeratin-18. Of the 24 patients treated at the maximum tolerated dose (1,000 mg/m(2)/d), 50% achieved stable disease.

CONCLUSIONS

I.v. belinostat is well tolerated, exhibits dose-dependent pharmacodynamic effects, and has promising antitumor activity.