Potentially toxic amyloid beta-peptide (Abeta) in Alzheimer's disease (AD) is generated from a family of Abeta-containing precursor proteins (APP), which is regulated via the 5'-untranslated region (5'-UTR) of its mRNA. We analyzed 5'-UTRs of the APP superfamily, including amyloid plaque-forming and non-amyloid plaque-forming species, and of prions (<em>27</em> different DNA sequences). A "CAGA" sequence proximal to the "ATG" start codon was present in a location unique to APP genes of amyloid plaque-forming species and absent in all other genes surveyed. This CAGA box is immediately upstream of an <em>interleukin</em>-1-responsive element (acute box). In addition, the proximal CAGA box is predicted to appear on a stem-loop structure in both human and guinea pig APP mRNA. This stem-loop is part of a predicted bulge-loop that encompasses a known iron regulatory element (IRE). Electrophoretic mobility shift with segments of the APP 5'-UTR showed that a region with the proximal CAGA sequence binds nuclear proteins, and this UTR fragment is active in a reporter gene functional assay. Thus, the 5'-UTR in the human APP but not those of APP-like proteins contains a specific region that may participate in APP regulation and may determine a more general model for amyloid generation as seen in AD. The 5'-UTR of human APP contains several interesting control elements, such as an acute box element, a CAGA box, an IRE, and a transforming growth factor-beta-responsive element, that could control APP expression and provide suitable and specific drug targets for AD.

In neutrophils, growth-related protein-alpha (CXCL1) and <em>interleukin</em>-8 (CXCL8), are potent chemoattractants (Cytokine 14:<em>27</em>-36, 2001; Biochemistry 42:2874-2886, 2003) and can stimulate myeloperoxidase release via activation of the G protein-coupled receptors CXCR1 and CXCR2. The role of CXCR1 and CXCR2 in the pathogenesis of inflammatory responses has encouraged the development of small molecule antagonists for these receptors. The data presented herein describe the pharmacology of 2-hydroxy-N,N-dimethyl-3-{2-[[(R)-1-(5-methyl-furan-2-yl)-propyl]amino]-3,4-dioxo-cyclobut-1-enylamino}-benzamide (Sch5<em>27</em>123), a novel antagonist of both CXCR1 and CXCR2. Sch5<em>27</em>123 inhibited chemokine binding to (and activation of) these receptors in an insurmountable manner and, as such, is categorized as an allosteric antagonist. Sch5<em>27</em>123 inhibited neutrophil chemotaxis and myeloperoxidase release in response to CXCL1 and CXCL8 but had no effect on the response of these cells to C5a or formyl-methionyl-leucyl-phenylalanine. The pharmacological specificity of Sch5<em>27</em>123 was confirmed by testing in a diversity profile against a panel of enzymes, channels, and receptors. To measure compound affinity, we characterized [(3)H]Sch5<em>27</em>123 in both equilibrium and nonequilibrium binding analyses. Sch5<em>27</em>123 binding to CXCR1 and CXCR2 was both saturable and reversible. Although Sch5<em>27</em>123 bound to CXCR1 with good affinity (K(d) = 3.9 +/- 0.3 nM), the compound is CXCR2-selective (K(d) = 0.049 +/- 0.004 nM). Taken together, our data show that Sch5<em>27</em>123 represents a novel, potent, and specific CXCR2 antagonist with potential therapeutic utility in a variety of inflammatory conditions.

Amiodarone-induced thyrotoxicosis (AIT) occurs both in abnormal thyroid glands (nodular goiter, latent Graves' disease) (type I AIT) or in apparently normal thyroid glands (type II AIT). Differentiation of the two forms is crucial, because type I AIT responds well to methimazole and potassium perchlorate combined treatment, whereas type II AIT is effectively managed by glucocorticoids. Differential diagnosis is often difficult, although thyroid radioactive iodine uptake is usually low-to-normal in type I and low-suppressed in type II, and serum <em>interleukin</em>-6 levels are normal/slightly elevated in type I, markedly elevated in type II. Color flow Doppler sonography (CFDS) is a technique that shows intrathyroidal blood flow and provides real-time information on thyroid morphology and hyperfunction. To investigate the usefulness of CFDS in differentiating the two types of AIT, <em>27</em> consecutive AIT patients, 11 type I and 16 type II, were evaluated by CFDS before starting antithyroid treatment. Gender, age, severity of thyrotoxicosis, and cumulative amiodarone dose were similar in the two groups. All type II AIT patients had a CFDS pattern 0 (ie, absent vascularity), in agreement with the pathogenesis of the disease, due to thyroid damage. Likewise, nine patients with subacute thyroiditis, another destructive process of the thyroid gland, also had a CFDS pattern 0. Eleven patients with type I AIT had a CFDS pattern ranging from pattern I (presence of parenchymal blood flow with patchy uneven distribution) (7 patients, 64%) to pattern II (ie, mild increase of color flow Doppler signal with patchy distribution) (1 patient, 9%) and pattern III (markedly increased color flow Doppler signal with diffuse homogeneous distribution)(3 patients, <em>27</em>%), similar to that found in patients with untreated Graves' disease patients, thus indicating a hyper-functioning gland. Control subjects and euthyroid patients under long-term amiodarone treatment had absent thyroid hypervascularity and a CFDS pattern 0. These findings demonstrate that CFDS distinguishes type I and II AIT. Because of its rapidity and noninvasive features, CFDS represents a valuable tool for a quick differentiation between the two types of AIT. This can avoid any delay in initiating the appropriate treatment for a rapid control of thyrotoxicosis in patients whose tachyarrhythmias or other cardiac disorders make thyroid hormone excess extremely deleterious.

Some spontaneous preterm deliveries (PTD) are caused by occult infections of the fetal membranes (histologic chorioamnionitis [HCA]). High levels of infection-related markers, including some cytokines, sampled from maternal circulation in mid-pregnancy have been linked to PTD, but whether these specifically identify HCA has not been established. We have tested associations between 13 Th1, Th2 and Th17 cytokines and PTD with and without HCA in a prospective cohort study. The study sample included 926 Pregnancy Outcomes and Community Health Study subcohort women; women with medically indicated PTD or incomplete data excluded. A panel of cytokines was assessed using a multiplex assay in maternal plasma collected at 15-<em>27</em> weeks of gestation. Severe HCA was scored by a placental pathologist blinded to clinical variables. Multivariable polytomous logistic regression was used to estimate adjusted odds ratios (OR) per 1 standard deviation (S.D.) increase in cytokine levels using a 5 level outcome variable: PTD <35 weeks with HCA, PTD <35 weeks without HCA, PTD 35-36 weeks with HCA, PTD 35-36 weeks without HCA, and term (referent). <em>Interleukin</em> (IL)-1beta, IL-2, IL-12, interferon-gamma, IL-4, IL-6 and transforming growth factor-beta were all significantly associated with PTD <35 weeks with HCA, with ORs of 1.6-2.3 per S.D. increase. None of these were associated with PTD <35 weeks without HCA or PTD 35-36 weeks with HCA. Although the tissues of origin of circulating cytokines are unclear, the observed elevations across many cytokines among women who later delivered <35 weeks with HCA may represent a robust immune response to infection within gestational tissues. These results suggest that women with HCA could be identified using relatively non-invasive means.

Gram-negative sepsis is caused by endotoxin-induced release of tumor necrosis factor (TNF) and other cytokines. HA-1A is a human monoclonal antibody that binds specifically to endotoxin. HA-1A should prevent death in endotoxemic patients and reduce serum levels of TNF and <em>interleukin</em>-6 (IL-6). This hypothesis was tested in 82 septic patients who were randomly allocated to receive a single intravenous 100-mg dose of HA-1A or placebo. Pretreatment endotoxemia was detected in <em>27</em> patients (33%). Death occurred within 28 days of treatment in 8 (73%) of 11 placebo recipients and in 5 (31%) of 16 HA-1A recipients (P = .02). The median decrease in serum TNF level 24 h after treatment was 12 ng/L in patients given HA-1A and 0 ng/L in placebo recipients (n = 65; P = .04). For IL-6, this was 204 ng/L in patients given HA-1A and 44 ng/L in placebo recipients (n = 67; P = .4). Thus, HA-1A reduces mortality in septic patients with endotoxemia and lowers serum TNF levels.

<em>Interleukin</em> 1 (IL-1), IL-6, and tumour necrosis factor (TNF) alpha are pleiotropic cytokines produced predominantly by macrophages which have been implicated in the pathogenesis of rheumatoid arthritis (RA). Sulphasalazine has been shown to have disease modifying properties and to inhibit the production of cytokines in vitro. To evaluate the effect of sulphasalazine on cytokine production in vivo, serum cytokine levels were measured in a group of patients with RA entered into a randomised controlled trial. Serum levels of IL-1 alpha, IL-1 beta, IL-6, and TNF alpha were measured at baseline and at two monthly intervals for six months in 17 patients receiving sulphasalazine and in 22 patients treated with placebo. The two groups of patients had a similar age and sex distribution, had had RA for less than a year, had no joint erosions, and had not been treated previously with any other disease modifying drugs. In the 39 patients studied IL-1 alpha was detected >> 0.1 ng/ml) at baseline in 14 patients (median 0.24 ng/ml), IL-1 beta in 25 patients (median 1.0 ng/ml), TNF alpha in <em>27</em> patients (median 1.2 ng/ml), and IL-6 in 33 patients (median 0.44 ng/ml). In the group treated with sulphasalazine there was a progressive and significant decline in serum IL-1 alpha, IL-1 beta, and TNF alpha levels over the six month period (median levels at six months were < 0.1, 0.12, and 0.44 ng/ml respectively). <em>Interleukin</em> 6 levels were significantly reduced only at the four month time point (median level of 0.23 ng/ml). These reductions were associated with improvements in clinical and laboratory measures of disease activity. In contrast patients receiving the placebo showed no changes in serum cytokine levels and no improvement in clinical and laboratory indices of disease activity. These results suggest that sulphasalazine may exert its disease modifying effect partly by suppressing cytokine production in vivo.

To evaluate the role of <em>interleukin</em> (IL)-8 in meningococcal disease, a solid-phase double-ligand ELISA was used to quantitate IL-8 in sera and cerebrospinal fluid (CSF) from patients with meningococcal meningitis, bacteremia, or both with or without septic shock. IL-8 was demonstrated in sera from 28 of 62 patients; levels were significantly higher in patients with septic shock without meningitis (median, 36.1 ng/mL) than in patients with other manifestations (median, < 0.02 ng/mL), and 4 of 5 patients who died had high levels. IL-8 was detected in all <em>27</em> CSF samples. Serum IL-8 levels correlated highly significantly with those of IL-6 (r = .83) and tumor necrosis factor (TNF; r = .64), while the correlations between corresponding CSF levels were less pronounced (r = .43 and r = .38, respectively) but still significant. Serum IL-8 levels were highest in patients with a symptom history < 12 h. The elimination rate of IL-8 from serum varied and was similar to that of IL-6 and TNF. IL-8 appears to participate in the complex cytokine network during the initial phase of systemic meningococcal infections.

Lipopolysaccharide from Porphyromonas gingivalis (P-LPS), an important pathogenic bacterium, is closely associated with inflammatory destruction of periodontal tissues. P-LPS induces the release of cytokines and local factors from inflammatory cells, stimulates osteoclastic-cell differentiation, and causes alveolar bone resorption. However, the effect of P-LPS on osteoblastic-cell differentiation remains unclear. In this study, we investigated the effect of P-LPS extract prepared by the hot-phenol-water method, on the differentiation of primary fetal rat calvaria (RC) cells, which contain a subpopulation of osteoprogenitor cells, into osteoblastic cells. P-LPS extract significantly inhibited bone nodule (BN) formation and the activity of alkaline phosphatase (ALPase), an osteoblastic marker, in a dose-dependent manner (0 to 100 ng of P-LPS extract per ml). P-LPS extract (100 ng/ml) significantly decreased BN formation to <em>27</em>% of the control value and inhibited ALPase activity to approximately 60% of the control level on days 10 to 21 but did not affect RC cell proliferation and viability. P-LPS extract time-dependently suppressed the expression of ALPase mRNA, with an inhibitory pattern similar to that of enzyme activity. The expression of mRNAs for osteocalcin and osteopontin, matrix proteins related to bone metabolism, was markedly suppressed by P-LPS extract. Furthermore, P-LPS extract increased the expression of mRNAs for CD14, LPS receptor, and <em>interleukin</em>-1beta in RC cells. These results indicate that P-LPS inhibits osteoblastic-cell differentiation and suggest that LPS-induced bone resorption in periodontal disease may be mediated by effects on osteoblastic as well as osteoclastic cells.

Okadaic acid, a phosphatase inhibitor from a marine organism, mimics tumor necrosis factor/<em>interleukin</em>-1 (TNF/IL-1) in inducing changes in early cellular protein phosphorylation. A total of approximately 116 proteins exhibit significant and concordant changes in phosphorylation or dephosphorylation within 15 min in human fibroblasts activated by either okadaic acid, TNF, or IL-1. The fidelity of this mimicry by okadaic acid extends to the phosphorylation of the <em>27</em> hsp complex, stathmin, eIF-4E, myosin light chain, nucleolin, epidermal growth factor receptor, and other cdc2-kinase substrates (c-abl, RB, and p53). The okadaic acid-induced pattern of protein phosphorylation is distinct from that observed in cells treated with phorbol 12-myristate 13-acetate or with ligands like epidermal growth factor, cyclic AMP agonists, bradykinin, or interferons. Like TNF, okadaic acid also induces the transcription of immediate early response genes like c-jun and Egr-1 as well as the <em>interleukin</em>-6 genes. The overall early effects of okadaic acid uniquely parallel those of TNF/IL-1 and not those of other cytokines or ligands. Regulation of protein phosphatase inhibition is discussed as a mechanism for TNF/IL-1 signal transduction.

In order to evaluate the efficacy of dexamethasone (dex) in reducing the toxicity of therapy with lymphokine-activated killer (LAK) cells and <em>interleukin</em>-2 (IL-2), we treated six patients receiving this form of immunotherapy with intravenous (IV) dex, 4 mg every six hours. Compared with a control group of <em>27</em> patients not receiving dex with their immunotherapy, these corticosteroid-treated patients were able to tolerate the administration of more IL-2, yet experienced significantly less toxicity. Dyspnea, confusion, fever, mean peak serum creatinine, and bilirubin levels during treatment were significantly reduced in corticosteroid-treated patients, with a corresponding decrease in pruritus in this group as well. Overall weight gain was not different between groups, although a curtailment of weight gain temporally related to dex treatment was seen in some patients. Hematologic side effects, including anemia, eosinophilia, and thrombocytopenia, were not reduced by dex. These results suggest that dex can inhibit at least some of the toxic side effects of LAK cell and IL-2 therapy. Because of the concern that the therapeutic effect may also be abrogated, future studies combining corticosteroids with this form of immunotherapy should be undertaken with caution.

OBJECTIVE

To study the production of interleukin-13 (IL-13) in rheumatoid synovium and the effects of recombinant IL-13 on the phenotype and function of synovial fluid (SF) macrophages and T cells derived from patients with rheumatoid arthritis (RA).

METHODS

The presence of IL-13 in SF was studied using an IL-13-specific enzyme-linked immunosorbent assay (ELISA); the production of IL-13 was studied in SF mononuclear cells (SFMC) by reverse transcriptase-polymerase chain reaction. The effects of recombinant IL-13 on cytokine production by and phenotype of SFMC were evaluated using cytokine-specific ELISAs and flow cytometry, respectively. The effect of IL-13 on the proliferation of SFMC was determined by 3H-thymidine incorporation. The production and the effects of IL-13 were compared with those of IL-4.

RESULTS

IL-13 was present in 27 of 28 SF samples, and IL-13 messenger RNA (mRNA) was detectable in SFMC. Importantly, IL-13 levels were significantly higher than those of IL-4, and IL-13 protein and mRNA were expressed in several samples, although IL-4 synthesis was undetectable. Recombinant IL-13 significantly reduced the production of IL-1 beta and tumor necrosis factor alpha and the expression of CD16 and CD64 by SF macrophages, whereas the expression of HLA-DR and CD23 was increased. These effects on SF macrophages were similar to those observed with IL-4, but in contrast to IL-4, IL-13 had no growth-promoting effect on SF T cells.

CONCLUSIONS

IL-13 is consistently present in rheumatoid synovium. The ability of exogenous IL-13 to decrease the production of proinflammatory cytokines by SFMC suggests that it may have therapeutic potential in the treatment of patients with RA.

Nocturnal cytokine levels were measured serially in 12 healthy male volunteers for 12 h, including 8 h of polygraphically monitored nocturnal sleep. Plasma concentrations of <em>interleukin</em>-1 beta (IL-1 beta), <em>interleukin</em>-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha) were determined in 30-min intervals by enzyme-linked immunoadsorbent assays. In some subjects cytokines were not detectable at all. In the remaining volunteers (<em>27</em>% for IL-1 beta, 58% for IL-6 and TNF-alpha, respectively) occasional values near to the detection limits (DL) of the assays could be measured. With respect to IL-1 beta and IL-6, plasma levels above the DL were significantly more frequent during sleep than during the preceding time of wakefulness. No temporal association with NREM or REM episodes could be shown. TNF-alpha values above the DL were randomly distributed across the 12-h period investigated. It is concluded that in a considerable percentage of healthy subjects small amounts of cytokines are released at night. Release of IL-1 beta and IL-6 is temporally associated with sleep, whereas the release of TNF-alpha is not. It remains to be established whether nocturnal cytokine release reflects either an interaction between sleep and host defense mechanisms or a sleep-independent circadian rhythmicity.

T helper cells that produce <em>interleukin</em>-17 (IL-17) (Th17 cells) have recently been identified as the third distinct subset of effector T cells, the differentiation of which depends on specific transcription nuclear factor retinoic acid-related orphan nuclear receptor-gammat. Emerging data have suggested that Th17 cells play an important role in innate immunity, adaptive immunity and autoimmunity. Interestingly, there is a reciprocal relationship between Th17 cells and regulatory T cells (Treg), not only in development, but also in their effector function. Transforming growth factor (TGF)-beta induces Treg-specific transcription factor Forkhead box P3(FOXP3), while the addition of IL-6 to TGF-beta inhibits the generation of Treg cells and induces Th17 cells. It is proposed that the fine balance between Th17 and Treg cells is crucial for maintenance of immune homeostasis. In addition to IL-6, other factors such as retinoic acid, rapamycin, or cytokines (e.g., IL-2 and IL-<em>27</em>) could dictate the balance between Th17 and Treg cells. Since Treg cells play an important role in hepatic immunity with overregulation in chronic viral hepatitis and hepatic carcinoma, and inadequate inhibition in autoimmune liver diseases, graft rejection and acute liver failure, it is reasonable to assume that Th17 cells may play a reciprocal role in these diseases. Thus, future research on the Treg/Th17 balance may provide an opportunity to illustrate the pathogenesis of hepatic inflammation and to explore new therapeutic targets for immune-related liver diseases.

<em>Interleukin</em> (IL)-6 and -8 are important inflammatory cytokines in bacterial infections. Their serum and urine concentrations were measured in <em>27</em> neonates with urinary tract infection (UTI) at onset and the second week of therapy, as well as in 23 control neonates. Escherichia coli was isolated in 89% of cases. 99mTc-dimercaptosuccinic acid (99mTc-DMSA) scans were performed between the 10th and 90th days after UTI and showed pyelonephritic changes in 15 neonates (56%). Increased IL-6 and IL-8 concentrations were found in urine but not in serum within the first 24 h after presumptive diagnosis of UTI (P=.036 and.010, respectively), suggesting that the neonatal urinary tract can respond to uropathogens by producing inflammatory cytokines. Urine concentrations of IL-6 correlated with findings of renal changes in 99mTc-DMSA scans (P=.012) and thus may serve as a marker of renal parenchymal outcome. All neonates exhibited undetectable urine cytokine levels during the second week of therapy.

Autoimmune diseases such as rheumatoid arthritis are the consequence of a persistent imbalance between pro- and anti-inflammatory immune mechanisms leading to chronic inflammation. The action of several cytokines is at the basis of this complex process. This review is focused on the signalling events triggered by two major groups of cytokines, namely the IL-12 and IL-17 families, which in the past few years have been shown to have a prominent role in the pathogenesis of such diseases. In particular, we will focus on the signalling cascades set in motion by such cytokines and how this may relate to the pathogenesis of human immune and inflammatory disorders as knowledge of such cascades may help in the development of novel therapeutic approaches for such diseases.

Th1 immune response is essential in the protection against mycobacterial intracellular pathogens. Lipoproteins trigger both humoral and cellular immune responses and may be candidate protective antigens. We studied in BALB/c mice the immunogenicity and the protection offered by the recombinant <em>27</em>-kDa Mycobacterium tuberculosis lipoprotein and the corresponding DNA vaccine. Immunization with the <em>27</em>-kDa antigen resulted in high titers of immunoglobulin G1 (IgG1) and IgG2a with a typical Th1 profile and a strong delayed hypersensitivity response. A strong proliferation response was observed in splenocytes, and significant nitric oxide production and gamma interferon secretion but not <em>interleukin</em> 10 secretion were measured. Based on these criteria, the <em>27</em>-kDa antigen induced a typical Th1-type immune response thought to be necessary for protection. Surprisingly, in <em>27</em>-kDa-vaccinated mice (protein or DNA vaccines) challenged by M. tuberculosis H37Rv or BCG strains, there was a significant increase in the numbers of CFU in the spleen compared to that for control groups. Furthermore, the protection provided by BCG or other mycobacterial antigens was completely abolished once the <em>27</em>-kDa antigen was added to the vaccine preparations. This study indicates that the <em>27</em>-kDa antigen has an adverse effect on the protection afforded by recognized vaccines. We are currently studying how the <em>27</em>-kDa antigen modulates the mouse immune response.

In vivo expansion of haploidentical natural killer (NK) cell infusions with <em>interleukin</em>-2 (IL-2) can induce remission of refractory acute myeloid leukemia, but efficacy may be hampered by concurrent stimulation of host regulatory T cells. To overcome this limitation, we substituted the NK homeostatic factor IL-15 in 2 phase 1/2 trials. Forty-two patients received either intravenous (IV) (NCT01385423) or subcutaneous (SC) (NCT02395822) recombinant human IL-15 (rhIL-15) after lymphodepleting chemotherapy and haploidentical NK cells. Escalating doses of rhIL-15 (0.3-1.0 μg/kg) were given on 12 consecutive days in a phase 1 trial. Of 26 patients, 36% had robust in vivo NK-cell expansion at day 14, and 32% achieved complete remission. Hypothesizing that SC dosing of rhIL-15 would be safer and better tolerated, 16 patients received 10 once per day doses of SC rhIL-15 at 2.0 μg/kg on a phase 2 trial. NK-cell expansion at day 14 was seen in <em>27</em>% of the patients, and 40% achieved remission. rhIL-15 induced better rates of in vivo NK-cell expansion and remission compared with previous trials with IL-2, but it was associated with previously unreported cytokine release syndrome (CRS) after SC but not IV dosing. CRS was observed in 56% of patients given SC rhIL-15 (with concurrent neurologic toxicity in 5 of 9 patients) and was responsive to steroids and tocilizumab. SC administration was associated with slower pharmacokinetic clearance and higher levels of IL-6 than IV dosing. These novel trials testing the use of IL-15 to potentiate cell therapy suggest that dosing schedules based on pharmacokinetics and pharmacodynamics will preserve the therapeutic benefits of IL-15 and minimize CRS. These trials were registered at www.clinicaltrials.gov as #NCT01385423 and #NCT02395822.

The independent effects of exercise and weight loss on markers of inflammation (MOI) in obese individuals have not been clearly characterized. The objectives of this study were to: (i) identify the independent effects of exercise and weight loss on MOI and (ii) determine whether changes in MOI were associated with changes in fat distribution. Subjects were 126 healthy, premenopausal women, BMI <em>27</em>-30 kg/m(2). They were randomized to one of three groups: diet only, diet + aerobic-, or diet + resistance training until a BMI <25 kg/m(2) was achieved. Fat distribution was measured with computed tomography, and body composition with dual-energy X-ray absorptiometry. Serum concentrations of tumor necrosis factor (TNF)-α, soluble TNF receptor 1 (sTNF-R1), soluble TNF receptor 2 (sTNF-R2), C-reactive protein (CRP), and <em>interleukin</em> (IL)-6 were assessed. Results of repeated-measures ANOVA indicated a significant effect of time on MOI, such that MOI decreased with weight loss. Results of mixed-model analysis indicated that adjusting for intra-abdominal adipose tissue (IAAT) and total fat mass explained the decreases in TNF-α and sTNF-R1, whereas only total fat mass explained the decreases in sTNF-R2, IL-6, and CRP. In conclusion, weight loss was associated with decreases in MOI. The effect of weight loss appeared to be mediated by changes in total fat mass or IAAT. Addition of exercise did not alter the response, suggesting that weight loss has a more profound impact for reducing MOI in overweight women than exercise.

Myocarditis, a general inflammatory condition of the heart muscle, can result from a variety of etiologies, the most common being viral. Despite common pathogens, concomitant myocarditis and myositis remains a rare event. Although a common cause of respiratory illness, extrapulmonary infections with influenza are infrequent. We describe the case of a patient who presented to our centre with concomitant "seasonal" H1N1 influenza A myocarditis further complicated by pan-myositis. The patient's condition rapidly declined, eventually requiring biventricular mechanical support, in addition to multilimb fasciotomies. The cardiac support required was progressive, from a percutaneous left ventricular assist device, to extracorporeal membrane oxygenation, to eventual biventricular assist device support for bridge-to-transplantation. This case motivated a detailed review of the literature (a total of 29 cases were identified), in which we found that patients with influenza myocarditis/myositis were predominantly female (63%) and young (mean age 33.2 years) and continue to have a high incidence of morbidity and mortality (<em>27</em>%). As a result of its atypical pattern, the 2009 H1N1 pandemic strain has gained attention. From our review, we found 7 patients with of 2009 H1N1 pandemic influenza myocarditis. Serial serum cytokine analysis did not demonstrate a "cytokine storm," which has been associated with other virulent influenza strains. The PB1-F2 marker in particular has been associated with a vigorous cytokine response. The 2009 H1N1 and "seasonal" influenza strains lack this marker. In those patients with community-acquired influenza, <em>interleukin</em>-6 has been shown to correlate with symptoms. For patients with myocarditis resulting in shock, mechanical circulatory support has gained acceptance as a means to recovery or transplantation.

Dendritic cells (DCs) have been shown to enhance anti-tumor immune responses in several preclinical models. Furthermore, DC-like function can be elicited from peripheral blood monocytes cultured in vitro with <em>interleukin</em>-4 and granulocyte-macrophage colony-stimulating factor. For this reason, a phase 1 study was initiated at the Surgery Branch of the National Cancer Institute to test the toxicity and biological activity of the intravenous administration of peripheral blood monocyte-derived DCs. The DCs were generated by 5- to 7-day incubation in <em>interleukin</em>-4 (1,000 U/mL) and granulocyte-macrophage colony-stimulating factor (1,000 U/mL) of peripheral blood monocytes obtained by leukapheresis. Before administration, the DCs were pulsed separately with the HLA-A*0201-associated melanoma epitopes MART-1(<em>27</em>-35) and gp-100-209-2M. The DCs were administered four times at 3-week intervals. A first cohort of patients (n = 3) was treated with 6 x 10(7) DCs and a second cohort (n = 5) with 2 x 10(8) DCs (in either case, one half of the DCs were pulsed with MART-1(<em>27</em>-35) and the other half was pulsed with gp-100-209-2M). In a final cohort under accrual (n = 2) 2 x 10(8) DCs were administered in combination with <em>interleukin</em>-2 (720,000 IU/kg every 8 hours). The recovery of DCs after in vitro culture ranged from 3% to 35% (mean, 15%) of the original peripheral blood monocytes. Administration of DCs caused no symptoms at any of the doses, and the concomitant administration of <em>interleukin</em>-2 did not cause toxicity other than that expected for <em>interleukin</em>-2 alone. Monitoring of patients' cytotoxic T lymphocyte reactivity before and after treatment revealed enhancement of cytotoxic T lymphocyte reactivity only in one of five patients tested. Of seven patients evaluated for response, one had a transient partial response with regression of pulmonary and cutaneous metastases. A relatively large number of DCs can be safely administered intravenously. The poor clinical outcome of this study perhaps could be explained by the type of protocol used for DC maturation, the route of administration, or both. For this reason, this clinical protocol was interrupted prematurely, whereas other strategies for DC preparation and route of administration are being investigated at the authors' institution.

Background: COVID-19 is an ongoing threat to society. Patients who develop the most severe forms of the disease have high mortality. The interleukin-6 inhibitor tocilizumab has the potential to improve outcomes in these patients by preventing the development of cytokine release storm.

Methods: We conducted a retrospective, case-control, single-center study in patients with severe to critical COVID-19 disease treated with tocilizumab. Disease severity was defined based on the amount of oxygen supplementation required. The primary endpoint was the overall mortality. Secondary endpoints were mortality in non-intubated patients, and mortality in intubated patients.

Results: A total of 193 patients were included in the study. 96 patients received tocilizumab, while 97 served as control group. The mean age was 60 years. Patients over 65 years represented 43% of the population. More patients in the tocilizumab group reported fever, cough, and shortness of breath (83%, 80%, and 96% versus 73%, 69%, and 71%, respectively). There was a non-statistically significant lower mortality in the treatment group (52% versus 62.1% P = 0.09). When excluding intubated patients, there was statistically significant lower mortality in patients treated with tocilizumab (6 vs. 27% P = 0.024). Bacteremia was more common in the control group (24% vs 13% P = 0.43), while fungemia was the similar for both (3% vs 4% P = 0.72).

Conclusion: Our study showed a non-statistically significant lower mortality in patients with severe to critical COVID-19 disease who received tocilizumab. When intubated patients were excluded, the use of tocilizumab was associated with lower mortality.

<em>Interleukin</em>-17 (IL-17) gene models have been found in the sequenced genomes of Strongylocentrotus purpuratus and Caenorhabditis elegans. However, there have been no published reports on the empirical cloning and characterization of any <em>interleukin</em> cDNAs in invertebrates. From a Pacific oyster (Crassostrea gigas) hemocyte cDNA library, two clones were obtained that encoded a protein similar to vertebrate IL-17s. The putative oyster IL-17 homolog (CgIL-17) was <em>27</em>% identical to rainbow trout IL-17D, 21% to human IL-17D and 24% to an IL-17D-like gene model obtained from the annotation of the sea urchin genome. IL-17s from the oyster, sea urchin, trout and human, contained conserved cysteine residues found in all forms of IL-17 in mammals. Injection of bacteria into C. gigas oysters produced a large and rapid elevation in CgIL-17 transcript abundance in hemocytes, suggesting that this is a very early response gene to pathogens that may be responsible for the stimulation of other immune genes in the oyster.

Endometrial cells take part in embryo-maternal communication, as well as supporting the immune system in defending against invading pathogens. The aim of the present study was to examine the mRNA expression of factors that have been suggested to be involved in both events in the bovine endometrial epithelium, namely bovine granulocyte chemotactic protein 2 (CXCL5), <em>interleukin</em>-1 beta (IL1B), IL6, IL8, tumour necrosis factor (TNF), cyclooxygenase 2 (PTGS2) and haptoglobin (HP). Samples were collected in vivo from cows on Days 21-<em>27</em> postpartum by the cytobrush method to evaluate the correlation between inflammatory factors and uterine health (cows with signs of clinical or subclinical endometritis and healthy cows). Bovine uteri were collected at the abattoir to investigate oestrous cycle-dependent mRNA expression patterns. Real-time reverse transcription-polymerase chain reaction revealed that the expression of CXCL5, IL1B, IL8 and TNF mRNA was significantly higher in cows with subclinical or clinical endometritis compared with healthy cows. The expression of CXCL5, IL1B and IL8 mRNA was increased around ovulation compared with the luteal phase. There was no indication of either oestrous cycle-dependent expression or a correlation with uterine health for IL6, PTGS2 and HP transcripts. These results suggest that CXCL5, IL1B, IL8 and TNF may represent potential marker genes for the detection of cows with subclinical endometritis and for monitoring new therapeutic approaches.

We compared gene expression in blood neutrophils (polymorphonuclear leukocytes, or PMNs) collected from healthy subjects with those of cystic fibrosis (CF) patients devoid of bacterial colonization. Macroarray analysis of 1050 genes revealed upregulation of 62 genes and downregulation expression of <em>27</em> genes in CF blood PMNs. Among upregulated genes were those coding for vitronectin, some chemokines (particularly CCL17 and CCL18), some <em>interleukin</em> (IL) receptors (IL-3, IL-8, IL-10, IL-12), all three colony-stimulating factors (G-, M-, GM-CSF), numerous genes coding for molecules involved in signal transduction, and a few genes under the control of gamma-interferon. In contrast, none of the genes coding for adhesion molecules were modulated. The upregulation of six genes in CF PMNs (coding for thrombospondin-1, G-CSF, CXCL10, CCL17, IKKvarepsilon, IL-10Ra) was further confirmed by qPCR. In addition, the increased presence of G-CSF, CCL17, and CXCL10 was confirmed by ELISA in supernatants of neutrophils from CF patients. When comparison was performed between blood and airway PMNs of CF patients, there was a limited difference in terms of gene expression. Only the mRNA expression of amphiregulin and tumor necrosis factor (TNF) receptor p55 were significantly higher in airway PMNs. The presence of amphiregulin was confirmed by ELISA in the sputum of CF patients, suggesting for the first time a role of amphiregulin in cystic fibrosis. Altogether, this study clearly demonstrates that blood PMNs from CF patients display a profound modification of gene expression profile associated with the disease, suggesting a state of activation of these cells.