BACKGROUND

Morbid obesity is associated with insulin resistance (IR), chronic inflammation and premature atherosclerosis. Since vascular inflammation may contribute to the increased risk of cardiovascular morbidity and mortality of these patients, we studied circulating Interleukin-18 (L-18) and monocyte-chemoattractant-protein-1 (MCP-1) levels in 37 patients with morbid obesity before and after significant weight loss induced by bariatric surgery and their preoperative and postoperative associations with C-reactive protein (CRP) and IR-associated factors.

METHODS

High sensitivity assays were used to measure concentrations of fasting CRP, IL-18 and MCP-1. Differences between patients before and after bariatric surgery were analyzed by Student's paired t-test. To investigate the associations of the observed reductions of values, delta of parameters were calculated and preoperative, postoperative and delta data were tested by univariate and multivariate linear regression.

RESULTS

After a mean follow-up period of 26.5 months and a massive weight loss of 35 kg induced by bariatric surgery, circulating IL-18 levels decreased by 37% (P<0.001) and circulating MCP-1 levels by 47% (P<0.001). Multiple linear regression of delta values of IL-18 showed that only 2-hour glucose (P=0.008) remained independently and significantly associated with IL-18, whereas multiple linear regression analysis of delta values of MCP-1 revealed that only delta of HOMA-IR (P<0.001) remained independently and significantly associated with MCP-1, respectively.

CONCLUSIONS

Because both biomarkers have been shown to play an important role in the development and progression of atherosclerosis, the observations presented in this study could be of clinical relevance for morbidly obese patients undergoing bariatric surgery.

The present study examined the skeletal muscle expression of several genes related to the inflammatory process before and after a bout of downhill running. Twenty-nine males between the ages of 18 and <em>35</em> years performed a 45-min downhill (-17.5%) treadmill protocol at 60% of maximal oxygen consumption. Venous bloods samples and muscle biopsy samples from the vastus lateralis were donated prior to and at 3-h and 24-h postexercise, along with ratings of perceived muscle soreness. Serum creatine kinase (CK) was determined, as was skeletal muscle gene expression of <em>interleukin</em> (IL)-6, IL-8, IL-12 (p<em>35</em>), tumor necrosis factor-alpha (TNF-alpha), IL-1beta, cyclooxygenase 2 (COX2), and nuclear factor kappa B (NFkB) (p105/p50). Gene expression was analyzed using RT-PCR and compared with a standard housekeeping gene (beta-actin). Data were analyzed for statistical differences using multivariate analysis of variance with univariate follow-up. In addition, Pearson correlations were conducted to determine if any significant relationship exists between any of these transcripts and both CK and muscle soreness. Significant (p < 0.05) up-regulations in IL-6, IL-8, and COX2 mRNA expression were observed compared with baseline, whereas no significant changes for IL-12, IL-1beta, TNF-alpha, or NFkB were noted. Significant increases in IL-6 mRNA were observed at 3 h (p < 0.001) and 24 h (p = 0.043), whereas significant increases in IL-8 (p = 0.001) and COX2 (p = 0.046) mRNA were observed at 3-h postexercise. In addition, muscle soreness was significantly correlated with IL-8 at 24 h (r = -0.370; p = 0.048), whereas CK was significantly related to NFkB at baseline (r = -0.460; p = 0.012). These data indicate that increases in the mRNA expression of IL-6, IL-8, and COX2 occur in the vastus lateralis as a result of damaging eccentric exercise in young, recreationally trained males. Further, it appears that IL-8 transcription may play some role in inhibiting postexercise muscle soreness, possibly through regulation of angiogenesis.

We have previously shown that tumor necrosis factor (TNF)-alpha, a cytokine involved in asthma, enhances Ca2+ responsiveness to bronchoconstrictor agents in cultured human airway smooth muscle (ASM) cells. In the present study, we investigated the potential mechanism(s) by which TNF-alpha modulates ASM cell responsiveness to such agents. In human ASM cells loaded with fura 2, TNF-alpha and <em>interleukin</em> (IL)-1 beta significantly enhanced thrombin- and bradykinin-evoked elevations of intracellular Ca2+. In TNF-alpha-treated cells. Ca2+ responses to thrombin and bradykinin were <em>35</em>0 +/- 14 and 573 +/- 93 nM vs. 130 +/- 17 and 247 +/- 48 nM in nontreated cells, respectively (P < 0.0001). In IL-1 beta-treated cells, the Ca2+ response to bradykinin was <em>35</em>0 +/- 21 vs. 127 +/- 12 nM in nontreated cells (P < 0.0001). The time course for TNF-alpha potentiation of agonist-induced Ca2+ responses requires a minimum of 6 h and was maximum after 12 h of incubation. In addition, cycloheximide, a protein synthesis inhibitor, completely blocked the potentiating effect of TNF-alpha on Ca2+ signals. We also found that TNF-alpha significantly enhanced increases in phosphoinositide (PI) accumulation induced by bradykinin. The percentage of change in PI accumulation over control was 115 +/- 8 to 210 +/- 15% in control cells vs. 128 +/- 10 to 437 +/- 92% in TNF-alpha-treated cells for 3 x 10(-9) to 3 x 10(-6) M bradykinin. The PI turnover to 10 mM NaF, a direct activator of G proteins, was also found to be enhanced by TNF-alpha. The percentage of change in PI accumulation over control increased from 280 +/- <em>35</em>% in control cells to 437 +/- 92% in TNF-alpha-treated cells. Taken together, these results show that TNF-alpha can potently regulate G protein-mediated signal transduction in ASM cells by activating pathways dependent on protein synthesis. Our study demonstrates one potential mechanism underlying the enhanced Ca2+ response to bronchoconstrictor agents induced by cytokines in human ASM cells.

OBJECTIVE

Administration of female sex steroids in males after trauma-hemorrhage has salutary effects on the depressed immune responses.

METHODS

Randomized laboratory experiment.

METHODS

Male C3H/HeN mice were subjected to midline laparotomy and hemorrhagic shock (<em>35</em>+/-5 mm Hg for 90 minutes, then resuscitation) or sham operation and received subcutaneous 17beta-estradiol (40 microg/kg body weight) or corn oil vehicle at the beginning of resuscitation.

METHODS

At 24 hours after hemorrhage, the animals were killed and plasma 17beta-estradiol and IL-6, splenocyte interleukin (IL) 2, IL-3, and IL-10 production as well as splenic and peritoneal macrophage IL-1beta, IL-10, and IL-6 release were measured.

RESULTS

Splenocyte IL-2 and IL-3 release were significantly depressed after hemorrhage in vehicle-treated mice (P<.05, analysis of variance). Treatment with 17beta-estradiol after hemorrhage led to the restoration of splenocyte IL-2 and IL-3 release. The depressed proinflammatory cytokine (IL-1 and IL-6) release seen in splenic and peritoneal macrophages was restored in the 17beta-estradiol-treated hemorrhage group. In contrast, the sustained release of the anti-inflammatory cytokine IL-10 by splenocytes and splenic and peritoneal macrophages in vehicle-treated mice after hemorrhage was decreased in 17beta-estradiol-treated mice. The increase in circulating IL-6 levels after hemorrhage was significantly attenuated in 17beta-estradiol-treated mice. Although administration of 17beta-estradiol increased plasma 17beta-estradiol levels by approximately 100% in sham as well as hemorrhage groups, improved immune responses were seen only in posthemorrhage 17beta-estradiol-treated mice. There was no adverse effect of 17beta-estradiol treatment in the sham or posthemorrhage groups.

CONCLUSIONS

Since administration of a single dose of 17beta-estradiol in males after trauma-hemorrhage restores the immune functions and decreases circulating levels of IL-6, hormones with estrogenic properties should be considered as safe and novel therapeutic agents for restoring the immune responsiveness in male trauma victims.

Rat hepatic cells respond to <em>interleukin</em> (IL) -1, IL-6, and dexamethasone treatment by increasing the transcription rate of acute-phase plasma protein genes. The same conditions lead to changes in the expression of CAAT-enhancer binding protein (C/EBP) isoforms which are specific to the hepatic cell line. To identify the relationship between C/EBP isoforms and acute-phase protein gene activation, the hormone-specific expression of C/EBP alpha, beta, and delta was determined in H-<em>35</em> and HTC cells and was compared to acute-phase liver. C/EBP beta was found to be the principal isoform in hepatoma cells and to be strongly stimulated by cytokines and dexamethasone in H-<em>35</em> cells. Transactivating functions were observed for all three C/EBP isoforms by cotransfection of CAT gene reporter constructs containing cytokine and glucocorticoid response elements of acute-phase protein genes and expression plasmids for mouse C/EBP alpha, beta, and delta into rat and human hepatoma cells. The degree of C/EBP-mediated transactivation was, however, extremely variable among the different regulatory elements. Transcription run-on reactions with nuclei from transiently transfected H-<em>35</em> cells indicated that cotransfected C/EBP beta increases basal expression of reporter gene constructs as well as the dexamethasone-mediated stimulation of constructs containing the glucocorticoid response elements of the rat alpha 1-acid glycoprotein gene, but did not accelerate or enhance hormone-dependent transcription activation of reporter gene plasmids containing the IL-6 regulatory element of the beta-fibrinogen gene. Activation of the reporter gene constructs appeared to be temporally and quantitatively correlated with the amount of nuclear C/EBP as determined by two-dimensional Western and Southwestern blot analyses.

Eighty-five percent of male STR/ort mice develop osteoarthritic lesions of the knee joint by <em>35</em> weeks of age. We have developed a non-radioactive in-situ hybridization method using digoxigenin-labeled oligonucleotide probes to study the expression of the cytokines <em>interleukin</em> (IL) 1 alpha, Il-1 beta and IL-6 and the growth factors insulin-like growth factor-1 (IGF-1) and transforming growth factor beta (TGF beta 1) during the development of osteoarthritis (OA) in this model. Age- and sex-matched CBA mice, which do not develop OA, showed no detectable expression of any of the cytokines or growth factors studied. In contrast, 20-week-old STR/ort mice with no OA lesions showed positive expression [positive: (+)] for all the cytokines and growth factors studied. At <em>35</em> weeks of age, STR/ort mice with varying grades of OA showed positive (+) or strong (++) signals for both cytokines and growth factors throughout the tibial articular cartilage. The strongest signal was seen in areas where OA lesions were present. In areas of histologically-normal cartilage adjacent to the lesions, the signals were still positive but weaker. Fifty-week-old STR/ort mice with OA lesions showed a similar pattern of expression to <em>35</em>-week-old mice. Thirty-five or 50-week-old STR/ort mice with no OA lesions had much reduced expression compared with those with OA lesions. These mice may be indicative of those STR/ort mice which do not develop OA. The results seen in the STR/ort together with previous biochemical analyses are consistent with an up-regulation of anabolic growth factors and catabolic cytokines in the prelesional stages of OA with anabolic effects predominating. At later stages of OA, the effects of catabolic factors appear to predominate and osteoarthritic lesions become evident.

BACKGROUND

Patients with atopic dermatitis (AD) are prone to have skin infections. We aimed to investigate mRNA expression levels of various antimicrobial peptides and proteins (AMPs) in AD patients, and compare it with psoriasis vulgaris (PV) patients and healthy subjects.

METHODS

Skin biopsies were obtained from healthy subjects and patients with AD and PV. Quantitative real-time RT-PCR was used to determine the mRNA levels of human beta-defensin (hBD)-1, hBD-2, hBD-3, LL-37, psoriasin, RNase 7, interferon-gamma, and interleukin-10 (IL-10).

RESULTS

Except for LL-37, mRNA of hBDs, psoriasin, and RNase 7 was significantly higher expressed in AD (n = 42) and/or PV (n = 35) patients when compared to controls (n = 18). While PV lesions showed significantly higher mRNA hBD-2 levels than lesions of AD, the latter was associated with significantly higher mRNA levels of RNase 7 when compared to PV. A significant positive correlation of hBD expression was observed both in AD patients and PV patients. hBD mRNA levels of AD skin correlated with psoriasin and RNase 7 levels. hBD-1 mRNA expression correlated with AD activity and IL-10 mRNA expression.

CONCLUSIONS

Most AMPs investigated in this study proved to be overexpressed in AD as well as PV when compared to controls. However, a statistically significant difference in AMP mRNA expression between AD and PV was only found for hBD-2 and RNase 7. A moderate-to-strong linear relationship between the mRNA expression of particular AMPs appears to exist in AD, and to a lesser extent in PV as well.

We have established a sensitive ELISPOT assay measuring interferon gamma (IFN gamma) release on a single-cell basis to detect influenza peptide-specific CD8+ T cells in uncultured peripheral blood mononuclear cells (PBMC). Using this method, we studied the T cell response to HLA-A1 and HLA-A2.1 binding peptide epitopes derived from the MAGE-1 and MAGE-3 proteins, from the melanoma-associated antigens tyrosinase, Melan-A/MART-1 and gp100, and from influenza proteins in stage IV melanoma patients and healthy controls. In 18 of 24 HLA-A2-positive donors (75%), but only in 9 of 25 HLA-A2-positive melanoma patients (36%) T cells reactive with the influenza matrix peptide were demonstrated (p = 0.007). T cells responding to one or several of the melanoma-associated peptides were detected in 5 of 25 HLA-A2-positive patients with metastatic melanoma. Four of these 5 patients had been treated with <em>interleukin</em>-2- and IFN alpha-containing therapy. Two of the 24 healthy donors had T cells reactive with the MART-1 27-<em>35</em> peptide. No reactivity with the HLA-A1-binding peptides from MAGE-1 or MAGE-3 was detected in any of the HLA-A1-positive healthy controls or melanoma patients. These results show that the IFN gamma-ELISPOT assay is suitable to determine quantitatively T cells reactive with melanoma-associated and influenza peptide epitopes in uncultured PBMC. The failure to detect T cells responding to influenza in many melanoma patients with progressive disease may indicate an impairment of their T cell function.

The B1 cell surface molecule (CD20) is a <em>35</em>-kDa phosphoprotein expressed by B lineage cells during most stages of differentiation. Some monoclonal antibodies reactive with B1 induce activation while others, anti-B1a, inhibit B lymphocyte function. To further determine the requirement of B1 molecule function in proliferation and differentiation the effects of anti-B1a antibody binding on early cellular activation events were examined. Immunoglobulin secretion of lymphocyte cultures stimulated with pokeweed mitogen was maximally inhibited if the antibody (1-10 micrograms/ml) was added during the first 24 h of culture. However, even high antibody concentrations were unable to inhibit increases in free intracellular Ca2+ concentrations immediately following cross-linkage of cell surface immunoglobulin, or inhibit cell enlargement and the expression of transferrin and <em>interleukin</em> 2 receptors. Antibody binding to B1 inhibited RNA synthesis (37-80%) and progression through cell cycle following activation. In contrast, proliferation induced by phorbol myristate acetate was not inhibited by antibody binding to the B1 molecule. The findings that the earliest activation events and phorbol myristate acetate-induced proliferation were not inhibited by antibody binding to B1 suggest that inhibition is due to the blocking of a step of the activation process required for cell cycle progression and differentiation, rather than blocking initial signal transduction across the membrane or providing a negative or suppressive signal.

Eotaxin is a novel C-C chemokine with selective chemoattractant activity for eosinophils. We determined whether eotaxin could be produced by human airway smooth muscle (HASM) cells in culture and examined its regulation by <em>interleukin</em>-10 (IL-10) and the corticosteroid, dexamethasone. Stimulation of the cells with <em>interleukin</em>-1beta (IL-1beta) or tumour necrosis factor (TNFalpha) each at 10 ng ml(-1) induced the release of eotaxin protein with maximal accumulation by 24 h. Interferon-gamma (IFNgamma) alone at 10 ng ml(-1) had no effect and there was no synergy between these cytokines on the release of eotaxin. Reverse phase high performance liquid chromatographic (HPLC) analysis of supernatents from cells treated with TNFalpha (10 ng ml(-1) for 96 h showed immunoreactivity to eotaxin which eluted with the expected retention time of 34.5-<em>35</em> min. Both IL-1beta and TNFalpha-induced release of eotaxin was not inhibited by dexamethasone (1 microM), however IL-10 (10 ng ml(-1)) had a significant inhibitory effect. Dexamethasone and IL-10 did not inhibit the induction of eotaxin mRNA induced by IL-1beta or TNFalpha. Thus, human airway smooth muscle cells can release eotaxin and could be an important source of chemokine production during airway inflammatory events.

Acute inflammatory illnesses, including the sepsis syndrome, often include a component of coagulation. A human whole blood culture system was developed so that the relationship between coagulation activation and cytokine responses in the presence or absence of lipopolysaccharide (LPS) could be evaluated. In the absence of LPS stimulation, coagulation activation resulted in a novel pattern of cytokine production. During a 4-hour culture of coagulating blood, significant production of <em>interleukin</em>-8 (IL-8; >2,000 pg/mL) was observed, whereas other proinflammatory cytokines including IL-1 beta, IL-6, or tumor necrosis factor a were undetectable or less than <em>35</em> pg/mL. The cytokine profile was distinct from that of fully anticoagulated, LPS-stimulated blood, which showed levels of all the indicated proinflammatory cytokines>> or = 2,000 pg/mL over the same time period. Over 24 to 48 hours, the coagulation-induced cytokine response was characterized by marked and sustained IL-8 production, limited IL-6 generation (with kinetics delayed relative to IL-8), and minimal or undetectable tumor necrosis factor alpha levels. The magnitude of the whole blood IL-8 response correlated with the level of coagulation activation as determined by measurement of thrombin-antithrombin III complex formation. The combined stimuli of coagulation activation and LPS challenge induced a synergistic enhancement of IL-8 production but not of IL-6. Coagulation-induced cytokine production and the synergistic production of IL-8 by coagulation and LPS could be attenuated by hirudin or tissue factor pathway inhibitor (TFPI). Studies to elucidate mechanisms implicated (1) the TFPI third Kunitz and carboxy-terminus as important structural components for TFPI regulation of coagulation activation and (2) thrombin as a candidate mediator of the mononuclear cell cytokine response to coagulation activation. In summary, a unique aspect of the crosstalk between the coagulation and cytokine cascades in whole blood is shown with the identification of IL-8 as a key proinflammatory participant.

Adipocytokines are a subset of cytokines produced by adipose tissue and are associated with risk of type II diabetes and atherosclerosis. Levels of adipocytokines differ between Black and White Americans, even after adjustment for differences in adiposity, diseases associated with adipocytokines including type 2 diabetes and cardiovascular disease, and general socioeconomic status indicators such as income. We used a series of ancestry informative markers to estimate genetic ancestry in a population-based study of older Black Americans, and examined the association between genetic ancestry and adipocytokines and soluble receptors to help determine which of these may be most amenable to admixture mapping. We typed <em>35</em> ancestry informative markers in 1,241 self-reported Black Americans with available DNA from the Health, Aging, and Body Composition (Health ABC) study with available DNA and used a maximum likelihood approach to estimate percent European ancestry. We used linear regression models to determine the association between these adipocytokines and percent ancestry, and staged models to examine whether adiposity or other measures affected the associations of genetic ancestry and adipocytokines. Mean European ancestry was 22.3+/-15.9%. In multivariate adjusted models, the strongest associations observed were between higher European ancestry and <em>interleukin</em>-6 soluble receptor (IL-6 SR), C-reactive protein (CRP), and adiponectin levels, with <em>interleukin</em>-2 soluble receptor (IL-2 SR) and soluble tumor necrosis factor receptor II (TNF-alpha SR II) also showing more modest but significant associations. The association with adiponectin became stronger after adjustment for adiposity. These novel findings suggest that admixture mapping may identify genetic factors influencing the levels of IL-6 SR, CRP, IL-2 SR, and adiponectin.

Adhesive interactions between murine cerebrovascular endothelial cells (EC) which comprise the blood-brain barrier (BBB) and myelin basic protein (MBP)-specific encephalitogenic T lymphocytes were investigated. Adhesion was assessed by measuring the percent attachment of 51Cr-labeled T cells to EC monolayers. The basal level adhesion (20-<em>35</em>%) was significantly up-regulated by treating EC with recombinant murine gamma interferon (IFN-gamma), <em>interleukin</em>-1 alpha (IL-1 alpha) and/or tumor necrosis factor-alpha (TNF alpha). The ability of these cytokines to modulate adhesion was dose- and time-dependent and could be detected as early as 1 h after treatment. The expression of intercellular adhesion molecule-1 (ICAM-1) by EC was examined by immunofluorescence staining and ELISA. Although all unstimulated EC cultures expressed ICAM-1, treatment of EC with the above cytokines dramatically up-regulated the level of ICAM-1 expression in a dose- and time-dependent fashion similar to that observed in the adhesion assays. Treatment of EC with transforming growth factor-beta 1 (TGF beta) down-regulated the level of T cell adhesion on untreated EC in a dose-dependent manner. Pretreatment of EC with TGF beta also partially inhibited the up-regulation of adhesion induced by IFN-gamma, IL-1 alpha and/or TNF alpha. TGF beta had no effect on the up-regulation of ICAM-1 expression induced by IFN-gamma, IL-1 alpha and/or TNF alpha. These results indicate that in addition to ICAM-1, other molecules may be involved in adhesion of encephalitogenic T cells to the EC comprising the cerebral vasculature.(ABSTRACT TRUNCATED AT 250 WORDS)

BACKGROUND

Cachexia is common in chronic obstructive pulmonary disease (COPD) and is thought to be linked to an enhanced systemic inflammatory response.

OBJECTIVE

We investigated differences in the systemic inflammatory profile and polymorphisms in related inflammatory genes in COPD patients.

METHODS

A cross-sectional study was performed in 99 patients with COPD (Global Initiative for Chronic Obstructive Lung Disease stages II-IV), who were stratified by cachexia based on fat-free mass index (FFMI; in kg/m2: <16 for men and <15 for women) and compared with healthy control subjects (HCs). Body composition was determined by bioelectrical impedance analysis. Plasma concentrations and gene polymorphisms of interleukin 1beta (IL-1beta -511), IL-6 (IL-6 -174), and the tumor necrosis factor system (TNF-alpha -308 and lymphotoxin-alpha +252) were determined. Plasma C-reactive protein, leptin, and urinary pseudouridine (as a marker of cellular protein breakdown) were measured.

RESULTS

Fat mass, leptin, and pseudouridine were significantly different (P < 0.001) between noncachectic patients (NCPs) and cachectic patients (CPs: n = 35); the systemic inflammatory cytokine profile was not. NCPs had a body compositional shift toward a lower fat-free mass and a higher fat mass compared with HCs. CPs and NCPs had a greater systemic inflammatory response (P < 0.05) than did HCs, as reflected in C-reactive protein, soluble TNF-R75, and IL-6 concentrations. The overall distribution of the IL-1beta -511 polymorphism was significantly different between the groups (P < 0.05).

CONCLUSIONS

In COPD patients, who are characterized by an elevated systemic inflammatory response, cachexia is not discriminatory for the extent of increase in inflammatory status. This study, however, indicates a potential influence of genetic predisposition on the cachexia process.

Given that combination therapy with statin plus fibrate confers a risk of myopathy, it is worthwhile to determine whether statin or fibrate monotherapy is associated with greater clinical benefit in individuals with combined hyperlipidemia. In this randomized double-blind study, we compared the efficacy of simvastatin and fenofibrate on indexes of endothelial function (flow-mediated dilation (FMD) of the brachial artery) and inflammatory markers (plasma high-sensitivity C-reactive protein (CRP), <em>interleukin</em>-1 beta (IL-1 beta), soluble CD40, and soluble CD40 ligand (sCD40L) levels), as surrogate indicators of future coronary heart disease (CHD), in patients with combined hyperlipidemia. A total of 70 patients with plasma triglyceride levels between 200 and 500 mg/dl and total cholesterol levels of >200 mg/dl were randomly assigned to receive either simvastatin (20 mg/day) (n=<em>35</em>) or micronized fenofibrate (200 mg/day) (n=<em>35</em>) for 8 weeks. Treatment with simvastatin was associated with significantly greater reduction of total cholesterol and low-density lipoprotein cholesterol (LDL-C), while the decrease in triglycerides was significantly greater in patients receiving fenofibrate. Both fenofibrate and simvastatin markedly reduced plasma levels of high-sensitivity CRP, IL-1 beta, and sCD40L, and improved endothelium-dependent FMD without mutual differences. The changes in plasma inflammatory markers did not correlate with baseline clinical characteristics in both groups. However, the improvement in FMD with fenofibrate treatment correlated inversely with baseline high-density lipoprotein cholesterol (HDL-C) levels, whereas the improvement in FMD with simvastatin treatment was positively related to HDL-C levels. Accordingly, in the subgroup with a baseline HDL-C of < or =40 mg/dl, only fenofibrate significantly improved the endothelium-dependent FMD. On the other hand, in the subgroup with HDL-C >40 mg/dl, only treatment with simvastatin achieved significant improvement in FMD. The data here indicate that in patients with combined hyperlipidemia, both fenofibrate and simvastatin have comparative beneficial effects on various inflammatory markers and differential beneficial effects on endothelial function according to baseline HDL-C levels. These findings should be validated by additional prospective studies, in which patients are stratified by baseline HDL-C prior to randomization.

In chronic hepatitis B virus (HBV) infection, immune responses to hepatitis B core antigen (HBcAg) are weak. <em>Interleukin</em> (IL)-10 is a potent immunosuppressive cytokine which we reported recently to be secreted in response to HBcAg by peripheral blood mononuclear cells (PBMCs) from patients with chronic HBV infection or healthy controls. Using an enzyme-linked immunospot assay, we compared the ability of HBcAg to stimulate IL-10 production by PBMC with that of lipopolysaccharide (LPS), phytohaemagglutinin-P and hepatitis C virus-derived antigens in 16 patients with chronic HBV infection and six healthy controls. Frequencies of IL-10 spot-forming cells (SFC) in response to HBcAg were comparable to those obtained with LPS in patients with chronic HBV infection. Frequencies of IL-10 SFC in response to HBcAg or to LPS were significantly higher in patients with chronic HBV infection than in healthy controls. IL-10 SFC in response to HBcAg consisted of 26-<em>35</em>% T cells, 62-70% monocytes and less than 1% B cells in patients with chronic HBV infection. Only monocytes contributed to IL-10 production in controls. Frequencies of HBcAg stimulated IL-10 SFC representing T cells and monocytes were significantly higher in patients with elevated serum alanine aminotransferase (ALT) and detectable HBV DNA than in patients with normal ALT and undetectable HBV DNA. The potent ability of HBcAg to stimulate IL-10 production by PBMC may contribute importantly to immune tolerance toward HBV.

The murine lymphotoxin (LT) gene has been cloned and used to identify cDNA clones in a library prepared from activated murine T cell mRNA. A recombinant murine genomic library was screened with a human lymphotoxin cDNA probe, resulting in the isolation of the entire LT gene. The murine LT gene structure is similar to the human gene, containing three intervening sequences. An activated murine T cell cDNA library was prepared with poly(A)+ RNA isolated 7 hr after concanavalin A stimulation of an L3T4+ <em>interleukin</em> 2-dependent murine T cell clone. Two colonies of the cDNA library that contained inserts that hybridized with the murine LT gene probe were sequenced and were used to construct expression plasmids. The amino acid sequence deduced from the cDNA indicates that murine LT is highly homologous to human LT (74%) and is related to murine tumor necrosis factor (<em>35</em>% homology). The cDNA was transcribed and was translated in vitro, and was expressed in COS-1 cells. This has resulted in the production of LT biological activity.

OBJECTIVE

The peroxisome proliferator-activated receptor gamma (PPARgamma) is a member of the nuclear receptor superfamily of ligand-dependent transcription factors related to retinoid, steroid, and thyroid hormone receptors. The thiazolidinedione rosiglitazone is a PPARgamma ligand that modulates the transcription of target genes. The aim of this study was to investigate the effects of rosiglitazone on the inflammatory response in mice with collagen-induced arthritis (CIA).

METHODS

CIA was induced in DBA/1J mice by an intradermal injection of 100 microl of an emulsion of bovine type II collagen (CII; 100 microg) in Freund's complete adjuvant (CFA) at the base of the tail. On day 21, a second injection of CII in CFA was administered. Rosiglitazone (10 mg/kg/day) or vehicle (10% DMSO) was administered beginning on day 25 (arthritis onset) until day <em>35</em>. Clinical, radiographic, histopathologic, and laboratory assessments were performed.

RESULTS

Mice immunized with CII in CFA developed erosive arthritis of the hind paws. Macroscopic evidence of CIA first appeared as periarticular erythema and edema of the hind paws. The incidence of CIA was 100% by day 27 in the CII-challenged mice, and the severity progressed over the <em>35</em>-day study period. Radiographic evaluation revealed focal resorption of bone. Histopathologic features of CIA included erosion of cartilage at the joint margins. Rosiglitazone treatment ameliorated the clinical signs on days 26-<em>35</em> and improved the histologic findings in the joint and paw. The degree of oxidative and nitrosative damage was significantly reduced in rosiglitazone-treated mice, as indicated by elevation of malondialdehyde levels, formation of nitrotyrosine, and activation of poly(ADP-ribose) polymerase. Plasma levels of the proinflammatory cytokines tumor necrosis factor, interleukin-1beta, and interleukin-6 were also significantly reduced by rosiglitazone treatment.

CONCLUSIONS

These data demonstrate that rosiglitazone exerts an antiinflammatory effect on chronic inflammation and is able to ameliorate the tissue damage associated with CIA.

OBJECTIVE

Physical activity has recently been established as a potential modifier of the inflammatory process, suggesting that it mitigates inflammation and consequently reduces the incidence of several chronic diseases such as cardiovascular events.

METHODS

This study examined the association between different domains ofself-reported physical activity (work, transportation, household, and leisure time) and three inflammatory markers (fibrinogen, C-reactive protein (CRP), and <em>interleukin</em> 6 (IL-6)). Study subjects included 796 men and women aged <em>35</em>-74 yr with complete data for the main study variables who participated in the 1989/1990 MONItoring trends and determinants in CArdiovascular disease (MONICA) Augsburg Survey. Data were collected using the MONICA Optional Study on Physical Activity (MOSPA) questionnaire, and activity levels were classified into low, moderate, and vigorous physical activities.

RESULTS

Fibrinogen showed an inverse relationship with higher levels of work (Ptrend = 0.038), transportation (Ptrend = 0.025), leisure time (Ptrend = 0.013), and summary physical activity (Ptrend< 0.001). This relationship was still observed after adjusting for age and sex and further correction for body mass index, waist-to-hip ratio, smoking status, hypertension, diabetes, total-to-HDL cholesterol ratio, education, and self-reported limited physical activity due to health problems. IL-6 showed significant results for transportation (Ptrend = 0.031), leisure time (Ptrend = 0.016), and summary physical activity (Ptrend < 0.001), whereas CRP was inversely related with the summary activity (Ptrend = 0.003) in the fully adjusted model. No statistically significant inverse association between household physical activity and any of the investigated markers was found. We observed interactions between summary physical activity and smoking (fibrinogen: P = 0.003) as well as ex-smoking (CRP: P < 0.001; IL-6: P = 0.049).

CONCLUSIONS

These data indicate that beyond leisure time, work and transportation physical activity may reduce inflammation.

The production of human eosinophils in vitro from normal bone marrow by using murine eosinophil differentiation factor (mEDF/<em>interleukin</em> 5) is described. Eosinophil production was selective and first detectable after 14 days and reached a peak between 21 and <em>35</em> days when they were the predominant cell type (41% to 89%). Until day 14, all the eosinophils were typical myelocytes, developing thereafter into metamyelocytes and mature cells. All cell types had characteristic light- and electron-microscopic features, apart from the absence of granules with crystalline cores. The eosinophils produced were readily recovered, and both immature myelocytes and mature cells were functionally active in an antibody-dependent, cell-mediated cytotoxicity assay. mEDF added into the assay enhanced the cytotoxicity but to a lower degree than previously reported for peripheral blood eosinophils, which suggests that they may be partially activated. The possibility that eosinophils could be deactivated was tested by removing mEDF from the culture medium. The eosinophils retained viability and functional activity, however, and showed no increased ability to be activated by mEDF for up to six days after removing the mEDF. The liquid culture of human bone marrow was shown to be an alternative assay for eosinophil differentiation factors to colony formation.

Morin is a flavonoid present in fruits and Chinese herbs. Based on in vitro studies, morin has been reported to show various beneficial biological activities. However, there is growing evidence that conjugative metabolism is central to the biological fate of flavonoids. Therefore, the biological effects of morin could be primarily determined by its conjugated metabolites. In this study, the effects of morin and its sulfates/glucuronides on the production of nitric oxide (NO) and cytokines from lipopolysaccharide (LPS)-activated macrophages were individually investigated and compared. The results indicated that the 50% NO production was inhibited from LPS-activated RAW 264.7 cells by 1.25 mM morin and 1.25 microM morin sulfates/glucuronides. Meanwhile, the 50% inhibition concentration (IC50) values of morin and morin sulfates/glucuronides in activated peritoneal macrophages were 1.5 mM morin and 1.5 microM morin sulfates/glucuronides, respectively. In addition, 30% of the tumor necrosis factor-alpha (TNF-alpha) and <em>35</em>% of the <em>interleukin</em> (IL)-12 productions from activated macrophages were inhibited by 2-2.5 mM morin and 2-2.5 microM morin sulfates/glucuronides, respectively. Furthermore, phagocyte activities in the peripheral blood of those for mice dosed with morin for two months were about 65-70% of controls. Lower NO production and reduced macrophage phagocytic activities corresponded to LPS-resistant state. These findings indicated that morin may exhibit anti-inflammatory activity and reduced the incidence of experimental septic shock through decreasing the functions of macrophages and may regulate immune response through modulating the cytokine profiles. Therefore, morin could be a promising therapeutic candidate for inflammatory disease due to the strong activity of its metabolites.

Vascular response is an essential pathological mechanism underlying various inflammatory diseases. This study determines whether IL-<em>35</em>, a novel responsive anti-inflammatory cytokine, inhibits vascular response in acute inflammation. Using a mouse model of LPS-induced acute inflammation and plasma samples from sepsis patients, we found that IL-<em>35</em> was induced in the plasma of mice after LPS injection as well as in the plasma of sepsis patients. In addition, IL-<em>35</em> decreased LPS-induced proinflammatory cytokines and chemokines in the plasma of mice. Furthermore, IL-<em>35</em> inhibited leukocyte adhesion to the endothelium in the vessels of lung and cremaster muscle and decreased the numbers of inflammatory cells in bronchoalveolar lavage fluid. Mechanistically, IL-<em>35</em> inhibited the LPS-induced up-regulation of endothelial cell (EC) adhesion molecule VCAM-1 through IL-<em>35</em> receptors gp130 and IL-12Rβ2 via inhibition of the MAPK-activator protein-1 (AP-1) signaling pathway. We also found that IL-27, which shares the EBI3 subunit with IL-<em>35</em>, promoted LPS-induced VCAM-1 in human aortic ECs and that EBI3-deficient mice had similar vascular response to LPS when compared with that of WT mice. These results demonstrated for the first time that inflammation-induced IL-<em>35</em> inhibits LPS-induced EC activation by suppressing MAPK-AP1-mediated VCAM-1 expression and attenuates LPS-induced secretion of proinflammatory cytokines/chemokines. Our results provide insight into the control of vascular inflammation by IL-<em>35</em> and suggest that IL-<em>35</em> is an attractive novel therapeutic reagent for sepsis and cardiovascular diseases.

OBJECTIVE

Transforming growth factor-beta (TGF-beta) has been shown to antagonise interleukin-1 (IL-1) effects in different systems. Investigations were carried out to study whether TGF-beta 1 modulates IL-1 induced inflammation and IL-1 effects on articular cartilage in the murine knee joint.

METHODS

IL-1, TGF-beta 1 or both factors together were injected into the knee joint. Inflammation was studied in whole knee histological sections. Patellar cartilage proteoglycan synthesis was measured using 35S-sulphate incorporation while patellar cartilage glycosaminoglycan content was determined with automated image analysis on joint sections.

RESULTS

Co-injection of TGF-beta 1 and IL-1 resulted in synergistic attraction of inflammatory cells. In contrast, TGF-beta 1 counteracted IL-1 induced suppression of articular cartilage proteoglycan synthesis. Proteoglycan depletion was similar shortly after the last injection of IL-1 or IL-1/TGF-beta 1, but accelerated recovery was found with the combination at later days. This protective effect of TGF-beta 1 could not be demonstrated in older mice.

CONCLUSIONS

TGF-beta 1 aggravates IL-1 induced knee joint inflammation, but counteracts the deleterious effects of IL-1 on articular cartilage proteoglycan synthesis and content. The data indicate that TGF-beta 1 could play an important part in articular cartilage restoration after IL-1 induced proteoglycan depletion.

OBJECTIVE

We have previously shown that SB265610 (1-(2-bromo-phenyl)-3-(7-cyano-3H-benzotriazol-4-yl)-urea) behaves as an allosteric, inverse agonist at the C-X-C chemokine (CXCR)2 receptor. The aim of this study was to determine whether SB265610, in addition to two other known antagonists, bind to either of the two putative, topographically distinct, allosteric binding sites previously reported in the Literature.

METHODS

Ten single point mutations were introduced into the CXCR2 receptor using site-directed mutagenesis. Three CXCR2 antagonists were investigated, SB265610, Pteridone-1 (2-(2,3 difluoro-benzylsulphanyl)-4-((R)-2-hydroxy-1-methyl-ethylamino)-8H-pteridin-7-one) and Sch527123 (2-hydroxy-N,N-dimethyl-3-{2-[[(R)-1-(5-methyl-furan-2-yl)-propyl]amino]-3,4-dioxo-cyclobut-1enylamino}-benzamide), and the effect of these mutations on their binding affinity and ability to inhibit <em>interleukin</em>-8-stimulated binding of [(<em>35</em>)S]GTPgammaS was examined.

RESULTS

Seven of the nine mutations introduced into the C-terminal domain and intracellular loops of the receptor produced a significant reduction in affinity at least one of the antagonists tested. Of those seven mutations, three produced a significant reduction in the affinity of all three antagonists, namely K320A, Y314A and D84N. In all but one mutation, the changes observed on antagonist affinity were matched with effects on inhibition of <em>interleukin</em>-8-stimulated [(<em>35</em>)S]GTPgammaS binding.

CONCLUSIONS

These antagonists bind to a common intracellular, allosteric, binding site of the CXCR2 receptor, which has been further delineated. As many of these mutations are close to the site of G protein coupling or to a region of the receptor that is responsible for the transduction of the activation signal, our results suggest a molecular mechanism for the inhibition of receptor activation.