Metallothioneins (MTs) are major zinc binding proteins in the CNS that could be involved in the control of zinc metabolism as well as in protection against oxidative stress. Mice lacking MT-I and MT-II (MT-I + II deficient) because of targeted gene inactivation were injected with kainic acid (KA), a potent convulsive agent, to examine the neurobiological importance of these MT isoforms. At <em>35</em> mg/kg KA, MT-I + II deficient male mice showed a higher number of convulsions and a longer convulsion time than control mice. Three days later, KA-injected mice showed gliosis and neuronal injury in the hippocampus. MT-I + II deficiency decreased both astrogliosis and microgliosis and potentiated neuronal injury and apoptosis as shown by terminal deoxynucleotidyl transferase-mediated in situ end labelling (TUNEL), detection of single stranded DNA (ssDNA) and by increased <em>interleukin</em>-1beta-converting enzyme (ICE) and caspase-3 levels. Histochemically reactive zinc in the hippocampus was increased by KA to a greater extent in MT-I + II-deficient compared with control mice. KA-induced seizures also caused increased oxidative stress, as suggested by the malondialdehyde (MDA) and protein tyrosine nitration (NITT) levels and by the expression of MT-I + II, nuclear factor-kappaB (NF-kappaB), and Cu/Zn-superoxide dismutase (Cu/Zn-SOD). MT-I + II deficiency potentiated the oxidative stress caused by KA. Both KA and MT-I + II deficiency significantly affected the expression of MT-III, granulocyte-macrophage colony stimulating factor (GM-CSF) and its receptor (GM-CSFr). The present results indicate MT-I + II as important for neuron survival during KA-induced seizures, and suggest that both impaired zinc regulation and compromised antioxidant activity contribute to the observed neuropathology of the MT-I + II-deficient mice.

OBJECTIVE

Early burn wound excision modulates the hypermetabolic response in severe pediatric burn injuries.

METHODS

Before-after trial.

METHODS

A 30-bed burn referral center in a private, university-affiliated hospital.

METHODS

We studied <em>35</em> severely burned children who were divided into 2 groups. One group (n = 20) was treated with early burn wound excision within 24 hours after the injury. The second group (n = 15) was treated conservatively with silver sulfadiazine in other burn facilities for 5 days, and burn wounds were surgically excised when patients were admitted to our burn center on day 6 after the injury. Data compiled included oxygen consumption and acute-phase protein, <em>interleukin</em> 1beta; <em>interleukin</em> 6, <em>interleukin</em> 10, tumor necrosis factor alpha, and anabolic hormone (growth hormone, insulinlike growth factor type 1) levels preoperatively and 24 hours and 5 days postoperatively.

METHODS

Acute-phase and hypermetabolic responses.

RESULTS

Early burn wound excision abrogated the hypermetabolic response in pediatric burn patients. Patients who underwent conservative treatment had a significantly more severe inflammatory and hypermetabolic response at the same time interval and significantly lower levels of anabolic hormones.

CONCLUSIONS

Early burn wound excision is a safe therapeutic approach that modulates the hypermetabolic response after burn injury. It was superior to the conservative treatment of silver sulfadiazine and delayed excision, and it should be considered when treating all severe full-thickness burns.

The cytokine <em>interleukin</em> 6 (IL6) has several effects on the central nervous system in addition to the well established regulation of the acute phase inflammatory response. Therefore, the distribution of IL6- and IL6 receptor mRNA in the rat brain has been investigated by in situ hybridization using [<em>35S</em>]-labeled oligonucleotides. The messages of both genes were found in the CA1-CA4 regions as well as in the dentate gyrus of the hippocampus, in the habenulae, the dorsomedial and the ventromedial hypothalamus, in the internal capsule, the optic tract and in the piriform cortex. These data indicate both neuronal and glial localization of IL6 and IL6 receptor and their involvement in an autocrine or paracrine action of the cytokine in centrally regulated functions including neuroendocrine control.

OBJECTIVE

Anti-inflammatory therapeutic approaches are considered for the management of type 2 diabetes and for the prevention of its complications. There is limited evidence regarding the effects of prebiotics on inflammation, especially in patients with type 2 diabetes. This trial aims to examine the effects of oligofructose-enriched inulin on glycemic status, inflammation markers, and metabolic endotoxemia in female patients.

METHODS

Over a period of 8 wk, 52 women with body mass indices of >25 kg/m(2) but (<em>35</em> kg/m(2) with type 2 diabetes were randomly assigned to either an intervention group, in which participants were given oligofructose-enriched inulin (n = 27, consuming 10 g/d of oligofructose-enriched inulin), or to a control group, in which participants were given maltodextrin (n = 25, consuming 10 g/d of maltodextrin). Fasting plasma glucose, glycosylated hemoglobin, high-sensitivity C-reactive protein, <em>interleukin</em>-6, tumor necrosis factor-α, interferon-γ, <em>interleukin</em>-10, and plasma lipopolysaccharide were measured before and after the intervention. Data were analyzed with the use of SPSS software version 13. Paired and unpaired Student t tests and analysis of covariance were used to compare quantitative variables.

RESULTS

Oligofructose-enriched inulin caused a significant decrease in the levels of fasting plasma glucose (19.2 mg/dL; 9.50%), glycosylated hemoglobin (1.0%; 8.40%), interleukin-6 (1.3 pg/mL; 8.15%), tumor necrosis factor-α (3.0 pg/mL; 19.80%) and plasma lipopolysaccharide (6.0 EU/mL; 21.95%) as compared with maltodextrin (P < 0.05). Decreases in levels of interferon-γ (0.3 pg/mL; 16.50%) and high-sensitivity C-reactive protein (3.9 ng/mL; 31.70%) and an increase in the level of interleukin-10 (0.4 pg/mL, 11.50%) were not significant in the oligofructose-enriched inulin group as compared with the maltodextrin group.

CONCLUSIONS

In women with type 2 diabetes and suboptimal daily dietary fiber intake, oligofructose-enriched inulin may help to modulate some inflammatory markers.

OBJECTIVE

This study was undertaken to examine whether oligohydramnios in women with preterm premature rupture of membranes is associated with evidence of fetal, amniotic, and maternal inflammatory responses.

METHODS

Amniotic fluid index was measured before the performance of amniocentesis in patients with preterm premature rupture of membranes. Fifty-nine patients who were delivered of preterm neonates (gestational age </=<em>35</em> weeks) within 3 days of amniocentesis were included in this study. Amniotic fluid was cultured for aerobic and anaerobic bacteria and for genital mycoplasmas. The intensity of the inflammatory response was evaluated by the following: presence of clinical and histologic chorioamnionitis; amniotic fluid concentrations of <em>interleukin</em> 6, <em>interleukin</em> 1beta, and tumor necrosis factor alpha; amniotic fluid white blood cell count; and <em>interleukin</em> 6 concentrations in umbilical cord plasma at birth. Proinflammatory cytokines were measured with specific and sensitive immunoassays.

RESULTS

Thirty-two percent (19/59) of patients had an amniotic fluid index </=5 cm. Patients with an amniotic fluid index </=5 cm had significantly higher rates of positive amniotic fluid culture results and clinical and histologic chorioamnionitis; higher median amniotic fluid concentrations of <em>interleukin</em> 6, <em>interleukin</em> 1beta, and tumor necrosis factor alpha; and higher median cord plasma concentrations of <em>interleukin</em> 6 than did those with an amniotic fluid index >5 cm (positive amniotic fluid culture result, 79% [15/19] vs 30% [12/40]; clinical chorioamnionitis, 37% [7/19] vs 5% [2/40]; histologic chorioamnionitis, 100% [17/17] vs 69% [24/<em>35</em>]; median amniotic fluid <em>interleukin</em> 6 concentration, 13.5 ng/mL; range, 0.2-142.2 ng/mL vs 3.0 ng/mL and 0.001-115.2 ng/mL; median amniotic fluid <em>interleukin</em> 1beta concentration, 348.0 pg/mL; range, 0.7->80, 000 pg/mL vs 36.6 pg/mL and 0-2075 pg/mL; median amniotic fluid tumor necrosis factor alpha concentration, 132.0 pg/mL; range, 0-1600 pg/mL vs 11.2 pg/mL and 0-1305 pg/mL; median cord plasma <em>interleukin</em> 6 concentration, 49.7 pg/mL; range, 4.4-7400 pg/mL vs 9. 1 pg/mL and 0-5211 pg/mL; P <.05 for each). There was no significant difference between the 2 groups of patients in the mean umbilical artery pH at birth.

CONCLUSIONS

Oligohydramnios in women with preterm premature rupture of membranes is associated with an inflammatory response in the fetal, amniotic, and maternal compartments.

OBJECTIVE

Traumatic joint injury can initiate early cartilage degeneration in the presence of elevated inflammatory cytokines (e.g., tumor necrosis factor (TNF)-α and interleukin (IL)-6). The positive/negative effects of post-injury dynamic loading on cartilage degradation and repair in vivo are not well-understood. This study examined the effects of dynamic strain on immature bovine cartilage in vitro challenged with TNF-α + IL-6 and its soluble receptor (sIL-6R) with/without initial mechanical injury.

METHODS

Groups of mechanically injured or non-injured explants were cultured in TNF-α + IL-6/sIL-6R for 8 days. Intermittent dynamic compression was applied concurrently at 10%, 20%, or 30% strain amplitude. Outcome measures included sulfated glycosaminoglycan (sGAG) loss (dimethylmethylene blue (DMMB)), aggrecan biosynthesis ((35)S-incorporation), aggrecanase activity (Western blot), chondrocyte viability (fluorescence staining) and apoptosis (nuclear blebbing via light microscopy), and gene expression (qPCR).

RESULTS

In bovine explants, cytokine alone and injury-plus-cytokine treatments markedly increased sGAG loss and aggrecanase activity, and induced chondrocyte apoptosis. These effects were abolished by moderate 10% and 20% strains. However, 30% strain amplitude greatly increased apoptosis and had no inhibitory effect on aggrecanase activity. TNF + IL-6/sIL-6R downregulated matrix gene expression and upregulated expression of inflammatory genes, effects that were rescued by moderate dynamic strains but not by 30% strain.

CONCLUSIONS

Moderate dynamic compression inhibits the pro-catabolic response of cartilage to mechanical injury and cytokine challenge, but there is a threshold strain amplitude above which loading becomes detrimental to cartilage. Our findings support the concept of appropriate loading for post-injury rehabilitation.

<em>Interleukin</em>-12 (IL-12) is a pleiotropic cytokine, formerly termed cytotoxic lymphocyte maturation factor (CLMF) or natural killer cell stimulatory factor (NKSF), which is produced primarily by stimulated macrophages. IL-12 is a disulfide-linked heterodimeric cytokine composed of a <em>35</em>-kDa light chain (p<em>35</em>) and a 40-kDa heavy chain (p40). Unlike most other cytokines, simultaneous transfection of mammalian cells with two different genes is necessary for the production of biologically active IL-12. IL-12 exerts a variety of biological effects on human T and natural killer (NK) cells in vitro, in addition to its ability to promote cytolytic activity, including direct stimulation of the production of IFN-gamma and other cytokines from peripheral blood T and NK cells. The recent finding that IL-12 directs the development of a TH1 type immune response from naive T cells demonstrates the critical role of IL-12 in regulating the immune response. The characteristics of IL-12 function described above strongly suggest its potential usefulness in cancer therapy. Indeed, our studies demonstrate that IL-12 exerts potent antitumor effects following systemic or local administration. We have shown that IL-12 delivered by retroviral vectors allows high-level expression and effective eradication of established tumor in multiple murine tumor models including MCA207 sarcoma. Successful therapy is associated with acquisition of a state of long-term, specific and protective immunity to subsequent challenge with tumor. We have recently received approval from the Recombinant DNA Advisory Committee to proceed with IL-12 gene therapy in humans.

OBJECTIVE

As a proinflammatory cytokine, interleukin-17 (IL-17) contributes to the inflammation of many autoimmune diseases. We examined IL-17 levels in serum and tissues from patients with chronic hepatitis B virus infection (HBV), and especially evaluated the role of IL-17 in the pathogenesis and progression of liver fibrosis.

METHODS

Whole venous blood was obtained from four patient groups: chronic hepatitis B (CHB, n = 47), liver cirrhosis (LC, n = 49), primary hepatocellular carcinoma (PHC, n = 44), chronic liver failure (CLF, n = 33), and a normal control group (n = 20). HBsAg was positive in all patients. Liver biopsy samples were acquired from asymptomatic HBsAg carriers (ASC, n = 35), CHB (n = 57), and LC (n = 31) patients. We performed ELISA to measure IL-17 levels in serum samples, and used reverse RT-PCR to measure IL-17 mRNA levels in peripheral blood mononuclear cells (PBMC). IL-17 protein expression was detected in liver biopsy tissues by immunohistochemistry.

RESULTS

Compared to normal controls, serum IL-17 protein and mRNA levels were significantly higher in the four infection groups. LC patients exhibited the highest serum IL-17 and PBMC mRNA levels. No significant differences were found between the other three groups. High levels of IL-17 were also observed in tissues from CHB and LC patients, compared to ASC. IL-17 expression was mainly located in the portal area and was positively correlated with inflammation grade and fibrosis stage.

CONCLUSIONS

IL-17 expression was found to be increased with increasing degrees of liver fibrosis. This suggests that IL-17 may not only induce the inflammation, but also contribute to disease progression and chronicity.

UNASSIGNED

The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/5306959258322482.

External apical root resorption (EARR) can be an undesirable sequela of orthodontic treatment. Previous studies have suggested that EARR has a substantial genetic component. Linkage and association were examined between polymorphisms of the <em>interleukin</em> IL-1 (IL-1A and IL-1B) genes and EARR in <em>35</em> white American families. Buccal swab cells were collected for DNA isolation and analysis. The EARR in the maxillary central incisors, the mandibular central incisors, and the mesial and distal roots of the mandibular first molar were analyzed separately and together by using both linkage and association methods of analysis. Highly significant (P =.0003) evidence of linkage disequilibrium of IL-1B polymorphism with the clinical manifestation of EARR was obtained. The analysis indicates that the IL-1B polymorphism accounts for 15% of the total variation of maxillary incisor EARR. Persons homozygous for the IL-1B allele 1 have a 5.6 fold (95% CI 1.9-21.2) increased risk of EARR greater than 2 mm as compared with those who are not homozygous for the IL-1 beta allele 1. Data indicate that allele 1 at the IL-1B gene, known to decrease the production of IL-1 cytokine in vivo, significantly increases the risk of EARR. These findings are consistent with an interpretation of EARR as a complex condition influenced by many factors, with the IL-1B gene contributing an important predisposition to this common problem. Defining genetic contributions to EARR is an important factor in understanding the contribution of environmental factors, such as habits and therapeutic biomechanics.

OBJECTIVE

To evaluate ongoing metabolic changes during a 48-h competitive run and a 48-h recovery period, with focus on potential health risks exemplified by heart and skeletal muscle damage biomarkers and oxidative stress-related indices.

METHODS

Blood samples were taken before the race, after 12, 24, and 48 h of running, and after 24 and 48 h of recovery from male amateur runners (N = 7, age <em>35</em>-59 years, VO2max mean ± SD 57.0 ± 4.0 ml kg(-1) min(-1), total distance covered 183-320 km). The samples were analyzed for morphology, acid-base and electrolyte balance, iron status, lipid profile, <em>interleukin</em>-6, high-sensitivity C-reactive protein, N-terminal pro-brain-type natriuretic peptide, high-sensitivity cardiac troponin T, non-enzymatic antioxidants, activities of selected enzymes including antioxidant enzymes, and total antioxidant status.

RESULTS

The sustained ultra-endurance run caused hypocapnic alkalosis with slight hyperkalemia and hypocalcemia, but no hyponatremia. Blood biochemistry showed severe muscle but not liver damage, and an acute inflammatory response. These effects were evidenced by leukocytosis, several fold rises in interleukin-6 and high sensitivity C-reactive protein, extreme elevations in serum levels of muscle enzymes, and marked increases in cardiac biomarker levels. Most of the changes dissolved during the 48 h post-race recovery. Neither the iron pool, nor erythropoiesis, nor pro-oxidant/antioxidant balance were substantially affected.

CONCLUSIONS

The changes consequent on the ultra-endurance run do not pose a serious health risk in men who begin their endeavor with ultra-endurance running in mid-life. There is some circumstantial evidence that hyperventilatory hypocapnia may modulate inflammatory response by stimulating the release of interleukin-6 from working skeletal muscles.

<em>Interleukin</em>-17-expressing CD4+ T helper 17 (Th17) cells are considered as critical regulators of multiple sclerosis disease activity. However, depending on the species and pro-inflammatory milieu, Th17 cells are functionally heterogeneous, consisting of subpopulations that differentially produce <em>interleukin</em>-17, interferon-gamma and granulocyte macrophage colony-stimulating factor. In the current study, we studied distinct effector phenotypes of human Th17 cells and their correlation with disease activity in multiple sclerosis patients. T helper memory populations single- and double-positive for C-C chemokine receptor 6 (CCR6) and CXC chemokine receptor 3 (CXCR3) were functionally assessed in blood and/or cerebrospinal fluid from a total of 59 patients with clinically isolated syndrome, <em>35</em> untreated patients and 24 natalizumab-treated patients with relapsing-remitting multiple sclerosis, and nine patients with end-stage multiple sclerosis. Within the clinically isolated syndrome group, 23 patients had a second attack within 1 year and 26 patients did not experience subsequent attacks during a follow-up of >5 years. Low frequencies of T helper 1 (Th1)-like Th17 (CCR6+CXCR3+), and not Th17 (CCR6+CXCR3-) effector memory populations in blood strongly associated with a rapid diagnosis of clinically definite multiple sclerosis. In cerebrospinal fluid of clinically isolated syndrome and relapsing-remitting multiple sclerosis patients, Th1-like Th17 effector memory cells were abundant and showed increased production of interferon-gamma and granulocyte macrophage colony-stimulating factor compared to paired CCR6+ and CCR6-CD8+ T cell populations and their blood equivalents after short-term culturing. Their local enrichment was confirmed ex vivo using cerebrospinal fluid and brain single-cell suspensions. Across all pro-inflammatory T helper cells analysed in relapsing-remitting multiple sclerosis blood, Th1-like Th17 subpopulation T helper 17.1 (Th17.1; CCR6+CXCR3+CCR4-) expressed the highest very late antigen-4 levels and selectively accumulated in natalizumab-treated patients who remained free of clinical relapses. This was not found in patients who experienced relapses during natalizumab treatment. The enhanced potential of Th17.1 cells to infiltrate the central nervous system was supported by their predominance in cerebrospinal fluid of early multiple sclerosis patients and their preferential transmigration across human brain endothelial layers. These findings reveal a dominant contribution of Th1-like Th17 subpopulations, in particular Th17.1 cells, to clinical disease activity and provide a strong rationale for more specific and earlier use of T cell-targeted therapy in multiple sclerosis.

We describe here the derivation of a rat monoclonal antibody (mAb) against mouse CD40 (designated 3/23), which stains 45-50% of spleen cells of adult mice, approximately 90% of which are B cells. Interestingly, some 5-10% of both CD4+ and CD8+ T cells in the spleens of (some, but not all) adult, unimmunized mice are also CD40+, whereas CD40+ cells were not detectable in the thymus, even following collagenase digestion. Some <em>35</em>-40% of lymphoid cells in the bone marrow of adult mice are CD40+ and virtually all of these are B220+, and hence of the B cell lineage: triple-color flow cytometry showed that CD40 is expressed at low levels on some 30% of pre-B cells, at intermediate levels on 80% of immature B cells and on essentially all mature B cells in the bone marrow. These results, therefore, suggest that in the mouse CD40 is expressed relatively late during the process of B cell differentiation. The mAb induced marked up-regulation of major histocompatibility complex class II molecules, CD23 and B7.2 antigens on mature B cells. It also stimulated modest levels of DNA synthesis in mature B cells by itself: this was markedly enhanced by suboptimal concentrations of mitogenic (but not non-mitogenic) anti-mu and anti-delta mAb, and moderately enhanced by co-stimulation with <em>interleukin</em>-4. Hypercross-linking of CD40 (using biotinylated mAb and avidin) also enhanced the proliferative response to anti-CD40.

OBJECTIVE

The objective of this study was to determine the distribution of two biovars of Ureaplasma urealyticum (parvo and T960) in human amniotic fluid and to examine whether the magnitude of the intrauterine inflammatory response and pregnancy outcomes are different between patients with microbial invasion of the amniotic cavity with "parvo biovar" and those with "T960 biovar".

METHODS

This cohort included 77 preterm singleton pregnancies (gestational age < 37 weeks) in whom U. urealyticum was detected from amniotic fluid using the polymerase chain reaction (PCR). Amniotic fluid was obtained by transabdominal amniocentesis. Amniotic fluid was cultured for aerobic and anaerobic bacteria as well as mycoplasmas. U. urealyticum was biotyped by PCR methods. Amniotic fluid inflammatory response was determined by amniotic fluid white blood cell count and interleukin-6 concentration.

RESULTS

1) The "parvo biovar" was detected in 82% (63/77) and "T960 biovar" was in 18% (14/77) of cases; 2) U. urealyticum was isolated by conventional culture method from amniotic fluid in 56% (35/63) of cases with positive for "parvo biovar" and in 50% (7/14) of cases with positive for "T960 biovar"; 3) There were no significant differences in the median gestational age at amniocentesis, gestational age at delivery, birth weight, amniotic fluid white blood cell count, amniotic fluid interleukin-6 concentration and the rates of clinical chorioamnionitis, histologic chorioamnionitis, funisitis and neonatal morbidity between patients in the two biovar groups.

CONCLUSIONS

1) The "parvo biovar" is more frequently isolated from amniotic fluid of preterm gestations than the "T960 biovar"; 2) Biovar diversity of U. urealyticum in amniotic fluid was not associated with different pregnancy outcome and magnitude of the intraamniotic inflammatory response.

Between 20% and <em>35</em>% of subjects with asthma experience asthma exacerbations during periods of stress. The biological mechanisms underlying these exacerbations are not clearly understood, and the role of psychologic factors in the pathophysiology of asthma remains controversial. We investigated the ability of psychologic stress to modulate airway inflammation and airway hyperresponsiveness (AHR) to methacholine in a murine model of asthma. Animals were exposed to a stressor daily for 3 (short-term stress) or 7 (long-term stress) days. After allergen challenge, AHR was assessed through plethysmography, and bronchoalveolar lavage cells were counted as a measure of inflammation. After short-term stress, inflammatory cell number was decreased compared with unstressed animals, whereas levels of <em>interleukin</em> (IL)-6, IL-9, and IL-13 were increased. Administration of a corticosteroid receptor antagonist, before stress, prevented the decrease in inflammatory cell numbers. In contrast, animals stressed for 7 consecutive days showed a significant increase in inflammatory cell numbers, which was independent of the glucocorticoid response, but no change in cytokine levels. AHR was not altered in stressed animals. Our results indicate that repeated exposure to stress over the long term engages different mechanisms than short-term stress and can exacerbate the chronic inflammatory responses of the airway.

OBJECTIVE

Adequate restoration of intravascular volume remains an important maneuver in the management of the surgical patient. Influence of different volume replacement regimens on inflammation/endothelial activation in elderly surgical patients was assessed.

METHODS

Prospective, randomized study.

METHODS

Surgical intensive care unit of a university-affiliated hospital.

METHODS

Sixty-six patients >65 years undergoing major abdominal surgery.

METHODS

Ringer's lactate (RL; n=22), normal saline solution (NS; n=22) or a low-molecular HES (mean molecular weight 130 kD) with a low degree of substitution (0.4; HES 130/0.4; n=22) were administered after induction of anesthesia until the 1st postoperative day (POD) to keep central venous pressure between 8-12 mmHg.

RESULTS

C-reactive protein, <em>interleukins</em> (IL-6, IL-8), adhesion molecules [endothelial leukocyte adhesion molecule-1 (ELAM-1) and intercellular adhesion molecule-1 (ICAM-1)] were measured prior to volume therapy at the end of surgery, 5 h after surgery and at the morning of the 1st POD. RL patients received 10,150+/-1,660 ml of RL, NS patients 10,220+/-1,770 ml of NS and the HES-treated group 2,850+/-300 ml of HES 130/0.4 and 2,810+/-<em>35</em>0 ml of RL. Hemodynamics were similar in all groups. CRP, IL-6 and IL-8 plasma levels increased significantly higher in both crystalloid groups (IL-6 in the NS group: increase to 407+/-33 pg/ml; RL: increase to 377+/-<em>35</em> pg/dl) than in the HES-130 treated group (IL-6: increase to 197+/-20 pg/dl). Plasma levels of ELAM-1 and ICAM remained almost unchanged in the HES 130-, but significantly increased in the RL- and NS-treated patients.

CONCLUSIONS

In elderly patients, markers of inflammation and endothelial injury and activation were significantly higher after crystalloid- than after HES 130/0.4-based volume replacement regimens.

OBJECTIVE

Despite the substantial clinical efficacy of tumor necrosis factor alpha (TNFalpha) antagonist therapy in patients with rheumatoid arthritis (RA), some patients respond poorly to such agents. Since an interferon (IFN) signature is variably expressed among RA patients, we investigated whether plasma type I IFN activity might predict the response to TNF antagonist therapy.

METHODS

RA patients (n = <em>35</em>), the majority of whom were Hispanic, from a single center were evaluated before and after initiation of TNF antagonist therapy. As controls, 12 RA patients from the same center who were not treated with a TNF antagonist were studied. Plasma type I IFN activity was measured using a reporter cell assay, and disease status was assessed using the Disease Activity Score in 28 joints (DAS28). Levels of <em>interleukin</em>-1 receptor antagonist (IL-1Ra) were determined in baseline plasma samples using a commercial enzyme-linked immunosorbent assay. The clinical response was classified according to the European League Against Rheumatism criteria for improvement in RA.

RESULTS

Plasma type I IFN activity at baseline was significantly associated with clinical response (odds ratio 1.36 [95% confidence interval 1.05-1.76], P = 0.020), with high baseline IFN activity associated with a good response. Changes in DAS28 scores were greater among patients with a baseline plasma IFNbeta/alpha ratio >0.8 (indicating elevated plasma IFNbeta levels). Consistent with the capacity of IFNbeta to induce IL-1Ra, elevated baseline IL-1Ra levels were associated with better therapeutic outcomes (odds ratio 1.82 [95% confidence interval 1.1-3.29], P = 0.027).

CONCLUSIONS

The plasma type I IFN activity, the IFNbeta/alpha ratio, and the IL-1Ra level were predictive of the therapeutic response in TNF antagonist-treated RA patients, indicating that these parameters might define clinically meaningful subgroups of RA patients with distinct responses to therapeutic agents.

OBJECTIVE

The serine protease urokinase-type plasminogen (uPA) and its receptor (uPAR) appear to have a major function in tumor invasion and metastasis. Interleukin-8 (IL-8) acts as an angiogenic factor in solid cancer. The purpose of this study was to determine whether the expression of IL-8 and the uPAR gene correlates with clinicopathological parameters in human gastric carcinoma.

METHODS

We examined the expression of uPAR mRNA and IL-8 mRNA using Northern blot analysis and RT-PCR in 35 gastric carcinomas and the surrounding normal mucosa. Macroscopic and histopathological tumor findings and survival rates were obtained from the patient records.

RESULTS

uPAR and IL-8 mRNA expression levels were higher in most neoplasms compared to the corresponding normal mucosal tissue. A correlation between uPAR and IL-8 expression in tumors was observed (r = 0.447, p < 0.01). uPAR mRNA expression in the tumors correlated well with lymph node metastasis (p < 0.02), and its stage (p < 0.01). The IL-8 MRNA expression in the tumors correlated with diffuse-type gastric cancer (p < 0.05). The survival rates of patients with tumors displaying high uPAR expression levels were significantly lower (p < 0.04) than those of patients without uPAR expression, but patients with IL-8 expression only showed a tendency to a lower survival rate.

CONCLUSIONS

These results suggest that uPAR and IL-8 may be important prognostic factors in human gastric carcinomas.

In <em>35</em> patients fulfilling the criteria of systemic inflammatory response syndrome (SIRS) of infectious origin, tumor necrosis factor-alpha (TNF-alpha), <em>interleukin</em>-6 (IL-6), tumor necrosis factor-soluble receptor (TNF-sR), and <em>interleukin</em>-12 (IL-12), C-reactive protein (CRP) levels and the Acute Physiology, and Chronic Health Evaluation III score (APACHE III) were determined on days 1 to 7, 14, 21, and 28. The Mortality Probability Models (MPM) II sepsis score was assessed at the time of admission into the study. The MPM II sepsis score correlated with IL-6 plasma levels on day 1. The APACHE III scores correlated with plasma levels of TNF-sR on days 2-7, with IL-6 levels on days 3-7, and with CRP levels on days 4-7 (p < 0.01). The MPM II sepsis score, the APACHE III score, and the IL-6, TNF-sR, and CRP levels were significantly different between survivors and nonsurvivors and between patients with and without shock (p < 0.05). A significant decrease of the APACHE III scores, IL-6, and CRP levels was observed over the study period in the survivor group only (p < 0.05), while neither the dynamics of TNF-alpha nor IL-12 plasma levels contributed to the risk estimation of mortality.

BACKGROUND

Alteration in intestinal permeability may be an important factor in the pathogenesis of both the progression of some chronic liver diseases and the onset of some complications in patients with liver cirrhosis.

OBJECTIVE

To investigate the relationships between intestinal permeability, portal hypertension, alcohol use, plasma levels of pro-inflammatory cytokines, and nitric oxide, expressed as s-nitrosothiols, and nitrite levels in patients with various types and degrees of chronic liver diseases.

METHODS

134 healthy volunteers and 83 patients with chronic liver damage entered the study. Intestinal permeability was assessed with the lactulose/mannitol test. Plasma levels of tumour necrosis factor-alpha, interleukin-6, and nitrite and total s-nitrosothiols were determined.

RESULTS

Intestinal permeability was altered in patients with advanced liver disease and impaired in 15-35% of patients without cirrhosis. Independent factors for intestinal permeability alteration were age, portal hypertension, alcohol use, and diabetes. Plasma levels of inflammatory cytokines and nitrosothiols were significantly higher in patients with altered intestinal permeability.

CONCLUSIONS

An intestinal permeability evaluation in patients with chronic liver diseases might clarify the significance of intestinal permeability in the pathophysiology of both the progression of liver damage, and the occurrence of complications that accompany liver cirrhosis.

Several cytokines including gamma-interferon, tumor necrosis factor alpha, <em>interleukin</em> 1 beta (IL-1 beta), and <em>interleukin</em> 6 (IL-6) are pyrogenic and can inhibit lipogenic processes. Because patients with lymphoma often suffer from fever, weight loss, and night sweats (B symptoms), the etiology of which is unknown, the authors investigated serum levels of these cytokines in normal volunteers and in patients with Hodgkin's and non-Hodgkin's lymphoma. Sixty serum samples from patients with Hodgkin's disease (28 patients) or non-Hodgkin's lymphoma (32 patients), as well as 20 samples from normal volunteers, were collected. The majority of patients had advanced (Stage III or IV) or relapsed disease. The assay for gamma-interferon was a specific and sensitive radioimmunoassay (lower limit of detection = 0.1 unit/ml); the assays for tumor necrosis factor alpha, IL-1 beta, and IL-6 were enzyme-linked immunoassays with lower limits of sensitivity of 10 pg/ml, 20 pg/ml, and 22 pg/ml, respectively. There were no statistically significant differences in gamma-interferon, tumor necrosis factor alpha, or IL-1 beta levels between lymphoma patients and normal subjects. In contrast, 20 of 57 patients (<em>35</em>%) with lymphoma as compared with 0 of 19 normal volunteers (0%) had detectable serum IL-6 levels (P < 0.005, chi 2 test). Interestingly, 17 of 29 lymphoma patients with B symptoms (59%) as opposed to 3 of 28 lymphoma patients without B symptoms (11%) had detectable serum IL-6 levels (P < 0.001, chi 2 test); the median IL-6 level was 28.9 pg/ml (B symptoms present) versus undetectable (no B symptoms) (P < 0.005, Mann-Whitney U test). Analyzing Hodgkin's and non-Hodgkin's lymphoma groups separately revealed similar results. IL-6 levels showed no significant correlation with time from diagnosis, beta 2-microglobulin, or lactate dehydrogenase levels. However, analysis by the method of Kaplan and Meir demonstrated that the median survival of Hodgkin's disease patients with detectable IL-6 levels >> or = 22 pg/ml) was 10 mo, whereas the median survival has not been reached at a median follow-up time of 37.5 mo in those patients with lower values (Wilcoxon P value = 0.0012). There were too few patients in each subset of non-Hodgkin's lymphoma to determine the correlation between IL-6 and survival but, considered as a single group, a statistically significant correlation was not found.(ABSTRACT TRUNCATED AT 400 WORDS)

OBJECTIVE

To evaluate the effectiveness of curative radiotherapy (RT) plus recombinant interleukin-2 (rIL-2) immunotherapy regarding the treatment results for angiosarcoma of the scalp. Curative resection of angiosarcoma of the scalp is usually difficult because of the diffuse, clinically undetectable local spread. RT is a rational therapeutic approach, because a wide region of the dermis can be treated, while sparing the underlying normal tissues. Recently, the effectiveness of immunotherapy with rIL-2 has also been reported in the treatment of angiosarcoma of the scalp.

METHODS

The data of 20 patients with angiosarcoma of the scalp treated with curative RT plus rIL-2 immunotherapy between January 1988 and June 2002 were retrospectively analyzed. The total radiation dose was 70.3 +/- 6.9 Gy. The fractions were 2-3 Gy daily, given 5 d/wk. rIL-2 immunotherapy was performed by transcatheter arterial administration in 10 patients, systemic administration in 11 during the course of RT, and intratumoral injection in 10 during and/or after RT; 12 patients received a combination of two. Five patients underwent limited surgery, and concomitant pacilitaxel chemotherapy was also used in 2 patients.

RESULTS

The median survival time for overall, local recurrence-free, and distant metastasis-free survival was 36.2, 11.1, and 17.8 months, respectively. Local recurrence developed in 7 patients (35%), 4 of whom also had evidence of distant metastases. An additional 7 patients (35%) developed distant metastases alone. Recurrence within the radiation field was recognized in 2 patients with systemic rIL-2 administration alone (p < 0.05). Arterial or intratumoral administration combined with systemic administration of rIL-2 resulted in better distant metaststasis-free survival rates (p < 0.05).

CONCLUSIONS

Curative RT plus rIL-2 immunotherapy provided an efficient, effective means of treating angiosarcoma of the scalp. Arterial or intratumoral administration combined with systemic administration of rIL-2 may prolong survival. Additional studies with detailed treatment protocols are recommended.

BACKGROUND

Dendritic cells (DC) play critical roles in the initiation and modulation of immune responses and may determine the balance between tolerance and immunity. Viral interleukin-10 (vIL-10), encoded by the Epstein-Barr virus, is highly homologous to the "immunosuppressive" cytokine, mammalian IL-10. It impairs antigen-presenting cell function but lacks certain immunostimulatory properties of mammalian IL-10. We accomplished the following: (1) evaluated the effects of vIL-10 protein on DC phenotype and function, (2) transduced mouse bone marrow-derived DC to express vIL-10, and (3) assessed the impact of transgene expression on DC allostimulatory activity.

METHODS

DC progenitors propagated from bone marrow of B10 (H2b) mice in granulocyte-macrophage colony-stimulating factor plus IL-4 were repeatedly transduced by centrifugation, using retroviral supernatant obtained from the BOSC 23 ecotropic packaging cell line. To evaluate transduction efficiency, DC were transduced with the retroviral vector MFG-enhanced green fluorescence protein as a marker gene. Transgene and key cell surface molecule expression were examined by flow cytometry. The level of vIL-10 gene product in the culture supernatant was quantitated by ELISA. DC function was assessed by evaluation of the ability of DC to induce allogeneic (C3H;H2k) T-cell proliferation and cytotoxic T lymphocytes in primary mixed leukocyte reactions. Secondary mixed leukocyte reactions were used to test for T-cell hyporesponsiveness.

RESULTS

The early addition of vIL-10 protein to cultures inhibited DC maturation and function. vIL-10 gene transfer was achieved with an approximate transduction efficiency of 35 to 40%. Transduced DC expressed vIL-10 at a level of 40 ng/10(6) cells/48 hr. In comparison with controls, vIL-10-transduced cells showed decreased surface expression of major histocompatibility complex class II and costimulatory molecules, reduced ability to stimulate T-cell proliferation and cytotoxic T lymphocyte generation, and potential to induce alloantigen-specific hyporesponsiveness.

CONCLUSIONS

DC can be effectively transduced to express vIL-10 and limit their ability to stimulate in vitro. These genetically engineered antigen-presenting cells may have therapeutic potential to inhibit undesired immune responses to allo- or autoantigens.

From the finding of micro-organisms or inflammatory mediators, or both, in amniotic fluid (AF), it has been proposed that intrauterine infection is one cause of preterm labour (PTL, intact fetal membranes). This theory, however, remains unproved, i.e. the accumulation of micro-organisms and inflammatory mediators in AF after labour is in progress may be the consequence, not the cause, of labour both at term and preterm. This study was conducted to evaluate this possibility by a comparison of the concentrations of <em>interleukin</em> (IL)-1beta and IL-6 in AFs collected before and during PTL (<34 weeks gestation) with those in AFs collected at term (before labour and from the forebag and upper compartments of the amniotic sac during labour). The concentrations of IL-1beta and IL-6 in AF were also analysed as a function of the duration of labour (term or preterm) before fluid collection. In addition, studies were conducted to define the source of IL-1beta in AF. A total of 666 AFs were evaluated. IL-1beta was not detected (<50 pg/ml) in AFs collected before the onset of labour at any stage of gestation (n = 320), including 170 fluids obtained at term. During labour, IL-1beta was detected (>50 pg/ml) in 58 out of 106 (54.7%), 17 out of 64 (26.6%) and 60 out of 176 (34%) of AF samples obtained during PTL, term labour (upper compartment) and term labour (forebag) respectively. AF sampling, as well as labour and delivery, were completed in <18 h in all term pregnancies. However, labour (with cervical dilation) was in progress for >18 h before AF was collected in 39 out of 106 (37%) PTL pregnancies. The incidence of IL-1beta-positive samples among AFs collected before 18 h of PTL (23 out of 67; 34%) was indistinguishable from that in AFs collected during labour at term. However, in AFs collected after >18 h PTL, the incidence of IL-1beta-positive samples was <em>35</em> out of 39 (89.7%) The concentrations of IL-1beta (pg/ml; mean +/- SEM) in AFs collected during PTL (2680 +/- 730; n = 106) were greater than those in AFs collected from the upper compartment and forebag during term labour (436 +/- 244, n = 64; and 468 +/- 119, n = 176) respectively; this difference, however, was attributable to very high concentrations of IL-1beta in AFs in which PTL was in progress for >18 h before AF collection (6021 +/- 1832; n = 39). The concentrations of IL-6 in AF were correlated with those of IL-1beta (P < 0.001). We conclude that IL-1beta and IL-6 accumulate in AF in a similar proportion of pregnancies during the first 18 h of term and preterm labour. Therefore, the accumulation of these cytokines in AF cannot be taken as evidence for a role for infection in the pathogenesis of PTL.

BACKGROUND

Standard cytokine detection methods are unable to determine which cells are the producing cells. We report on the extent and under which conditions the multilabeling capability of flow cytometry (FCM) can bring new advances into the field.

METHODS

Five different cytokines, <em>interleukin</em>-2 (IL-2), -4, -5, -10 and interferon-gamma (IFN-gamma), were assessed simultaneously under five ex vivo stimulation conditions in peripheral blood mononuclear cells from five healthy volunteers in a 5-day kinetic study. A second group of <em>35</em> volunteers was assessed for IFN-gamma and IL-2 production.

RESULTS

This study showed that (a) intracytoplasmic cytokines were almost undetectable within unstimulated cells, (b) intracytoplasmic cytokines were detected only in CD69(+) T lymphocytes, and (c) intracytoplasmic IL-2 and IFN-gamma were dramatically upregulated after stimulation with phorbol myristate acetate (PMA)-ionomycin in a biphasic response or with PMA-phytohemagglutinin (one major peak only at 18 h) but to a lesser extent with other stimuli such as monoclonal antibodies. Th2 cytokines were detected at a later time point and at lower levels. PMA/ionomycin stimulation after 4 h and 18 h of culture in <em>35</em> other volunteers individualized several subgroups according to the frequency of IFN-gamma- or IL-2-producing cells--IFN-gamma delayed producers (n = 10/<em>35</em>), IFN-gamma low producers (n = 8/<em>35</em>), and IL-2 delayed producers (n = 16/<em>35</em>)--as opposed to IFN-gamma or IL-2 normal producers.

CONCLUSIONS

FCM appears to be a good tool to examine cell cytokine status in pathology (allergy, autoimmune disease, etc.) provided that optimal stimulation conditions and multiple time-point cultures are used. It also seems to be a relevant method to define new Th subsets further.