Chronic inflammation is a characteristic feature of aging, and the relationship between cellular senescence and inflammation, although extensively studied, is not well understood. An overlapping pathway screen identified human polynucleotide phosphorylase (hPNPase(old-<em>35</em>)), an evolutionary conserved 3',5'-exoribonuclease, as a gene up-regulated during both terminal differentiation and cellular senescence. Enhanced expression of hPNPase(old-<em>35</em>) via a replication-incompetent adenovirus (Ad.hPNPase(old-<em>35</em>)) in human melanoma cells and normal human melanocytes results in a characteristic senescence-like phenotype. Reactive oxygen species (ROS) play a key role in the induction of both in vitro and in vivo senescence. We now document that overexpression of hPNPase(old-<em>35</em>) results in increased production of ROS, leading to activation of the nuclear factor (NF)-kappaB pathway. Ad.hPNPase(old-<em>35</em>) infection promotes degradation of IkappaBalpha and nuclear translocation of NF-kappaB and markedly increases binding of the transcriptional activator p50/p65. The generation of ROS and activation of NF-kappaB by hPNPase(old-<em>35</em>) are prevented by treatment with a cell-permeable antioxidant, N-acetyl-l-cysteine. Infection with Ad.hPNPase(old-<em>35</em>) enhances the production of <em>interleukin</em> (IL)-6 and IL-8, two classical NF-kappaB-responsive cytokines, and this induction is inhibited by N-acetyl-l-cysteine. A cytokine array reveals that Ad.hPNPase(old-<em>35</em>) infection specifically induces the expression of proinflammatory cytokines, such as IL-6, IL-8, RANTES, and matrix metalloproteinase (MMP)-3. We hypothesize that hPNPase(old-<em>35</em>) might play a significant role in producing pathological changes associated with aging by generating proinflammatory cytokines via ROS and NF-kappaB. Understanding the relationship between hPNPase(old-<em>35</em>) and inflammation and aging provides a unique opportunity to mechanistically comprehend and potentially intervene in these physiologically important processes.

BACKGROUND

There is contradictory information regarding the prognostic importance of adipocytokines, hepatic and inflammatory biomarkers on the incidence of type 2 diabetes. The objective was to assess the prognostic relevance of adipocytokine and inflammatory markers (C-reactive protein - CRP; interleukin-1beta - IL-1β; interleukin-6- IL-6; tumour necrosis factor-α - TNF-α; leptin and adiponectin) and gamma-glutamyl transpeptidase (γGT) on the incidence of type 2 diabetes.

METHODS

Prospective, population-based study including 3,842 non-diabetic participants (43.3% men, age range 35 to 75 years), followed for an average of 5.5 years (2003-2008). The endpoint was the occurrence of type 2 diabetes.

RESULTS

208 participants (5.4%, 66 women) developed type 2 diabetes during follow-up. On univariate analysis, participants who developed type 2 diabetes had significantly higher baseline levels of IL-6, CRP, leptin and γGT, and lower levels of adiponectin than participants who remained free of type 2 diabetes. After adjusting for a validated type 2 diabetes risk score, only the associations with adiponectin: Odds Ratio and (95% confidence interval): 0.97 (0.64-1.47), 0.84 (0.55-1.30) and 0.64 (0.40-1.03) for the second, third and forth gender-specific quartiles respectively, remained significant (P-value for trend = 0.05). Adding each marker to a validated type 2 diabetes risk score (including age, family history of type 2 diabetes, height, waist circumference, resting heart rate, presence of hypertension, HDL cholesterol, triglycerides, fasting glucose and serum uric acid) did not improve the area under the ROC or the net reclassification index; similar findings were obtained when the markers were combined, when the markers were used as continuous (log-transformed) variables or when gender-specific quartiles were used.

CONCLUSIONS

Decreased adiponectin levels are associated with an increased risk for incident type 2 diabetes, but they seem to add little information regarding the risk of developing type 2 diabetes to a validated risk score.

The objective of this study was to evaluate the effects of adding essential oils to the diet of weaned pigs on performance, nutrient utilization, immune response and intestinal health. A total of 96 weaning pigs (8.37±1.58 kg) were allotted to one of three dietary treatments. The treatments consisted of an unsupplemented basal diet (negative control, NC) or similar diets supplemented with 0.01% of an essential oil product which contained 18% thymol and cinnamaldehyde (EOD) as well as a diet supplemented with 0.19% of an antibiotic mixture which provided 150 ppm chlortetracycline, 80 ppm colistin sulfate and 50 ppm kitasamycin (positive control, PC). Each treatment was provided to eight pens of pigs with four pigs per pen. Over the entire <em>35</em> d experiment, ADG and fecal score were improved (p<0.05) for pigs fed the PC and EOD compared with the NC. Dry matter and crude protein digestibility as well as lymphocyte proliferation for pigs fed the PC and EOD diets were increased significantly compared with NC (p<0.05). IGF-I levels in plasma were significantly increased (p<0.05) in pigs fed the PC diet compared with pigs fed the NC diet. <em>Interleukin</em>-6 concentration was lower (p<0.05) and the tumor necrosis factor-α level was higher (p<0.05) in the plasma of pigs fed the EOD diet than the NC diet. Plasma total antioxidant capacity level increased (p<0.05) in pigs fed the EOD diet compared with pigs fed the NC. Villus height to crypt depth ratio in the jejunum was greater (p<0.05) in pigs fed the PC and EOD diets than the NC. The numbers of E. coli in the cecum, colon and rectum were reduced (p<0.05) in pigs fed the PC and EOD diets compared with the control. In the colon, the ratio of Lactobacilli to E. coli was increased (p<0.05) in pigs fed the EOD diet compared with NC diet. Total aerobe numbers in the rectum were decreased (p<0.05) in pigs fed the PC and EOD diets compared with the control. Collectively, these results indicate that blends of essential oils could be a candidate for use as an alternative to traditional antibiotics in weaning pig diets.

BACKGROUND

We recently demonstrated that a blunt chest trauma, a strong inducer of the posttraumatic systemic inflammatory response and one of the most critical injuries in polytrauma patients, significantly delayed fracture healing in rats, possibly by the interaction of the systemic inflammation with early regeneration processes locally at the fracture site. The underlying cellular mechanisms, however, have as yet remained unknown. Therefore, the aim of this study was to analyze the cellular and morphologic composition of the early fracture callus after a blunt chest trauma.

METHODS

Rats received an osteotomy of the right femur stabilized by an external fixator in combination with a blunt chest trauma or not. The animals were killed after 3, 7, and <em>35</em> days, and the fracture calli were analyzed histologically for new tissue formation, polymorphonuclear leucocytes, macrophages, osteoclasts, and the presence of the proinflammatory cytokine <em>interleukin</em> 6.

RESULTS

The blunt chest trauma considerably increased the number of polymorphonuclear leucocytes in the callus by Day 3 compared with animals with isolated fractures. The number of macrophages was significantly reduced by the thoracic trauma at Days 3 and 7. The number of osteoclasts was not changed at any postoperative time point. After 3 days, the blunt chest trauma led to a significantly stronger <em>interleukin</em> 6 staining within the periosteal callus in zones of intramembranous ossification. During the time of cortical bridging at Day <em>35</em>, the amount of newly formed bone was significantly decreased after blunt chest trauma.

CONCLUSIONS

Our results suggest that the systemic posttraumatic inflammation induced by a thoracic trauma disturbed the inflammatory balance during the early healing stage by altering the recruitment of inflammatory cells and cytokine expression locally at the fracture site and thus impaired fracture healing. These findings provide new insights in the pathomechanisms of impaired fracture healing in patients experiencing severe trauma.

BACKGROUND

C-reactive protein (CRP), an acute phase protein that is released in response to inflammatory stimuli, is implicated in Alzheimer's disease (AD). However, the role of CRP in memory deficits associated with AD remains unclear.

OBJECTIVE

Experiments were carried out to determine whether CRP impaired memory and altered neurochemical measures associated with AD.

METHODS

The effects of intra-cerebroventricular administration of CRP or beta-amyloid peptide 25-<em>35</em> (Abeta(25-<em>35</em>)) on memory performance were evaluated using rat Morris water-maze and step-through passive avoidance tests; the levels of inflammatory cytokines (<em>interleukin</em>-1beta (IL-1beta), IL-6, and tumor necrosis factor (TNF-alpha)), endogenous CRP, and markers of the endogenous production of Abeta, including amyloid precursor protein (APP), presenilins (PS-1 and PS-2), and beta-site of APP cleaving enzyme (BACE), were also determined in brain regions using real-time reverse transcriptase polymerase chain reaction (RT-PCR) and Western blotting analysis.

RESULTS

Treatment with CRP (25.6 microg/rat) or Abeta(25-<em>35</em>) (10 microg/rat) 2 weeks ahead produced impairment of long-term memory in both animal tests. Real-time RT-PCR revealed increases in messenger RNA levels of APP, IL-1beta, IL-6, TNF-alpha, and CRP in the cerebral cortex and hippocampus and those of PS-1 and PS-2 in the cerebral cortex produced by treatment with CRP or Abeta(25-<em>35</em>). Immunoblotting analysis showed that while expression of APP was increased in both the cerebral cortex and the hippocampus, expression of IL-1beta, BACE, and TNF-alpha was increased only in the hippocampus.

CONCLUSIONS

The results suggest that CRP contributes to memory loss and early phase of pathogenesis of AD. CRP can be a novel target for therapeutic intervention of AD.

OBJECTIVE

Immune activation may have an important pathogenic role in the irritable bowel syndrome (IBS). While little is known about immunologic function in functional dyspepsia (FD), we have observed an association between cytokine secretion by peripheral blood mononuclear cells (PBMCs) and symptoms in IBS. Upper gastrointestinal inflammatory diseases are characterized by enhanced small bowel homing α4-, β7-integrin, chemokine receptor 9 (CCR9) positive T lymphocytes. We hypothesized that increased cytokine release and elevated circulating small bowel homing T cells are linked to the severity of symptoms in patients with FD. Thus, we aimed to (i) compare cytokine release in FD and healthy controls (HCs), (ii) quantify "gut homing" T cells in FD compared with HC and patients with IBS, and (iii) correlate the findings to symptom severity and gastric emptying.

METHODS

PBMC from 45 (Helicobacter pylori negative) patients with FD (Rome II) and <em>35</em> matched HC were isolated by density gradient centrifugation and cultured for 24 h. Cytokine production (tumor necrosis factor (TNF)-α, <em>interleukin</em> (IL)-1β, IL-6, IL-10) was measured by enzyme-linked immunosorbent assay. CD4+ α4β7+CCR9+ T cells were quantified by flow cytometry in FD, HC and 23 patients with IBS. Gastric emptying was measured by scintigraphy. Symptom severity was assessed utilizing the standardized Gastrointestinal Symptom Score.

RESULTS

FD patients had significantly higher TNF-α (107.2 ± 42.8 vs. 58.7 ± 7.4 pg/ml), IL-1β (204.8 ± 71.5 vs. 80.2 ± 17.4 pg/ml), and IL-10 (218 ± 63.3 vs. 110.9 ± 18.5 pg/ml) levels compared with HC, and enhanced gut homing lymphocytes compared with HC or IBS. Cytokine release and CD4+α4β7+CCR9+ lymphocytes were correlated with the symptom intensity of pain, cramps, nausea, and vomiting. Delayed gastric emptying was significantly associated (r = 0.78, P = 0.021) with CD4+α4β7+CCR9+ lymphocytes and IL-1β, TNF-α, and IL-10 secretion.

CONCLUSIONS

Cellular immune activation with increased small bowel homing T cells may be key factors in the clinical manifestations of H. pylori-negative FD.

We investigated the relationship of plasma adipocytokine concentrations with VLDL apolipoprotein B (apoB)-100 kinetics in men. Plasma adiponectin, leptin, resistin, <em>interleukin</em>-6 (IL-6), and tumor necrosis factor-alpha (TNF-alpha) concentrations were measured using enzyme immunoassays and insulin resistance by homeostasis model assessment (HOMA) score in 41 men with BMI of 22-<em>35</em> kg/m(2). VLDL apoB kinetics were determined using an intravenous infusion of 1-[(13)C]leucine, gas chromatography-mass spectrometry, and compartmental modeling. Visceral and subcutaneous adipose tissue mass (ATM) were determined using magnetic resonance imaging, and total ATM was measured by bioelectrical impedance. In univariate regression, plasma adiponectin and leptin concentrations were inversely and directly associated, respectively, with plasma triglyceride; HOMA score; and visceral, subcutaneous, and total ATMs. Conversely, adiponectin and leptin were directly and inversely correlated, respectively, with VLDL apoB catabolism and HDL cholesterol concentration (P < 0.05). Resistin, IL-6, and TNF-alpha were not significantly associated with any of these variables. In multivariate regression, adiponectin was the most significant predictor of plasma VLDL apoB concentration (P = 0.001) and, together with total or subcutaneous ATM, was an independent predictor of VLDL apoB catabolism (P < 0.001); HOMA score was the most significant predictor of VLDL apoB hepatic secretion (P < 0.05). Leptin was not an independent predictor of VLDL apoB kinetics. In conclusion, plasma VLDL apoB kinetics may be differentially controlled by adiponectin and insulin resistance, with adiponectin regulating catabolism and insulin resistance regulating hepatic secretion in men. Total body fat may also independently determine the rate of VLDL catabolism, but leptin, resistin, IL-6, and TNF-alpha do not have a significant effect in regulating apoB kinetics.

The prevalence of relative adrenal insufficiency (RAI) in critically ill cirrhosis patients with severe sepsis is over 60% and associated features include poor liver function, renal failure, refractory shock, and high mortality. RAI may also develop in noncritically ill cirrhosis patients but its relationship to the clinical course has not yet been assessed. The current study was performed in 143 noncritically ill cirrhosis patients admitted for acute decompensation. Within 24 hours after hospitalization adrenal function, plasma renin activity, plasma noradrenaline and vasopressin concentration, and serum levels of nitric oxide, <em>interleukin</em>-6 and tumor necrosis factor alpha were determined. RAI was defined as a serum total cortisol increase <9 μg/dL after 250 μg of intravenous corticotropin from basal values (<em>35</em> μg/dL. Patients were followed for 3 months. RAI was detected in 26% of patients (n = 37). At baseline, patients with RAI presented with lower mean arterial pressure (76 ± 12 versus 83 ± 14 mmHg, P = 0.009) and serum sodium (131 ± 7 versus 1<em>35</em> ± 5 mEq/L, P = 0.007) and higher blood urea nitrogen (32 ± 24 versus 24 ± 15 mg/dl, P = 0.06), plasma renin activity (7.1 ± 9.9 versus 3.4 ± 5.6 ng/mL*h, P = 0.03), and noradrenaline concentration (544 ± 334 versus 402 ± 316 pg/mL, P = 0.02). During follow-up, patients with RAI exhibited a higher probability of infection (41% versus 21%, P = 0.008), severe sepsis (27% versus 9%, P = 0.003), type-1 hepatorenal syndrome (HRS) (16% versus 3%, P = 0.002), and death (22% versus 7%, P = 0.01).

CONCLUSIONS

RAI is frequent in noncritically ill patients with acute decompensation of cirrhosis. As compared with those with normal adrenal function, patients with RAI have greater impairment of circulatory and renal function, higher probability of severe sepsis and type-1 HRS, and higher short-term mortality.

OBJECTIVE

Gain-of-function (GOF) mutations in the signal transducer and activator of transcription 1 (STAT1) result in unbalanced STAT signaling and cause immune dysregulation and immunodeficiency. The latter is often characterized by the susceptibility to recurrent Candida infections, resulting in the clinical picture of chronic mucocutaneous candidiasis (CMC). This study aims to assess the frequency of GOF STAT1 mutations in a large international cohort of CMC patients.

METHODS

STAT1 was sequenced in genomic DNA from 57 CMC patients and <em>35</em> healthy family members. The functional relevance of nine different STAT1 variants was shown by flow cytometric analysis of STAT1 phosphorylation in patients' peripheral blood cells (PBMC) after stimulation with interferon (IFN)-α, IFN-γ or <em>interleukin</em>-27 respectively. Extended clinical data sets were collected and summarized for 26 patients.

RESULTS

Heterozygous mutations within STAT1 were identified in <em>35</em> of 57 CMC patients (61%). Out of 39 familial cases from 11 families, 26 patients (67%) from 9 families and out of 18 sporadic cases, 9 patients (50%) were shown to have heterozygous mutations within STAT1. Thirteen distinct STAT1 mutations are reported in this paper. Eight of these mutations are known to cause CMC (p.M202V, p.A267V, p.R274W, p.R274Q, p.T385M, p.K388E, p.N397D, and p.F404Y). However, five STAT1 variants (p.F172L, p.Y287D, p.P293S, p.T385K and p.S466R) have not been reported before in CMC patients.

CONCLUSIONS

STAT1 mutations are frequently observed in patients suffering from CMC. Thus, sequence analysis of STAT1 in CMC patients is advised. Measurement of IFN- or IL-induced STAT1 phosphorylation in PBMC provides a fast and reliable diagnostic tool and should be carried out in addition to genetic testing.

OBJECTIVE

This study was performed to identify morphologic features of cardiac myxomas related to embolism and to provide a better understanding of the biology of these tumors, mainly in relation to their interleukin (IL)-6 expression and DNA content.

METHODS

A total of 37 cardiac myxomas were reviewed retrospectively in a clinicopathologic study that included the correlation of echocardiographic and pathologic findings in 25 cases, together with immunohistochemical evaluation of IL-6 expression and flow cytometric DNA analysis of 35 tumors.

RESULTS

There were 24 female patients and 13 male patients. The mean (+/- SD) age was 52 +/- 15 years. Fifty-four percent of patients presented with dyspnea, 51% presented with increased erythrocyte sedimentation rate (ESR), and 27% presented with embolic episodes, which were significantly associated with villous surface tumors. Atrial fibrillation was registered in 19% of patients and was significantly associated with large left atrial myxomas. Echocardiography proved to be a reliable method for preoperative diagnosis and for predicting tumor size and morphology. There was no perioperative mortality or long-term recurrences. The frequency of early surgical complications was associated with a longer mean ischemic time. Seventeen percent of tumors had abnormal DNA content, and 74% of tumors showed immunohistochemical expression of IL-6. Neither of these factors showed a significant association with embolism or constitutional illness.

CONCLUSIONS

Villous surface myxomas are related to embolism, and large left atrial tumors are related to atrial fibrillation. Echocardiography is a reliable method with which to predict tumor size and morphology. Myxoma cells usually express IL-6, and some tumors have abnormal cellular DNA content. Surgical excision of the tumor is a safe and effective treatment.

Recent reports indicate that after a peripheral nerve injury, the uninjured contralateral nerve is also affected. Because cytokines play an important role in the peripheral nerve injury, we studied the expression of five different mRNAs (<em>interleukin</em>-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha), <em>interleukin</em>-10 (IL-10), transforming growth factor-beta1 (TGF-beta1) and <em>interleukin</em>-4 (IL-4)) in the contralateral, non-operated, left sciatic nerve when the right rat sciatic nerve was transected. This study extended up to 42 days after the transection. No IL-4 expression was noted. During the first 3 days, high expression of the other studied cytokines was noted in the endoneurium. At day 7, the expression diminished to the control levels. After this, a cyclic expression pattern appeared, which was most pronounced in the endoneurium at <em>35</em> days. We also show that the expression pattern in the endoneurium is different from that in the surrounding epi- and perineurium. Also, our present study shows clearly that contralateral nerves are poor controls after injury.

BACKGROUND

Chondroitin sulfate (CS) and glucosamine sulfate (GS) are symptomatic slow-acting drugs for osteoarthritis (OA) widely used in clinic. Despite their widespread use, knowledge of the specific molecular mechanisms of their action is limited. The aim of this work is to explore the utility of a pharmacoproteomic approach for the identification of specific molecules involved in the pharmacological effect of GS and CS.

METHODS

Chondrocytes obtained from three healthy donors were treated with GS 10 mM and/or CS 200 μg/mL, and then stimulated with interleukin-1β (IL-1β) 10 ng/mL. Whole cell proteins were isolated 24 hours later and resolved by two-dimensional electrophoresis. The gels were stained with SYPRORuby. Modulated proteins were identified by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF/TOF) mass spectrometry. Real-time PCR and Western blot analyses were performed to validate our results.

RESULTS

A total of 31 different proteins were altered by GS or/and CS treatment when compared to control. Regarding their predicted biological function, 35% of the proteins modulated by GS are involved in signal transduction pathways, 15% in redox and stress response, and 25% in protein synthesis and folding processes. Interestingly, CS affects mainly energy production (31%) and metabolic pathways (13%), decreasing the expression levels of ten proteins. The chaperone GRP78 was found to be remarkably increased by GS alone and in combination with CS, a fact that unveils a putative mechanism for the reported anti-inflammatory effect of GS in OA. On the other hand, the antioxidant enzyme superoxide dismutase 2 (SOD2) was significantly decreased by both drugs and synergistically by their combination, thus suggesting a drug-induced decrease of the oxidative stress caused by IL-1β in chondrocytes.

CONCLUSIONS

CS and GS differentially modulate the proteomic profile of human chondrocytes. This pharmacoproteomic approach unravels the complex intracellular mechanisms that are modulated by these drugs on IL1β-stimulated human articular chondrocytes.

OBJECTIVE

To evaluate the presence and relative concentrations of cytokines, known to be involved in the inflammatory cascade, in acute anterior cruciate ligament (ACL) injury.

METHODS

We evaluated an extensive cytokine profile in synovial fluid from 12 patients with acute ACL injury undergoing arthroscopy compared with 15 control subjects using a BioPlex assay (Bio-Rad Laboratories, Hercules, CA) to measure the concentration of 17 inflammatory cytokines.

RESULTS

In patients with acute ACL injury compared with asymptomatic control subjects, the following cytokines were identified at significantly increased concentrations (P < .001, Mann-Whitney U test) compared with control samples: <em>interleukin</em> 6 (105 ± 72 v 0 ± 0 pg/ml), interferon γ (1,544 ± 608 v 9 ± 7.5 pg/ml), macrophage inflammatory protein 1β (16 ± 3.8 v 0.3 ± 0.2 pg/ml), and monocyte chemotactic protein 1 (<em>35</em> ± 13 v 0.5 ± 0.4 pg/ml). There was no case of a cytokine exhibiting increased levels in asymptomatic compared with symptomatic knee samples.

CONCLUSIONS

This investigation identified 4 specific cytokines (interleukin 6, interferon γ, monocyte chemotactic protein 1, and macrophage inflammatory protein 1β) out of a panel of 17 inflammatory molecules for which the levels were consistently elevated in the context of ACL injury compared with non-painful, non-acutely injured knees in a volunteer population.

METHODS

Level IV, prognostic case series.

OBJECTIVE

Activated granulocytes and inflammatory mediators of the innate immune response play fundamental roles in the pathogenesis of acute pancreatitis. We studied whether polymorphisms of interleukin-8 (IL-8) and Toll-like receptor 4 (TLR4) genes correlate with the severity of acute pancreatitis.

METHODS

Patients with acute pancreatitis (n = 92) were grouped according to the severity of the disease on the basis of the Ranson scores. Healthy blood donors (n = 200) served as controls. The IL-8 -251 gene polymorphism was analyzed by amplification-refractory mutation system; the single-nucleotide polymorphisms (Asp299Gly and Thr399Ile) of TLR4 were investigated by using a real-time polymerase chain reaction method with melting point analysis.

RESULTS

The IL-8 A/T heterozygote mutant variants were detected with a significantly higher frequency among the patients with severe pancreatitis than among the healthy blood donors (60 vs. 42%; p = 0.0264, odds ratio = 2.071, 95% confidence interval = 1.101-3.896), while the frequency of the normal allelic genotype (TT) was higher among the patients with mild pancreatitis than in the group with severe pancreatitis (35 vs. 16%; p = 0.051, odds ratio = 2.917, 95% confidence interval = 1.089-7.811). There was no significant correlation between TLR4 polymorphisms and the acute pancreatitis itself, but nonsignificantly increased frequencies of Asp299Gly and Thr399Ile heterozygotes among patients with severe infected pancreatic necrosis could be observed relative to the patients with mild pancreatitis.

CONCLUSIONS

Determination of the frequency of IL-8 polymorphism in acute pancreatitis may be informative and may provide further evidence concerning the role of IL-8 in the severe form of this disease. The possible role of TLR4 polymorphism in the outcome of severe acute pancreatitis requires further investigations in a larger series of patients.

For identification of risk factors for bloodstream infection (BSI) among neonatal intensive care unit patients, prospective 6-month studies in three neonatal intensive care units were conducted. BSI was diagnosed in 42 of 376 (11.2%) enrolled infants. Pathogens included coagulase-negative staphylococci, Candida sp., Group B streptococci and Gram-negative species. Patients with BSIs were more likely to die during their neonatal intensive care unit stay than were patients who did not acquire BSIs (6 of 42 vs. 11 of 334, P = 0.007). BSI rate was highest in infants with birth weight < 1500 g (relative risk (RR) = 6.8, P < 0.001), those treated with H-2 blockers (RR = 4.2, P < 0.001) or theophylline (RR = 2.8, P < 0.001) and those with admission diagnoses referable to the respiratory tract (RR = 3.7, P < 0.001). Infants who developed BSI were more severely ill on admission than other infants (median physiologic stability index 13 vs. 10 (P < 0.001) and were of lower gestational age (28 vs. <em>35</em> weeks, P < 0.001). In logistic regression analysis, risk of BSI was independently associated only with very low birth weight, respiratory admission diagnoses and receipt of H-2 blockers. Risk of isolation of a pathogen from blood culture was independently associated with Broviac, umbilical vein or peripheral venous catheterization>> 10, 7 or 3 days, respectively, at one insertion site. Rate of isolation of a pathogen was higher (9 of 59 (15%)) within 48 hours of a measurable serum <em>interleukin</em> 6 concentration than an <em>interleukin</em> 6 level of 0 pg/ml (10 of 159 (6%), P = 0.04).(ABSTRACT TRUNCATED AT 250 WORDS)

Many poxviruses express a <em>35</em>-40-kDa secreted protein, termed "T1" (for leporipoxviruses) or "<em>35</em>kDa" (for orthopoxviruses), that binds CC-chemokines with high affinity but is unrelated to any known cellular proteins. Many previously identified poxvirus cytokine-binding proteins display strict species ligand-binding specificity. Because the T1 and <em>35</em>kDa proteins share only 40% amino acid identity, we compared the abilities of purified myxoma virus-T1 (M-T1) and vaccinia virus (strain Lister)- and rabbitpox virus-<em>35</em>kDa proteins to inhibit human CC-chemokines in vitro. All three proteins were equally effective in preventing several human CC-chemokines from binding to target chemokine receptors and blocking subsequent intracellular calcium release. The inhibitory affinities were comparable (Ki = 0.07-1.02 nM). These proteins also displayed similar abilities to inhibit (IC50 = 6.3-10.5 nM) human macrophage inflammatory protein-1alpha-mediated chemotaxis of human monocytes. None of the viral proteins blocked <em>interleukin</em>-8-mediated calcium flux or chemotaxis of human neutrophils, confirming that the biological specificity of the T1/<em>35</em>kDa family is targeted inhibition of CC-chemokines. Despite the significant sequence divergence between the leporipoxvirus T1 and orthopoxvirus <em>35</em>kDa proteins, our data suggest that their CC-chemokine binding and inhibitory properties appear to be species nonspecific and that the critical motifs most likely reside within the limited regions of conservation.

Although studies indicate that simple hemorrhage induces profound depression of cell-mediated immunity and enhances the host's susceptibility to sepsis, the mechanism for this remains unknown. Since the Kupffer cells (KC) are positioned to have constant exposure to various immunomodulators and antigens released during hypotension, we have examined whether antigen presentation by KC, a critical component in eliciting an antigen specific immune response or those processes associated with it, are depressed following hemorrhage. C3H/HeN mice were bled to and maintained at a mean BP of <em>35</em> mmHg for 60 min, and then resuscitated with their own blood and adequate fluids. The mice were killed at varying periods of time after hemorrhage to obtain KC from the liver, and assessed for their capacity to present antigen to a sensitized clone Th/cell line (D10.G4.1). Hemorrhaged mice exhibited a marked decrease in antigen presenting capacity beginning as little as 2 h and lasting up to 3-5 days post-hemorrhage. The ability of KC to express mouse <em>interleukin</em> 1 (mIL-1) showed a significant decline at 2 h following hemorrhage, but this effect was not apparent at 24 h post-hemorrhage. In contrast, KC capacity to produce IL-1, IL-6 and tumour necrosis factor (TNF) (cytokines which can co-stimulate T cell antigen presentation) was markedly enhanced during the first 24 h following hemorrhage. A marked decrease was observed in both the mean of the average fluorescence per KC and the percent of Ia antigen-positive KC which persisted for at least 3 days after hemorrhage. The ability of ibuprofen (a cyclooxygenase blocker) to partially restore the antigen presenting capacity of KC from hemorrhaged mice in vitro indicates that prostaglandins are involved in this dysfunction. Thus, the depression of KC antigen presentation, as well as the enhanced capacity of these cells to release inflammatory mediators (TNF, IL-1, IL-6 and prostanoids) which may produce cell and organ dysfunction, could contribute to the host's enhanced susceptibility to sepsis following hemorrhage.

OBJECTIVE

To determine the relation between lower airway infection and inflammation, respiratory symptoms, and lung function in infants and young children with cystic fibrosis (CF).

METHODS

A prospective study of children with CF aged younger than 3 years, diagnosed by a newborn screening programme. All were clinically stable and had testing as outpatients. Subjects underwent bronchial lavage (BL) and lung function testing by the raised volume rapid thoracoabdominal compression technique under general anaesthesia. BL fluid was cultured and analysed for neutrophil count, interleukin 8, and neutrophil elastase. Lung function was assessed by forced expiratory volume in 0.5, 0.75, and 1 second.

RESULTS

Thirty six children with CF were tested on 54 occasions. Lower airway infection shown by BL was associated with a 10% reduction in FEV(0.5) compared with subjects without infection. No relation was identified between airway inflammation and lung function. Daily moist cough within the week before testing was reported on 20/54 occasions, but in only seven (35%) was infection detected. Independent of either infection status or airway inflammation, those with daily cough had lower lung function than those without respiratory symptoms at the time of BL (mean adjusted FEV(0.5) 195 ml and 236 ml respectively).

CONCLUSIONS

In young children with CF, both respiratory symptoms and airway infection have independent, additive effects on lung function, unrelated to airway inflammation. Further studies are needed to understand the mechanisms of airway obstruction in these young patients.

The pI-6.8 species of normal human <em>interleukin</em> 1 (IL-1) has been isolated by ion-exchange and reverse-phase high-performance liquid chromatography. The isolated material had a molecular weight of 18,000, and had a specific bioactivity of 1.7 X 10(7) half-maximal U/mg in the murine thymocyte proliferation assay, values similar to those obtained for murine P388D1-derived IL-1 (12), and human IL-1 isolated by a previously published purification protocol (15). Amino-terminal sequence analysis revealed a single N-terminal, and resulted in the identification of 30 of the first <em>35</em> amino acid residues. Sequence of three CNBr cleavage fragments of purified IL-1 resulted in the identification of an additional 38 residues. All of the sequences agree exactly with those deduced from complementary DNA (cDNA) by Auron, et al. (18), demonstrating that this cloned cDNA, though considerably different from the cDNA reported for murine IL-1 (12), nevertheless codes for the pI-6.8 species of human IL-1. The evidence also shows that the precursor protein for human IL-1 is largely processed at the N-terminal end. Little or no processing occurs at the carboxy-terminal end. Sequence homology with interferon-inducing factor (26) suggests that the pI-6.8 species of human IL-1 is a member of a gene family. Although equally potent in the murine thymocyte proliferation assay, murine IL-1 and the pI-6.8 species of human IL-1 are structurally distinct. Further study will answer the interesting question as to the relationship of the other charged species of human IL-1 to these distinct IL-1 classes.

OBJECTIVE

Allograft rejection involves multiple effector mechanisms. <em>Interleukin</em>(IL)-12 family members play a critical role in influencing helper T-cell differentiation and inflammatory processes, and their respective role in orchestrating inflammation of autoimmune or infectious origin starts to be unravelled. We highlight recent findings on the function of the different IL-12 family members: IL-12p70, IL-23, IL-27 and IL-<em>35</em> and discuss their possible involvement in influencing the balance between graft rejection and tolerance.

RESULTS

The capacity of dendritic cells to produce IL-12 and IL-23 strongly influences the outcome of CD4 T-cell responses. While the IL-12/interferon-gamma axis has classically been involved in autoimmune pathologies and acute graft rejection, it is now clear that it also displays immunoregulatory properties. In contrast, IL-23 promotes the function of proinflammatory IL-17-producing cells in both mice and humans. Both IL-27 and IL-<em>35</em> have recently emerged as important regulators of adaptive immune responses.

CONCLUSIONS

The contribution of the IL-12/interferon-gamma pathway to acute graft rejection may be more complicated than initially thought. As our understanding of the IL-12 family is rapidly growing and changing, the respective role of its members in orchestrating innate and adaptive immune responses toward alloantigens should be addressed.

OBJECTIVE

Chorioamnionitis is associated with increased risk for cerebral palsy (CP) in term infants. A functional polymorphism in the interleukin-6 (IL-6) gene has been implicated in newborn brain injury. We studied whether the IL-6 -174 G/C polymorphism confers increased risk for CP in term infants.

METHODS

This population-based case-control study included 334,333 live-born infants born at>>or=36 weeks gestation within Kaiser Permanente Medical Care Program from 1991 to 2002. Case patients (n = 250) were identified from electronic records and confirmed by chart review, and comprised all infants with spastic or dyskinetic CP not caused by developmental abnormalities who had a neonatal blood specimen available for study. Control patients (n = 305) were randomly selected from the study population.

RESULTS

Compared with genotype GG, the less common CC genotype was associated with increased risk for overall CP (odds ratio [OR], 2.6; 95% confidence interval [CI], 1.5-4.6), quadriparetic CP (OR, 4.1; 95% CI, 1.8-9.3), and hemiparetic CP (OR, 2.7; 95% CI, 1.3-5.7), after controlling for race. The C allele conferred increased risk for CP in both recessive and additive genetic models. In multivariate analysis controlling for race, independent risk factors for CP included CC genotype compared with GG (OR, 2.4; 95% CI, 1.3-4.4), clinical chorioamnionitis (OR, 4.6; 95% CI, 2.1-10.4), maternal age>>or= 35 (OR, 2.6; 95% CI, 1.6-4.1), and male sex (OR, 1.6; 95% CI, 1.1-2.4).

CONCLUSIONS

Our data suggest that a functional polymorphism in the IL-6 gene is a risk factor for CP among term and near-term infants.

OBJECTIVE

Idiopathic vaginal bleeding, a common complication of pregnancy, increases the risk of small-for-gestational age (SGA) neonate, preeclampsia and preterm delivery and can be the only clinical manifestation of intra-amniotic infection and/or inflammation (IAI). Placenta previa is thought to be protective against ascending intrauterine infection, yet an excess of histologic chorioamnionitis has been reported in this condition. The aim of this study was to determine the frequency and clinical significance of IAI in women with placenta previa and vaginal bleeding in the absence of preterm labor.

METHODS

A retrospective cohort study including <em>35</em> women with placenta previa and vaginal bleeding <37 weeks of gestation who underwent amniocentesis was undertaken. Patients with multiple gestations were excluded. Intra-amniotic infection was defined as a positive culture for microorganisms, and intra-amniotic inflammation as an elevated amniotic fluid <em>interleukin</em> (IL)-6 concentration. IL-6 concentrations were determined by ELISA in 28 amniotic fluid samples available. Non-parametric statistics were used for analysis.

RESULTS

1) The prevalence of intra-amniotic infection was 5.7% (2/<em>35</em>), and that of IAI was 17.9% (5/28); 2) the gestational age at delivery was lower in patients with IAI than in those without IAI [29.4 weeks, interquartile range (IQR): 23.1-34.7 vs. <em>35</em>.4 weeks, IQR: 33.9-36.9; P=0.028]; and 3) patients with placenta previa and IAI had a higher rate of delivery within 48 h (80% (4/5) vs. 19% (4/21); P=0.008) than those without IAI.

CONCLUSIONS

Patients with placenta previa presenting with vaginal bleeding have intra-amniotic infection in 5.7% of the cases, and IAI in 17.9%. IAI in patients with placenta previa and vaginal bleeding is a risk factor for preterm delivery within 48 h.

Oestrogen (17β-oestradiol, E₂) is a highly effective treatment for experimental autoimmune encephalomyelitis (EAE) that may potentiate Foxp3+ regulatory T (Treg) cells, which in turn limit the expansion of encephalitogenic T-cell specificities. To determine if Treg cells constitute the major non-redundant protective pathway for E₂, we evaluated E₂ protection of EAE after targeted deletion of Foxp3 expression in Foxp3-DTR mice. Unexpectedly, E₂-treated Foxp3-deficient mice were completely protected against clinical and histological myelin oligodendrocyte glycoprotein (MOG)-<em>35</em>-55 peptide-induced EAE before succumbing to diphtheria toxin-induced mortality. This finding indicated the presence of alternative E₂-dependent EAE-protective pathways that could compensate for the lack of Treg cells. Further investigation revealed that E₂ treatment inhibited proliferation and expression of CCL2 and CXCL2, but enhanced secretion of <em>interleukin</em>-10 (IL-10) and IL-13 by MOG-<em>35</em>-55-specific spleen cells. These changes occurred concomitantly with increased expression of several chemokines and receptors, including CXCL13 and CXCR5, and the negative co-activation molecules, PD-L1 and B7.2, by B cells and dendritic cells. Furthermore, E₂ treatment resulted in higher percentages of spleen and lymph node T cells expressing IL-17, interferon-γ and tumour necrosis factor-α, but with lower expression of CCR6, suggesting sequestration of MOG-<em>35</em>-55 peptide-specific T cells in peripheral immune organs. Taken together, these data suggest that E₂-induced mechanisms that provide protection against EAE in the absence of Foxp3+ Treg cells include induction of regulatory B cells and peripheral sequestration of encephalitogenic T cells.

BACKGROUND

COPD is characterized by a multi-component character involving a state of low-grade systemic inflammation and an increased prevalence of cardiovascular co-morbidity. The role of circulating leptin and other adipokines in the involvement of the systemic inflammation in COPD is only studied scarcely.

OBJECTIVE

To investigate gender related differences in the adipokine metabolism in relation to systemic inflammatory biomarkers in clinically stable subjects with COPD.

METHODS

In total, 91 clinically stable COPD patients and <em>35</em> healthy control subjects, matched for body mass index (BMI) with the COPD subjects, were included. Lung function measurement and body composition were performed in patients with COPD. In the total group, plasma concentration of the adipokines (leptin, adiponectin and resistin) and systemic inflammatory biomarkers C-reactive protein (CRP), <em>interleukin</em> 6 (IL-6), tumor necrosis factor α (TNFα), and its soluble receptors 55 and 75 (sTNFα-R55, R75) were analyzed.

RESULTS

The COPD group was characterized by increased levels of CRP, IL-6 and leptin. Plasma adiponectin and resistin concentrations were not different between the COPD and the control group. Within the COPD group, there was a significant interaction between gender and BMI on the leptin/fat mass ratio. In COPD women, a significant correlation between leptin and CRP was present.

CONCLUSIONS

In men with clinically stable COPD, leptin, adiponectin and resistin appear to be physiologically regulated, while in women, leptin metabolism is altered. Leptin secretion is increased in COPD women when compared to healthy women and compared to COPD men, and to a greater extent in overweight women with COPD.