Search

Intracellular signalling in blood leucocytes shows multiple aberrations in acute pancreatitis (AP) complicated by organ dysfunction (OD). We studied whether the aberrations associate with severity of AP and occur in sepsis complicated by OD. The study comprises 14 sepsis patients (11 with shock), 18 AP patients (nine mild; six moderately severe; three severe) and 28 healthy volunteers. Within 48 h after admission to hospital, phosphorylation of nuclear factor-ĸB (NF-ĸB), signal transducers and activators of transcription (STATs) 1,3, and extracellular signal-regulated kinases 1/2 were measured from stimulated or non-stimulated leucocytes using phosphospecific whole blood flow cytometry. In sepsis, as compared with healthy subjects, phosphorylated NF-ĸB levels of monocytes promoted by bacterial lipopolysaccharides, tumour necrosis factor or Escherichia coli cells were lower (P < 0.001 for all), pSTAT1 levels of monocytes promoted by IL-6 were lower (P < 0.05 for all), and STAT3 was constitutively phosphorylated in monocytes, neutrophils and lymphocytes (P < 0.001 for all). In AP, severity was associated with proportions of pSTAT1-positive monocytes and lymphocytes promoted by IL-6 (P < 0.01 for both), constitutive STAT3 phosphorylation in neutrophils (P < 0.05), but not with any of the pNF-ĸB levels. Monocyte pSTAT3 fluorescence intensity, promoted by IL-6, was lower in sepsis and AP patients with OD than in AP patients without OD (P < 0.001). Collectively, signalling aberrations in sepsis with OD mimic those described previously in AP with OD. Possibility that aberrations in STAT1 and STAT3 pathways provide novel markers predicting evolution of OD warrants studies including patients presenting without OD but developing it during follow-up.

Activated protein C (APC), an endogenous anticoagulant, has antithrombotic, fibrinolytic and anti-inflammatory properties. We recently conducted a controlled study (APCAP, activated protein C in severe acute pancreatitis) of APC treatment of patients with severe acute pancreatitis (SAP). Here we studied the effect of APC on the pivotal coagulation parameters of the surviving patients in the APCAP study.The study consisted of 20 patients of whom 10 patients had received APC and 10 patients had received placebo. Coagulation parameters, physiological anticoagulants, thrombograms and circulating levels of IL-6 and CRP were determined on admission and at days 1, 3-4 and 6-7.During follow-up, the temporal levels of prothrombin time (PT) decreased and the temporal levels of thromboplastin time (TT) increased in placebo group (p0.001 for both), but not in APC group. The temporal levels of antithrombin (AT) increased less in APC group than in placebo group (p = 0.011). The shapes of the SAP patients' thrombograms were strongly deranged and were marginally affected by APC treatment.Coagulopathy in SAP, a complex phenomenon, is not alleviated by APC treatment. Rather, the patients receiving APC are heading toward normal homeostasis of coagulation slower than patients receiving placebo.

Objective:New biomarkers are needed to better predict the severity of acute pancreatitis. CD73/ecto-5’-nucleotidase is an enzyme that generates adenosine, which dampens inflammation and improves vascular barrier function in several disease models. CD73 also circulates in a soluble form in the blood.

It has been proposed that the Akt kinase pathway provides a regulatory mechanism to limit the inflammatory response. We examined the activation of Akt upon LPS challenge in monocytes of patients with rheumatoid arthritis (RA), and correlated it with disease activity. Twelve subjects with recent-onset, DMARD-naive RA, thirteen patients with chronic, DMARD therapy -nonresponding RA and 27 healthy volunteers provided whole blood samples for phosphospecific flow cytometric measurement of unstimulated and LPS-stimulated Akt phosphorylation at serine 473 in monocytes, determined in relative fluorescence units (RFU). Activation capability, i.e. responsiveness of monocytes, was determined as the difference between stimulated and unstimulated samples, and compared between groups using Mann-Whitney test. CRP and ESR, swollen and tender joint counts, patients’ global assessment of disease activity, DAS28 score, and plasma IL-6 determined by ELISA were correlated with Akt activation using Spearman method. Median (interquartile range) Akt activation capability was significantly lower in DMARD-naive (379 RFU [285, 432],], p=0.016) and even lower in DMARD-nonresponding RA (258 RFU [213, 338], p

Background The components of nucleosomes, which contain DNA and histones, are released into the circulation from damaged cells and can promote inflammation. We studied whether the on-admission levels of circulating nucleosomes predict the development of severe acute pancreatitis (AP), in particular among the patients who present without clinical signs of organ dysfunction. Methods This is a prospective study of 74 AP patients admitted to Helsinki University Hospital from 2003 to 2007. Twenty-three patients had mild, 27 moderately severe, and 24 severe AP as defined by the revised Atlanta criteria. 14/24 severe AP patients had no sign of organ dysfunction on admission (modified marshall score

Reactive arthritis (ReA) is a sterile joint inflammation triggered by a remote infection and associated with human leucocyte antigen (HLA)-B27. Its pathogenesis is unknown, but abnormal response to microbial structures or endogenous inflammatory mediators may be involved. We studied responses in leucocyte signalling profiles in patients with previous ReA after a full recovery.The study comprised 10 HLA-B27-positive healthy subjects with a history of Yersinia enterocolitica-triggered ReA (B27+ReA+) and 20 healthy reference subjects, of whom 10 carried HLA-B27 (B27+ReA-) and 10 did not (B27-ReA-). Phosphospecific fluorescent monoclonal antibodies and flow cytometry were used to determine activation of nuclear factor kappa B (NF-κB), signal transducers and activators of transcription (STATs) 1, 3, 5, and 6, and two mitogen-activated protein (MAP) kinases, p38 and extracellular signal-regulated kinase (ERK)1/2, in monocytes, lymphocytes, lymphocyte subsets, and neutrophils. B27+ReA+ and B27-ReA- whole-blood samples were incubated with Yersinia with or without infliximab to study the role of tumour necrosis factor (TNF) in lymphocyte subset activation. Samples of the three subject groups were studied using soluble bacterial or endogenous stimuli. Fluorescence levels were determined as relative fluorescence units (RFU) and the proportion of positively fluorescing cells.The intracellular activation of circulating leucocytes in response to soluble stimuli was consistently comparable in B27+ReA+, B27+ReA-, and B27-ReA- subjects. Infliximab inhibited Yersinia-induced lymphocyte NF-κB phosphorylation similarly in B27+ReA+ and B27-ReA- groups.ReA susceptibility is not reflected in leucocyte signalling profiles elicited by phlogistic stimuli. However, the possibility remains that aberrations occur in response to combinations of stimuli, such as those associated with leucocyte adhesion.

The study aimed to determine the effect of the activated protein C on the course of systemic inflammation in the APCAP (activated protein C in acute pancreatitis) trial where we randomized 32 patients with severe acute pancreatitis to receive either recombinant activated protein C (drotrecogin alfa activated) (n=16) or placebo (n=16) for 96 hours. In the present study, we present the time course of the patients’ plasma or serum levels of soluble markers (IL-8, IL-6, IL-10, IL-1ra, sE-selectin, PCT) and monocyte and neutrophil cell surface (CD11b, CD14, CD62L, HLA-DR) markers of systemic inflammatory response during the first 14 days after the randomization. The results of the intervention and placebo groups were comparable showing that recombinant APC treatment did not alter the course of systemic inflammation in severe acute pancreatitis. Our finding is in accordance with the clinical findings in the APCAP trial indicating that the intervention did not affect evolution of multiple organ dysfunctions.

Tumour necrosis factor (TNF) receptor-associated periodic syndrome (TRAPS) is a dominantly inherited autoinflammatory disease caused by heterozygous mutations in the TNFRSF1A gene encoding for the TNF receptor 1 (TNFR1). TRAPS is a multi-faceted and heterogeneous disease which commonly manifests as recurrent episodes of high fever accompanied by abdominal pain, pleurisy, migratory rash, and myalgia. Disease attacks occur spontaneously or may be elicited by minor triggers. Because of a vigorous and sustained acute-phase response it may be complicated by systemic AA amyloidosis. Therapeutically interleukin-1 blockade seems even more promising than TNF blockade. Studies on the pathogenesis of TRAPS have shown TNFα-dependent cellular signalling to be defective, an enigmatic finding considering the hyperinflammatory phenotype of the disease. Several studies indicate that most mutated receptors never reach the cell surface but are misfolded and trapped in the endoplasmic reticulum, where they may elicit an intracellular inflammatory response, and thus lead to constitutional expression of proinflammatory cytokines. The aim of this review is to describe the current understanding of the pathogenesis of TRAPS by integrating recent clinical and laboratory data.

Background: Preeclampsia and preterm labor often underlie preterm birth, and are associated with maternal inflammation. In preterm infants, respiratory distress syndrome (RDS) and mechanical ventilation are associated with systemic inflammation. Objective: We aimed to study whether early-onset preeclampsia or preterm labor modulate the systemic inflammation affecting preterm infants with RDS. Methods: We recruited mechanically ventilated infants with gestational ages Results: As compared with infants born after preterm labor, infants born after early-onset preeclampsia had higher CD11b expression on days 1–6 on both neutrophils and monocytes. In addition, infants born after early-onset preeclampsia had higher CRP concentrations on days 2–6 (all p < 0.05). Conclusions: As compared with infants born after preterm labor to mothers without preeclampsia, infants born after early-onset preeclampsia presented with a stronger postnatal systemic inflammatory reaction. Antenatal exposure to preeclampsia may induce fetal leukocyte priming and regulation of inflammation, and thereby modify postnatal inflammatory reactions and morbidity.

Ilmakunnas M, Hockerstedt K, Makisalo H, Siitonen S, Repo H, Pesonen EJ. Hepatic IL-8 release during graft procurement is associated with impaired graft function after human liver transplantation. Clin Transplant 2010: 24: 29–35. © 2009 John Wiley & Sons A/S. Abstract: In experimental models, brain death induces inflammatory cascades, leading to reduced graft survival. Thus far, factors prior to graft preservation have gained less attention in clinical setting. We studied pre-preservation inflammatory response and its effects on graft function in 30 brain dead liver donors and the respective recipients. Before donor graft perfusion, portal and hepatic venous blood samples were drawn for phagocyte adhesion molecule expression and plasma cytokine determinations. Donor intensive care unit stay correlated with donor C-reactive protein (R = 0.472, p = 0.013) and IL-6 (R = 0.419, p = 0.026) levels, and donor (R = 0.478, p = 0.016) and recipient gamma-glutamyl transferase (R = 0.432, p = 0.019) levels. During graft procurement, hepatic IL-8 release was observed in 17/30 donors. Grafts with hepatic IL-8 release exhibited subsequently higher alkaline phosphatase [319 (213–405) IU/L vs. 175 (149–208) IU/L, p = 0.006] and bilirubin [101 (44–139) μmol/L vs. 30 (23–72) μmol/L, p = 0.029] levels after transplantation. Our findings support the concept that inflammatory response in the brain dead organ donor contributes to the development of graft injury in human liver transplantation.

OBJECTIVES: To study in patients with acute pancreatitis (AP) the plasma soluble form of the receptor for advanced glycation end products (sRAGE) and high-mobility group box chromosomal protein 1 (HMGB1) levels, followed-up for 12 days after hospitalization, in relation to the occurrence of organ failure and mortality. METHODS: Thirty-eight patients with severe AP and organ failure (grade 2). A control group (127 patients) consisted of 38 patients with severe AP without organ failure (grade 1) and 89 patients with mild AP (grade 0). Plasma samples for determination of HMGB1 and sRAGE levels were collected on admission and on days 1 and 2, days 3 and 4, and days 7 and 12 after admission. RESULTS: The median of the highest sRAGE levels was higher in grade 2 patients (472 pg/mL; interquartile range [IQR], 259-912) than in grade 0 plus grade 1 patients (349 pg/mL; IQR, 209-544; P = 0.024). Among the patients with detectable HMGB1, the median of the highest HMGB1 levels was 117 ng/mL (IQR, 56-212; n = 24) in grade 2 patients and 87 ng/mL (IQR, 54-161; n = 62) in grade 0 plus grade 1 patients (P = 0.310). CONCLUSIONS: We demonstrate that sRAGE level, but not HMGB1 level, is significantly higher in AP patients who develop organ failure than in AP patients without organ failure who recover.

In this Case Study, the authors describe a patient with Poncet disease, a rare form of polyarthritis associated with tuberculosis infection. They discuss their application of interferon-γ-release assays and PCR specific for DNA of theMycobacteria tuberculosis complex to make the diagnosis, despite a negative M. tuberculosisculture. Background. A 55-year-old, HLA-B27-positive Finnish woman presented with migratory, sterile polyarthritis. Investigations. Physical examination, chest radiography, serologic testing, microscopy, M. tuberculosis-specific interferon γ enzyme-linked immunospot (ELISPOT) assay, smear and culture of synovial fluid for acid-fast bacilli, and PCR. Diagnosis. The patient's assayed blood and synovial fluid lymphocytes were reactive, and the numbers of M. tuberculosis-specific T cells, as determined by ELISPOT, were twofold to sixfold higher in synovial fluid than in blood. Cultures for acid-fast bacilli remained negative, while PCR specific for DNA of the M. tuberculosis complex was positive in synovial fluid cells. These results suggested that the patient had either active or latent M. tuberculosis infection. This finding, coupled with the presence of polyarthritis, led to a diagnosis of Poncet disease. Management. Standard antituberculous therapy consisting of isoniazid, rifampicin and pyrazinamide was given for 2 months, followed by isoniazid and rifampicin for 4 months. Inflamed joints were treated with methylprednisolone injections. The polyarthritis resolved within 4 months of initiating antituberculous therapy.

Secondary immune response to tetanus toxoid (TT), monocyte function, and generation of suppressor cells were studied in patients with previous yersinia arthritis (YA) and healthy controls. A comparison of HLA-B27 positive and negative subjects revealed that leukocytes from the former showed significantly higher responses to TT but not to a variety of other antigens in a lymphocyte proliferation test, and higher rates of migration in a leukocyte migration inhibition assay in the absence of TT. The enhanced migration of leukocytes supports the concept that hyper-reactive neutrophils contribute to inflammatory symptoms in HLA-B27 positive subjects, irrespective of previous YA. No correlation was found between HLA-DR specificities and YA. However, HLA-DR2/HLA-B7 was associated with high suppressor cell activity and low serum levels of anti-TT-antibodies. This accords with the view that HLA-DR antigens play a role in the regulation of immune response.

Monosomy 7, a deletion of the long arm of chromosome 7, was shown in the neutrophils of peripheral blood in 5 of 6 patients with a myelodysplastic syndrome or leukemia who had monosomy 7 in their bone marrow cells. In a chemiluminescence assay the production of reactive oxygen species by neutrophils from patients was increased in whole blood and decreased in purified cells, which suggests that neutrophils with monosomy 7 tolerate poorly the cell purification procedures used. The in vitro migration of purified neutrophils obtained from patients with monosomy 7 was impaired. It is known that patients with monosomy 7 have an increased susceptibility to infections. It is possible that neutrophils with monosomy 7 are too easily triggered to a full-scale respiratory burst and thereby the cells exhaust their ability to eliminate invading microbes efficiently.

Background. Experimental models of hepatic ischemia/reperfusion injury have implicated a pathophysiologic role for neutrophils in subsequent hepatocellular damage. In human liver transplantation, however, the effect of reperfusion-induced neutrophil activation on initial graft function is not clear. Methods. In 38 patients undergoing liver transplantation, neutrophil CD11b and L-selectin expression, neutrophil count, and plasma lactoferrin levels were measured. To assess changes within the graft during initial reperfusion, samples of blood entering and leaving the graft were obtained simultaneously, and transhepatic ratio calculated (hepatic vein/portal vein; 1 denotes no change, 1 an increase across the liver). Graft steatosis, postoperative liver function, and outcome were recorded. Associations between neutrophil activation markers and outcome measures were evaluated. Results. Substantial hepatic neutrophil activation occurred during initial reperfusion, demonstrated by concomitant L-selectin shedding and CD11b upregulation (transhepatic ratios 0.9 [0.7-1.0]; 1.4 [0.9-1.9]; both P

Objectives: Genotype assessment has been suggested to be a tool for predicting disease severity in acute pancreatitis (AP). To study this hypothesis, we performed genotype analysis of tumor necrosis factor (TNF) −308 A/G, CD14 −159C/T, and HSPA1B +1267 A/G polymorphisms. Methods: This is a case-control association study of 397 patients with AP (214 of whom had an alcohol-induced AP) and 300 controls. The control group comprised 218 subjects with detailed data of alcohol consumption, 70 of whom were heavy drinkers (daily alcohol intake >40 g), and 92 blood donors. The severity of AP was determined according to the Atlanta classification. Genotyping was performed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry-assisted genotyping method. Results: Major allele frequency in TNF gene was 0.87 for patients with AP and 0.86 for controls. For CD14, the gene major allele frequency was 0.60 for patients and 0.63 for controls. For HSPA1B, the major allele frequencies were 0.52 for patients and 0.49 for controls, respectively. The allele frequencies did not differ significantly between AP patients with organ failure and those with mild disease, patients with alcohol-induced AP, or those with biliary AP. The patients with septic infectious complications (n = 47) had genotype distribution no different from those with mild, uncomplicated disease (n = 245). Conclusions: The TNF, CD14, and HSPA1B polymorphisms studied seem not to play a role in determining the severity of AP or the risk of alcohol-induced AP and thus do not serve as a tool for predicting disease severity.

Complement activation occurs in patients with acute pancreatitis (AP) and may contribute to the development of organ failure. Because a number of enzymes are released during AP that could influence the complement inhibitor CD59, the purpose of this study was to examine serum levels of CD59 in relation to severity of AP.Twelve patients with severe AP had organ failure (referred to as the grade 2 group). For each of them, we found 2-3 age-matched AP patients who served as controls (n=27). Of these, a total of 13 had mild AP (grade 0 group) and 14 severe AP without organ failure (grade 1 group). Blood samples were collected at admission and on days 1 and 3-7 post-admission. Grade 2 patients were compared with grade 0 and grade 1 patients. CD59 levels were measured by a sandwich enzyme immunoassay.At admission, median CD59 levels were significantly higher (p = 0.002) in grade 2 patients (median 104.2 ng/ml, range 26.1-186.3) than in grade 0 patients (37.3, range 30.3-75.9) and grade 1 patients (38.6, range 19.9 96.1). CD59 levels remained higher in grade 2 patients than in grade 0 and 1 patients on day 1 (p = 0.001) and days 3-7 (p = 0.002). The CD59 levels correlated significantly (p0.05) with C-reactive protein (CRP) levels (R = 0.40) and APACHE II scores (R = 0.32) on admission.Organ failure and severity of AP are associated with elevated serum levels of CD59.

Monocyte membrane CD14 (mCD14) and soluble CD14 (sCD14) both associate with poor outcome in sepsis. Because the value of combined use of the markers is unknown we measured both in patients with severe community acquired infections. The study comprised 142 acutely ill patients with community acquired pneumonia and/or blood culture-positive sepsis. Expression of mCD14 was measured, on admission to hospital, by whole blood flow cytometry and sCD14 by ELISA. There was no significant correlation between mCD14 and sCD14. Patients in the lowest tertile of mCD14 were 9.79 times (95% CI 1.31- >50, p =0.006) more likely to die than patients in the middle/highest tertiles. Survival rates in the highest and middle/lowest tertiles of sCD14 levels were comparable. After stratification by sCD14, patients in the lowest tertile of mCD14 were 14.4 times (95% CI 1.90-39.44) more likely to die than patients in the middle/highest tertiles. A significant positive correlation was detected between C-reactive protein and sCD14 levels, providing evidence that sCD14 may serve as an acute phase reactant. In conclusion, low monocyte mCD14 level, unlike the concurrent sCD14 level, predicts 28-d mortality in patients with community acquired infections.

In reperfusion injury activation of coagulation and inflammation contribute to organ dysfunction. Activated protein C (APC) exhibits anticoagulant and anti-inflammatory properties in models of reperfusion injury. We hypothesized that APC could be cardioprotective after ischemia and cardiopulmonary bypass (CPB).20 pigs, undergoing 120 min of CPB and aortic cross-clamping, were randomized to receive 1 mg of human APC or placebo to the last cardioplegic solution given 15 min before declamping to the systemic circulation. After aortic declamping the heart was supported by continuing CPB for 30 min followed by 30 min surveillance. Thrombin-antithrombin complexes, neutrophil L-selectin expression in blood and myeloperoxidase activity (MPO) of myocardial biopsies were measured.There was no indication of APC-induced increased bleeding. Thrombin levels were significantly lower in the APC group than in the placebo group and so were the rates of thrombin formation during the first 3 min of reperfusion and between 10 and 30 min after declamping. There were no differences in MPO or in the proportion of L-selectin (+) to L-selectin (-) neutrophils between groups. Significant systolic hypotension in the APC group was observed at 30 and 45 min compared with the placebo group which associated with the increased mortality observed in the APC group (p = 0.019).Human APC in cardioplegic solution during CPB in pigs, decreased reperfusion induced thrombin formation with no associated bleeding. No anti-inflammatory effects of human APC were seen. However, in this setting, APC caused hemodynamic deterioration. The observed phenomenon could be explained by systolic hypotension potentially produced by the release of vasoactive substances generated by the APC activation of PARs in the endothelium.

Obesity is a risk factor for a severe form of acute pancreatitis (AP). Because the underlying mechanisms are poorly known, we studied relationship between the severity of AP and plasma levels of leptin and adiponectin, 2 adipokines regulating the course of systemic inflammation.The study comprises 12 patients with severe AP and 12 control patients with mild AP matched by age (+/-10 years), body mass index (+/-3 kg/m), sex, and etiology of AP. Quantikine Human Adiponectin and Quantikine Human Leptin Immunoassays (RD Systems, Minneapolis, Minn) were used to measure the adipokine levels in the patients' plasma on admission and during the hospital stay.Median leptin concentrations on admission were 6.1 ng/mL (range, 1.6-72.9 ng/mL) in the severe AP group and 9.0 ng/mL (range, 2.5-36.3 ng/mL) in the mild AP group (P0.05). In severe AP, the value at days 2 to 4 (7.7 ng/mL; range, 1.6-13.9 ng/mL) did not differ from respective on-admission value (P0.05). In mild AP, the value at days 2 to 4 (3.8 ng/mL; range, 1.6-12.9 ng/mL) was lower than the respective on-admission value (P = 0.005). Adiponectin concentrations on admission were 5642 ng/mL (range, 1201-19,400 ng/mL) for severe AP and 6314 ng/mL (range, 1980-24,340 ng/mL) for mild AP (P0.05). Maximum variation of adiponectin level (the highest value minus the lowest value) was greater in severe AP than in mild AP (P = 0.001).In patients matched by age, sex, body mass index, and etiology, the on-admission plasma levels of adiponectin and leptin do not correlate with disease severity, suggesting that the adipokines do not affect the course of AP.

Objective: To explore the potential anti-inflammatory role of protein C pathway in ischemia-reperfusion injury during liver transplantation. Design: Prospective, observational clinical study. Setting: Tertiary teaching hospital. Patients: Fifty adult patients undergoing liver transplantation for acute liver failure or chronic liver disease. Interventions: To assess changes occurring across the transplanted liver, samples of blood entering and leaving the graft were obtained simultaneously from portal and hepatic veins. Plasma protein C and activated protein C levels, neutrophil and monocyte CD11b and L-selectin expression, and leukocyte differential counts were measured. Postoperative liver function and outcome of transplantation were recorded. Measurements and Main Results: During reperfusion, protein C became entrapped within the graft (portal vein 49% [20-96%]; graft caval effluent 25% [12-76%], p

OBJECTIVES Severe acute pancreatitis (AP) is frequently associated with immune suppression, which increases the risk of infections, organ failure, and death. Our aims were to measure monocyte function (ie, HLA-DR expression and tumor necrosis factor-alpha [TNF-alpha] production as markers of immune suppression) in patients with severe AP and to determine whether treatment of blood samples with granulocyte-macrophage colony-stimulating factor (GM-CSF) and/or interferon-gamma (IFN-gamma) corrected the functional defects of monocytes in vitro. METHODS The study consisted of 28 patients with severe AP who were treated at intensive care unit and in whom the proportion of HLA-DR-positive monocytes in the circulation was less than 70%, and 28 matched control subjects who were selected from healthy laboratory personnel. HLA-DR density was determined by whole blood flow cytometry. Monocyte TNF-alpha production in response to bacterial lipopolysaccharides (LPSs) was studied in a whole blood assay. Aliquots of blood were supplemented with IFN-gamma (all 28 patients), GM-CSF (the last 24 patients), or both (the last 12 patients). RESULTS The median proportion of HLA-DR-positive monocytes was 45% in patients (range, 18%-73%) and was 98% in controls (range, 86%-100%; P < 0.001). TNF-alpha levels in response to LPSs were lower in patients (545 pg/mL; range, 84-1990 pg/mL) than in controls (1415 pg/mL; range, 660-5490 pg/mL; P < 0.001). The proportion of HLA-DR-positive cells correlated positively with TNF-alpha levels (r = 0.56; P < 0.01). Both GM-CSF and IFN-gamma increased HLA-DR expression of monocytes in patients (98%; range, 74%-100% for GM-CSF; 99%; range, 86%-100% for IFN-gamma; both P < 0.001). The combination restored monocyte HLA-DR expression (99%; range, 96%-100%; P = 0.002). Compared with basal levels, GM-CSF increased TNF-alpha production of monocytes both in blood samples from patients (median, 1320 pg/mL; range, 35-8015 pg/mL) and controls (median, 3450 pg/mL; range, 1040-9835 pg/mL; both P < 0.001). IFN-gamma increased TNF-alpha production by monocytes in patients (683 pg/mL; range, 186-2705 pg/mL; P < 0.05) but not in controls (1658 pg/mL; range, 765-4755 pg/mL; P = 0.31). With the combination of GM-CSF and IFN-gamma, the TNF-alpha levels of monocytes in patients (3185 pg/mL; range, 545-8280 pg/mL) and in controls (2800 pg/mL; range, 1080-6860 pg/mL) were comparable. CONCLUSIONS The proportion of HLA-DR-positive monocytes correlates with TNF-alpha production, and they both reflect the degree of immune suppression. The low proportion of HLA-DR-positive monocytes in AP can be reversed in vitro by GM-CSF and/or IFN-gamma. The GM-CSF and IFN-gamma treatments also increase LPS-induced TNF-alpha production. By the combination of GM-CSF and IFN-gamma, but not by either agent alone, LPS-induced TNF-alpha production of monocytes was equally high in patients and in controls.

ObjectiveCardiopulmonary bypass elicits systemic inflammation. Depletion of circulating leukocytes might alleviate inflammatory response. We studied the effects of a leukocyte-depleting filter on phagocyte activation during cardiopulmonary bypass.MethodsFifty patients undergoing coronary artery bypass grafting were randomly allocated into an arterial line leukocyte filter group (n = 25) with a Pall LeukoGuard 6 leukocyte-depleting filter (LG6; Pall Biomedical, Portsmouth, United Kingdom) and a control group without any filter (n = 25). Blood sampling took place from arterial line at predetermined time points. In the filter group, the sample was taken immediately before the filter; to evaluate activation at the site, an additional sample was taken immediately after the filter. CD11b/CD18 and L-selectin expressions and basal production of hydrogen peroxide were determined with whole-blood flow cytometry, and plasma lactoferrin level was determined with enzyme-linked immunosorbent assay.ResultsNeutrophil CD11b expression was higher in the filter group than in the control group (P < .001). Likewise, monocyte CD11b expression, neutrophil hydrogen peroxide production, and lactoferrin plasma levels were all significantly higher, whereas neutrophil and monocyte counts and neutrophil L-selectin expression were all significantly lower in the filter group (all P < .001). At 5 minutes of CPB, CD11b expression increased across the filter on neutrophils (median difference 197 relative fluorescence units, range 45–431 relative fluorescence units, P < .001) and monocytes (median difference 26 relative fluorescence units, range −68–111 relative fluorescence units, P < .001).ConclusionThe LG6 arterial line leukocyte filter is ineffective in its principal task of diminishing phagocyte activation during cardiopulmonary bypass.

Late-onset hospital-acquired sepsis is common in extremely low birth-weight (1000 g) (ELBW) infants. The diagnosis is difficult since, at early stages of sepsis, routine laboratory tests are neither specific nor sensitive. In term infants with sepsis neutrophil surface expression of CD11b/CD18, a beta2-integrin, is significantly increased. Here we studied whether increased CD11b/CD18 density on blood neutrophils and monocytes serves as an early sepsis marker in ELBW infants. Blood samples were obtained from 30 ELBW infants on a daily basis for 3-4 postnatal weeks, and neutrophil and monocyte CD11b/CD18 expression was determined by flow-cytometry. Patients were assigned one of 3 groups: 1) an infected group, comprised of infants who had blood culture-positive sepsis and/or necrotizing enterocolitis, 2) a non-infected group, and 3) a potentially infected group, comprised of infants in whom infection was suspected but could not be confirmed microbiologically. One patient had blood culture contamination and was excluded from the analysis. In the infected group, CD11b expression gradually increased during the three days preceding sampling for blood culture. At the day of sampling, median expression of CD11b in neutrophils and monocytes was higher in the infected group than in the control group. For neutrophils the sensitivity and specificity were 1.00 and 0.56, respectively, and for monocytes, 0.86 and 0.94, respectively. From these data, we conclude that determination of CD11b/CD18 density on neutrophils and monocytes may improve diagnosis of late-onset sepsis in ELBW infants.

Immune suppression plays a role in the pathogenesis of acute pancreatitis. The purpose was to describe plasma anti-inflammatory cytokines and blood monocyte human leucocyte antigen (HLA)-DR expression, a cellular marker of immune suppression, in relation to clinical outcome in acute pancreatitis.We studied 74 patients with acute pancreatitis admitted within 72 h after symptom onset; 27 had mild disease and 47 severe disease, of whom 20 developed organ failure. Plasma cytokine concentrations and monocyte HLA-DR density were determined at admission and 1, 2, 3, 7, 14 and 21 days later.The levels of interleukin-1 receptor antagonist, interleukin-6 and interleukin-10 correlated inversely to monocyte HLA-DR expression; each marker correlated with disease severity. Interleukin-4, -11 and -13 levels were low. Organ failure occurred at median 36 h (range 8 to 158) after admission and was predicted at admission by the combination of interleukin-6 and interleukin-10 with sensitivity of 95%, specificity of 88% and positive likelihood ratio of 7.6 (95% confidence interval 3.3 to 17). Patients with secondary infections had a lower proportion of HLA-DR positive monocytes than did controls at day 14 (median: 32% versus 65%; n = 7) and at day 21 (median: 49% versus 83%; n = 6), P0.05 each. In the organ failure group, HLA-DR expression did not differ between survivors and non-survivors.Determining the severity of anti-inflammatory reaction at admission and monitoring the course of immune suppression provide a means for predicting clinical outcome in acute pancreatitis.

Immune suppression plays an important role in the pathogenesis of acute pancreatitis. Monocyte expression of HLA (human leucocyte antigen)-DR, a cellular marker of immune suppression, was determined in relation to the development of organ dysfunction in patients with acute pancreatitis. A total of 310 consecutive patients with acute pancreatitis, admitted to a university hospital within 72 h of pain onset, were studied; 194 (63%) had mild disease (group I), 87 (28%) had severe disease without organ dysfunction (group II), and 29 (9%) had severe disease with organ dysfunction (group III). HLA-DR expression, defined both as the proportion of monocytes that were HLA-DR-positive and as monocyte HLA-DR fluorescence intensity, was determined at admission, using whole-blood flow cytometry. Of the patients in group III, 13 (45%) developed organ dysfunction within 24 h of admission. The proportion of HLA-DR-positive monocytes and monocyte HLA-DR density were both related to the severity of pancreatitis (P

Mast cell tryptase, vascular endothelial growth factor (VEGF), and basic fibroblast growth factor (bFGF) possibly play a role in the pathogenesis of acute pancreatitis (AP). The aim is to describe their serum levels in relation to severity of AP.Seventy patients with AP were studied. Thirty-one had mild acute pancreatitis and 39 severe AP of whom 21 developed organ dysfunction. Serum concentration of tryptase was determined with fluoroimmunoassay (UniCAP), and VEGF and bFGF with ELISA at admission and on days 1, 2, and 7 post-hospitalization.The peak tryptase levels and tryptase levels at 2nd day after symptom onset, although mostly within normal range, were significantly higher in patients with organ dysfunction than in patients without organ dysfunction (6.6 microg/l (inter quartile range 4.8 to 12.6) versus 4.0 microg/l (2.7 to 6.2); P = 0.018 and 6.0 microg/l (4.4 to 7.6) versus 3.4 microg/l (2.3 to 4.8); P = 0.006, respectively). Median serum VEGF and bFGF concentrations increased during follow-up, were significantly higher on day 7 than on days 0, 1, and 2, but were not related to development of organ dysfunction.Mast cell activation, as defined by serum tryptase levels, may play a role in the development of remote organ dysfunction in patients with AP. However, neither tryptase nor the factors VEGF and bFGF serve as predictors of organ dysfunction in clinical AP.

Objective: To study the infectious backround of patients with a history of acute anterior uveitis (AAU) and healthy control subjects. Methods: Sixty four patients with previous AAU and 64 sex and age matched controls were studied. Serum antibodies to Salmonellae , Yersiniae , Klebsiella pneumoniae , Escherichia coli , Proteus mirabilis , Campylobacter jejuni , and Borrelia burgdorferi were measured using enzyme linked immunosorbent assay (ELISA), and antibodies to Chlamydia trachomatis and Chlamydia pneumoniae by microimmunofluorescence test. Peripheral blood mononuclear cells (PBMCs), separated by density gradient centrifugation, were studied for Salmonella and Yersinia antigens by means of an immunofluorescence test, and for C pneumoniae DNA with a polymerase chain reaction (PCR). Results: Neither prevalence nor levels of single microbial antibodies studied differed between the patients and control subjects, or between subgroups of patients created on the basis of clinical characteristics. In logistic regression analysis, the high number of recurrences (>10) of AAU was independently related to the presence of single or multiple bacterial antibodies (p=0.04). None of the PBMC samples of the patients were positive for Yersinia or Salmonella antigens. C pneumoniae PCR was positive in a patient who was negative for C pneumoniae antibodies. Conclusion: Although neither the prevalence nor the levels of single microbial antibodies studied differed between the patients and the controls, current data suggest that the presence of single or multiple antibodies in patients with many recurrences of AAU compared with patients with none or few recurrences may be a sign of repeated infections, antigen persistence, or raised innate immune responsiveness.

Objective. To study neutrophil activation in circulation as a sign of systemic inflammation in preterm infants with respiratory distress syndrome.Methods. The study comprised very low birth weight preterm infants who had respiratory distress syndrome and required intubation and mechanical ventilation (n = 51), 1-day-old preterm infants who had no need for mechanical ventilation (n = 12), term infants (n = 47), and adult volunteers (n = 25). Neutrophil surface expression of CD11b was quantified with flow cytometry.Results. In preterm infants with respiratory distress syndrome, neutrophil CD11b expression during the first day of life was higher than in cord blood (mean: 165 relative fluorescence units [RFU] [standard deviation [SD]: 53], n = 29 vs 83 RFU [SD: 21], n = 11; 95% confidence interval [CI] for difference: 59–106) or in preterm infants without mechanical ventilation (106 RFU [SD: 33], n = 12; 95% CI for difference: 17–90). CD11b expression decreased by age of 10 days. CD11b expression was lower in preterm cord than in term cord blood (95% CI for difference: 5–53). However, in preterm infants with respiratory distress syndrome aged 2 to 5 days, it was higher than in term infants of that age.Conclusions. The observations demonstrate an early transient postnatal neutrophil activation indicative of systemic inflammation that may contribute to the tissue injury in preterm infants with respiratory distress syndrome.

Upon activation, polymorphonuclear leucocytes (PMN) release bactericidal/permeability-increasing protein, (BPI) from their azurophil granules. BPI selectively binds to the lipopolysaccharide (LPS) on gram-negative bacteria and induces their death. This study examined plasma BPI concentration levels in healthy newborns and in newborns with clinical sepsis, and the ability of PMN from preterm and term infants to release BPI. We also studied the release of myeloperoxidase (MPO), and the surface expression of adhesion molecule CD11b on PMN. In infants with clinical sepsis, plasma BPI concentration was higher, 27.8 microg/L [8.6-883; median (range)] (n = 11), than in healthy term infants 8.9 microg/L (3.9-179) (n = 17), and in adults 7.3 microg/L (0.7 -18.4) (n = 15); p = 0.014, Kruskal-Wallis. In preterm infants (n = 8), the ability of PMN to release BPI in vitro after stimulation with PMA was 8.8, in term infants it was 15.9 (n = 29; p > 0.05 vs. preterm infants) and in adults 23.4 ng/10(6) PMN (n = 15; p = 0.024 and p > 0.05 vs. preterm and term infants, respectively). The corresponding values of MPO were 20.0 ng/10(6) (11.3-46.7) in preterms, 19.0 ng/10(6) (2.2-223.7) in terms, and 27.8 ng/10(6) (9.1-80.7) in adults; p = 0.67 between groups. In infants with clinical sepsis, CD11b level was higher, 292 RFU (234-403) than the basal CD11b expression levels in healthy newborn infants, 116 RFU (76-145); P = 0.0001. FMLP-stimulated PMN CD11b expressions in preterm cord blood, 1071 RFU (552-1286) and in term cord blood, 918 (567-1472) were on the same level, but lower than that in adult blood, 1592 (973-1946); p < 0.001, ANOVA. Our findings suggest that in preterm infants the ability to release BPI is lower than in adults and term infants. These findings suggest that premature neonates have an impaired ability to mobilize BPI, possibly contributing to their marked susceptibility to infections with Gram-negative bacteria.

Acute pancreatitis (AP) is a common disease with wide variation of severity. The diagnosis of AP is usually based on high serum amylase or lipase values but the accuracy of these methods is considered unsatisfactory. One in five of the patients develops a severe disease and carries a considerable risk of development of organ failure and high mortality. Early detection of patients with severe AP and especially those with increased risk of organ failure is importance since such patients seem to benefit from treatment in an intensive care unit started as soon as possible after presentation. In addition to enzymological methods, increasing interest has been focused on laboratory markers reflecting the level of inflammatory response in AP. At present, in routine clinical work the most commonly used severity marker is serum C-reactive protein, the concentration of which rises too slowly to be used for early prediction of severity. New therapies aiming at modifying the course of systemic inflammation in AP are being developed and therefore monitoring the patient's immune inflammatory status is needed. In this review article we present the current knowledge of laboratory tests, which has been evaluated for diagnostic and prognostic purposes in AP.

Systemic inflammation triggered by insults like sepsis and acute pancreatitis may play a role in development of indirect acute lung injury (ALI) and acute respiratory distress syndrome (ARDS). Because little is known about the course of systemic inflammation on the days preceding diagnosis of ARDS, we prospectively monitored immune inflammatory status in 52 patients at risk and we assessed the presence of ALI and ARDS on day 7 after admission to the intensive care unit. On admission, serum interleukin (IL) 8, IL-6, and soluble IL-2 receptor concentrations were significantly higher in patients with subsequent ALI (n = 18) than in patients without ALI (n = 30). During a 4-day follow-up, IL-8 and IL-6 levels of ALI patients remained high and those of non-ALI patients decreased. None of the markers discriminated ARDS patients (n = 9) from non-ARDS ALI patients (n = 9). Among 11 patients with acute pancreatitis, ALI patients had significantly higher IL-8, IL-6, and phagocyte CD11b expression levels than did non-ALI patients, whereas among 14 patients with massive transfusion, respective findings in ALI and non-ALI patients were comparable. Results give credence to the view that systemic inflammation plays a role in development of ALI triggered by pancreatitis, but not in that by massive transfusion. This finding, if confirmed in studies with sufficient statistical power, suggests that the patients with massive transfusion do not necessarily benefit from novel biotherapies aimed at altering the course of systemic inflammation.

Background: Soluble E-selectin (sE-selectin) is a marker of activation of vascular endothelium. Objective: To examine serum levels of sE-selectin in a cohort of 85 patients with early rheumatoid arthritis (RA) followed up for five years. Methods: sE-selectin levels were assessed annually using an enzyme linked immunosorbent assay (ELISA) and related to simultaneously obtained clinical and laboratory measures. Joint inflammation was evaluated by active joint count, functional status by Health Assessment Questionnaire (HAQ), and radiographic findings in hands and feet by the Larsen method. Laboratory tests included serum C reactive protein (CRP) level, erythrocyte sedimentation rate, blood haemoglobin level, white blood cell count (WBC), and platelet count. Area under the curve (AUC) was calculated for each variable, and Jonckheere's test for ordered alternatives was applied to assess significance of association between sE-selectin AUC tertiles and other variables. Baseline sE-selectin tertiles were related to change in Larsen score and HAQ score at five years. Odds ratios (OR) with 95% confidence interval (CI) were calculated using univariate and multivariate logistic regression. Results: sE-selectin levels were associated with CRP level (p=0.012), WBC (p=0.037), active joint count (p=0.019), progression of joint destruction (p=0.038), and HAQ score at five years (p=0.021), but not with extra-articular symptoms or comorbidity. Baseline sE-selectin levels in the third tertile predicted the HAQ score at five years (OR 4.18, 95% CI 1.15 to 15.22). sE-selectin levels of patients did not differ significantly from those of healthy control subjects. Conclusion: The degree of activation of vascular endothelium is associated with activity and outcome of early RA.

Objective. To examine CD14 and TNF &#102 gene polymorphisms in early arthritis in relation to clinical outcome. Methods. We studied 141 Caucasians who had had early arthritis 10 to 38 years earlier. We analysed CD14 (-159) and TNF &#102 (- 238, - 308, - 376) polymorphisms using a novel cycle minisequencing method. DNA pools from 370 Caucasian blood donors served as controls. Results. CD14 (-159)C &#77 T allele frequencies were comparable among patients and controls (39% vs 40%). Fifty men and 42 women had recovered while 24 men and six women had chronic spondyloarthropathy (SpA). Mutant T allele frequency was higher in the chronic SpA group than in the recovered group in women (75% vs 32%, relative risk 1.3, 95% confidence limit 1.1 to 1.6, P= 0.011), but not in men (38% vs 44%). All female patients with chronic SpA had CD14 (- 159)T allele and none had a possibly protective TNF &#102 (- 308)G &#77 A allele. Conclusion. Possession of CD14 (- 159)T allele does not increase risk of ReA but may increase susc...

To describe the incidence and nature of bloodstream infections (BSI) among children with juvenile idiopathic arthritis (JIA) followed-up prospectively from disease onset.The Social Insurance Institution's (SII) national register on individuals with reimbursement for medication of chronic diseases was used to identify children with JIA from 2004 through 2011 and their medications. The National Infectious Disease Register (NIDR) collects data of all blood culture positive samples from all microbiology laboratories in Finland. We combined the NIDR and SII registers to identify JIA patients with BSI. Clinical and laboratory data of each JIA-BSI patient were collected from hospital records.There were 1604 JIA patients and 6630 person-years of follow-up. Five patients had BSI. During the first 5 years after diagnosis the cumulative emergence of BSI was 0.38% [95% confidence interval (CI) 0.16% to 0.92%]. The incidence rates were 7.5/10 000 follow-up years for JIA (95% CI 2.4-17.6) and 2.8/10 000 follow-up years for the age-matched general population (95% CI 2.7-2.9). The standardised incidence ratio was 3.0 (95% CI 1.2 to 7.2). The causative bacteria were Streptococcus pneumoniae, Staphylococcus aureus, Escherichia coli and Fusobacterium necrophorum. Three patients were on anti-rheumatic drugs, including two on TNF inhibitors. All patients responded rapidly to antimicrobial therapy and recovered uneventfully.Although BSI is rare among children with JIA, the incidence is 3-fold higher than among the general population.

Despite convincing results of studies in vitro, less is known about the effects of antioxidants on in vivo redox balance in humans. We developed a novel parameter of in vivo redox balance, and studied it and its relation to dental infections in 51 patients on medication for coronary heart disease (CHD) and 39 random controls matched for age group, sex, social class and locality. In vivo redox balance was the ratio of plasma antioxidant capacity, as measured with radical-trapping assay, to neutrophil respiratory burst capacity, as measured with whole blood chemiluminescence assay. Dental infections were quantitated with four rating scales. CHD patients had higher values than controls. Patients on acetosalicylic acid (ASA), diuretics or beta blockers, but not the ones on calcium channel blocker, had significantly higher redox balance than non-users. Combination of calcium channel blockers and ASA was associated with redox balance similar to taking beta blockers or diuretics. Diuretics and ASA were independent determinants of redox balance in multivariate analyses. Redox balance did not correlate with severity of dental infections (Spearman's r 0.06 to 0.11). The results contrast experimental data indicating that calcium channel blockers are as antioxidants superior to other cardiovascular drugs. Total antioxidant capacity in parallel with oxygen species production capacity should be considered in attempts to solve the antioxidant paradox.

Background Neutrophil subpopulations with enhanced oxidative reactivity have been described in a number of clinical and in vitro settings. In the dichlorofluorescin (DCFH) oxidation assay, it is essential to maintain cellular viability and plasma membrane integrity through all stages of sample preparation. The process of erythrocyte lysing is crucial because a number of commercial lysing reagents can increase leukocyte membrane permeability. Methods We assessed viability [propidium iodide (PI) method], DCFH oxidation, and CD11b expression of resting or in vitro–stimulated neutrophils exposed to six different red cell lysing procedures. Results Formaldehyde-containing reagents (Optilyse B, FACS Lyse, and Erythrolyse) but not hypotonic shock or ammonium chloride (NH4Cl) solutions rendered 91.4–99.8% of resting neutrophils PI positive, with concomitant reductions in dichlorofluorescein (DCF) fluorescence, suggesting efflux of the fluorochrome. However, when stimulated with N-formyl-methionyl-leucyl-phenylalanine or Yersinia enterocolitica and then treated with FACS Lyse or Erythrolyse, up to 69.9% of neutrophils remained PI negative and exhibited enhanced DCF fluorescence. CD11b expression of PI-positive and -negative neutrophils was comparable, suggesting that they were activated equally. Conclusions FACS Lyse and Erythrolyse can modify neutrophil plasma membrane integrity, whereas hypotonic shock and NH4Cl solutions retain cellular viability and are lysing methods of choice in evaluation of neutrophil respiratory burst by DCFH oxidation assay. Cytometry 43:290–296, 2001. © 2001 Wiley-Liss, Inc.

Objective: To investigate whether marker(s) of systemic inflammation detect, at an early stage of acute pancreatitis, patients who may ultimately develop severe disease. Design: Prospective study. Setting: University hospital emergency unit. Patients: Thirty patients with mild acute pancreatitis (SEV 0 group) and 27 with severe acute pancreatitis. Of the latter, 11 did not develop organ failure (SEV 1 group), whereas the other 16 patients developed acute respiratory failure and 9 of them also developed renal failure (SEV 2 group). Interventions: Blood samples were collected at admission to the hospital (T 0 ), and at 12 hrs (T 12 ) and 24 hrs (T 24 ) after admission. Measurements and Main Results: The plasma concentrations of procalcitonin (PCT), soluble E-selectin (sE-selectin), soluble interleukin-2 receptor (slL-2R), and the serum concentration of C-reactive protein (CRP) were monitored. PCT levels at To were significantly higher in the SEV 1 group (median 0.4 nglmL, range 0.2-2.3) and the SEV 2 group (0.8 ng/mL, 0.2-73.5) than in the SEV 0 group (0.3 ng/mL, 0.1-3, p

A DDS type circuit structure for producing numerically programmable square wave clock signal is presented. The high speed D/A converter needed in conventional DDS systems is replaced by an N tap delay line time domain interpolator that effectively increases the sampling rate by a factor of N. Thus the circuit can be used up to full clock rate without image filtering. A prototype IC with clock frequency of 30 MHz, 5 bit interpolator and SFDR of −40 dBc up to 10 MHz and −33 dBc up to 15 MHz has been designed and tested.

BACKGROUND During experimental liver transplantation, neutrophil sequestration results in increased oxygen free radical production and correlates inversely with graft viability. Neutrophil activation in clinical liver transplantation is poorly understood. METHODS We assessed leukocyte sequestration and transhepatic differences of neutrophil and monocyte CD11b expression, neutrophil free radical production, and plasma concentrations of interleukin 6 and interleukin 8 in nine patients during liver transplantation. RESULTS Significant hepatic neutrophil sequestration occurred during initial graft rewarming with portal blood, after inferior vena cava declamping, and after hepatic artery declamping (all P

To obtain predictors of organ failure (OF), we studied markers of systemic inflammation [circulating levels of interleukin-6 (IL-6), IL-8, soluble IL-2 receptor (sIL-2R), soluble E-selectin and C-reactive protein, and neutrophil and monocyte CD11b expression] and routine blood cell counts in 20 patients with systemic inflammatory response syndrome and positive blood culture. Eight patients with shock due to community-acquired infection developed OF, whereas 11 normotensive patients and one patient with shock did not (NOF group). The first blood sample was collected within 48 h after taking the blood culture (T1). OF patients, as compared with NOF patients, had at T1 a lower monocyte count, a lower platelet count, higher levels of CD11b expression on both neutrophils and monocytes, and higher concentrations of IL-6, IL-8 and sIL-2R. C-reactive protein and soluble E-selectin concentrations did not differ between groups. No parameter alone identified all patients that subsequently developed OF. However, a sepsis-related inflammation severity score (SISS), developed on the basis of the presence or absence of shock and on the levels of markers at T1, identified each patient that developed OF. The maximum SISS value was 7. The range of SISS values in OF patients was 2–5, and that in NOF patients was 0–1. In conclusion, high levels of CD11b expression, depressed platelet and monocyte counts, and high concentrations of IL-6, IL-8 and sIL-2R predict OF in patients with community-acquired septic shock, and the combination of these markers may provide the means to identify sepsis patients who will develop OF.

Background To study the effect of nitecapone, a novel antioxidant, on cardiac neutrophil activation during cardiopulmonary bypass in patients. Methods In a double-blind, placebo controlled trial, 30 male patients undergoing coronary artery bypass grafting were randomly assigned to control (crystalloid cardioplegia, n = 15) and nitecapone groups (cardioplegia supplemented with nitecapone, n = 15). Leukocyte differential counts, neutrophil and monocyte CD11b and L-selectin expressions and neutrophil hydrogen peroxide production were measured in blood samples parallelly obtained from the coronary sinus and aorta before cardiopulmonary bypass and at 1, 5, and 10 min after aortic declamping. Myocardial myeloperoxidase activity was analyzed in biopsies taken at 1, 5, and 10 min after declamping. Results Transcoronary neutrophil difference (i.e., aorta--sinus coronarius) at 1 min after aortic declamping was significantly lower in nitecapone-treated patients (0.41 [-0.42-0.98] x 10(9) cells/l) than in controls (0.68 [-0.28-2.47] x 10(9) cells/l; P = 0.032). At 5 min after aortic declamping, significant transcoronary reduction of neutrophil hydrogen peroxide production and CD11b expression were observed in controls but not in nitecapone patients. At 24 h postoperatively, left ventricular stroke volume was better in nitecapone-treated patients (94 [51-118] ml) than controls (66 [40-104] ml; P= 0.018). Data are median [range]. Conclusion Nitecapone added to cardioplegia solution reduces cardiac neutrophil accumulation and transcoronary neutrophil activation during clinical cardiopulmonary bypass. Reflected by better left ventricular stroke volume, nitecapone treatment may be an additional way of reducing the deleterious effects of neutrophil activation during cardiopulmonary bypass.

Criteria of the systemic inflammatory response syndrome (SIRS) are known to include patients without systemic inflammation. Our aim was to explore additional markers of inflammation that would distinguish SIRS patients with systemic inflammation from patients without inflammation. The study included 100 acutely ill patients with SIRS. Peripheral blood neutrophil and monocyte CD11b expression, serum interleukin-6, interleukin-1β, tumour necrosis factor-α and C-reactive protein were determined, and severity of inflammation was evaluated by systemic inflammation composite score based on CD11b expression, C-reactive protein and cytokine levels. Levels of CD11b expression, C-reactive protein and interleukin-6 were higher in sepsis patients than in SIRS patients who met two criteria (SIRS2 group) or three criteria of SIRS (SIRS3 group). The systemic inflammation composite score of SIRS2 patients (median 1.5; range 0–8, n = 56) was lower than that of SIRS3 patients (3.5; range 0–9, n = 14, P = 0.013) and that of sepsis patients (5.0; range 3–10, n = 19, P< 0.001). The systemic inflammation composite score was 0 in 13/94 patients. In 81 patients in whom systemic inflammation composite scores exceeded 1, interleukin-6 was increased in 64 (79.0%), C-reactive protein in 59 (72.8%) and CD11b in 50 (61.7%). None of these markers, when used alone, identified all patients but at least one marker was positive in each patient. Quantifying phagocyte CD11b expression and serum interleukin-6 and C-reactive protein concurrently provides a means to discriminate SIRS patients with systemic inflammation from patients without systemic inflammation.

The aim of this study was to determine whether the +896 A→G substitution of the Toll-like receptor 4 ( TLR4 ) gene, causing the Asp299→Gly change in the extracellular domain of TLR4, influences treatment response in recent-onset rheumatoid arthritis. 169 patients with rheumatoid arthritis were genotyped from the Finnish Rheumatoid Arthritis Combination Therapy trial, in which they were treated either with only one disease-modifying antirheumatic drug (DMARD) with or without prednisolone (single group), or with three DMARDs and prednisolone (combination group). Patients homozygotic for the wild-type +896A allele were compared with those having the polymorphic G allele in terms of early clinical response (at 6 months) by the 28-joint Disease Activity Score (DAS28). 1 of 20 (5%; (95% (confidence interval (CI) 1 to 5)) patients of the single group with TLR4 +896AG or GG and 29 of 67 (43%; (95% CI 31 to 56)) patients with AA were in remission (p = 0.001). DAS28 of the single group with TLR4 +896AG or GG was higher than with AA (p = 0.019). In the combination group, remission rates and DAS28 values were comparable between the genotypes. The polymorphic TLR4 +896G allele may impair treatment response to single DMARD treatment in recent-onset rheumatoid arthritis.

Peripheral blood mononuclear cells of Crohn's disease (CD) patients with the common 1007fs mutation of the caspase recruitment domain-containing 15/nucleotide-binding oligomerization domain-containing 2 (CARD15/NOD2) gene show impaired nuclear factor kappa B (NF-κB) activation in response to muramyl dipeptide (MDP), as determined by Western blotting. We applied phospho-specific flow cytometry to examine NF-κB and p38 activation in whole blood monocytes of 16 CD patients with or without the 1007fs and previously described rare mutations of the CARD15 gene, and healthy reference subjects. Aliquots of whole blood were supplemented with MDP (0-1000 ng/mL), incubated for 10-40 min and processed for flow cytometry. Bacterial lipopolysaccharide (LPS) was used as a positive control agonist. We found that NF-κB and p38 phosphorylation induced by MDP was not detectable in monocytes of patients homozygous for the CARD15 1007fs mutation, while those induced by LPS were normal. We also determined MDP-induced NF-κB phosphorylation levels in nuclear extracts of mononuclear cells separated from blood using enzyme-linked immunosorbent assay (ELISA), and observed that the levels decreased in a 1007fs mutation-dose dependent manner. We conclude that phospho-specific whole blood flow cytometry provides a means to study phosphorylation of NF-κB and p38 in clinical samples and can be applied to screening of CD patients homozygous for the CARD15 1007fs mutation.

The pathogenesis of reactive arthritis (ReA) is incompletely understood but may involve aberration(s) in the host's innate immune response towards infecting microbes. We therefore studied the production of interleukin (IL)-1β, a marker of inflammasome activation, and of IL-6, IL-12, IL-23, and tumour necrosis factor (TNF)-α, promoters of T-cell differentiation, by peripheral blood mononuclear cells (PBMNs) and monocyte-derived macrophages from healthy subjects with a history of ReA.The study included 10 human leucocyte antigen (HLA)-B27-positive healthy subjects with previous ReA triggered by Yersinia enterocolitica O:3 infection and 20 healthy reference subjects, of whom 10 were HLA-B27 positive. PBMNs and macrophages were cultured for 18 h with bacterial lipopolysaccharide (LPS), muramyl dipeptide (MDP), Yersinia, or their appropriate combinations. PBMNs were also stimulated with monosodium urate (MSU) crystals. Cytokine levels were measured using an enzyme-linked immunosorbent assay (ELISA) and the Luminex system.IL-1β secretion was similar from cells of the ReA group and from the HLA-B27-positive and -negative reference groups. TNF-α production from macrophages upon co-stimulation of LPS and MDP increased in the order ReA groupHLA-B27-positive reference groupHLA-B27-negative reference group (p for a trend = 0.027). Similarly, Yersinia-induced TNF-α and IL-23 production increased in the same order (p for trend for TNF-α = 0.036; p for trend for IL-23 = 0.026).PBMNs and macrophages from healthy subjects with previous ReA show normal inflammasome activation and low TNF-α and IL-23 production. This low cytokine production may impair bacterial elimination and thereby contribute to the triggering of ReA.

Immaturity of the immune system renders newborns susceptible to infections. We searched for aberrations in leucocyte signalling profiles, using phospho-specific whole-blood flow cytometry, in cord blood of nine preterm (two born before 32nd gestational week) and nine full-term infants, born by caesarean section. Thirteen adults served as reference subjects. Monocyte NF-κB phosphorylation following tumour necrosis factor (TNF) or bacterial stimulation was higher in preterm neonates than in full-term neonates or adults, p38 phosphorylation following bacterial stimulation was higher in both preterm and full-term neonates than in adults, while STAT1 phosphorylation by IFN-γ or IL-6, STAT3 phosphorylation by IL-6 and STAT5 phosphorylation by GM-CSF were lower in both full-term and preterm neonates than in adults. Neutrophil STAT1 and STAT3 phosphorylation following IFN-γ stimulation and STAT5 phosphorylation following GM-CSF stimulation were lower in newborn neonates than in adults. In both CD3(+) CD4(+) and CD3(+) CD8(+) lymphocytes, NF-κB phosphorylation by TNF was higher and STAT5 phosphorylation by IL-2 was lower in preterm and full-term newborns than in adults. STAT6 phosphorylation by IL-4 was comparable in monocytes and lymphocytes of newborns and adults. The results suggest that innate immune signalling pathways responding to inflammatory stimuli are strongly functional in leucocytes of preterm neonates, which may render these neonates susceptible to inappropriate tissue injury. In leucocytes of both preterm and full-term newborns, responses needed against intracellular pathogens, and regulatory functions show immaturities, possibly contributing to worse control of infections.