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Ethnopharmacological Relevance: Zanthoxylum piperitum has been used as a traditional Asian medicine to treat hypertension, stroke, bruise and muscle pain. It has been known to induce detoxification; affect anti-bacterial, anti-oxidant, and tyrosinase activity; inhibit osteosarcoma proliferation; anti-osteoarthritis inflammation. In this study, we aim to identify the therapeutic effect of Z. piperitum 90% EtOH extract (ZPE-LR) on rheumatoid arthritis., Material and Methods: We investigated the anti-rheumatoid arthritis and -immunomodulatory activities of the ZPE-LR in collagen-induced arthritic (CIA) mice, a rheumatoid arthritis animal model. In order to assess the analgesic effects of ZPE-LR in vivo, acetic acid injection, formaldehyde injection, hot plate model was used. The mechanism for anti-inflammatory activity of ZPE-LR was identified with LPS-stimulated Raw 264.7 cells., Results: Pharmacologically, oral administration of ZPE-LR into CIA mice resulted in a significant and dose-dependent decrease in clinical arthritis score and paw swelling compared to untreated negative control. Pathologic examination showed that ZPE-LR prevented morphological change in cartilage and destruction of phalanges in CIA mice. This protective effect was associated with reduced pain, inflammatory mediators such as NO, TNF-α, IL-1β, and IL-6, as well as COX-2 and iNOS expression. Furthermore, reduction of phosphor-ERK and BDNF indicates a novel rheumatoid arthritis-regulating mechanism by ZPE-LR treatment., Conclusions: These data suggest that the administration of ZPE-LR remarkably inhibited CIA progression and might be helpful in suppressing inflammation and pain in rheumatoid arthritis., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Scorpionism is responsible for most accidents involving venomous animals in Brazil, which leads to severe symptoms that can evolve to death. Scorpion venoms consist of complexes cocktails, including peptides, proteins, and non-protein compounds, making separation and purification procedures extremely difficult and time-consuming. Scorpion toxins target different biological systems and can be used in basic science, for clinical, and biotechnological applications. This study is the first to explore the venom content of the unexplored scorpion species Rhopalurus crassicauda , which inhabits exclusively the northernmost state of Brazil, named Roraima, and southern region of Guyana. Here, we pioneer the fractionation of the R. crassicauda venom and isolated and characterized a novel scorpion beta-neurotoxin, designated Rc1, and a monomeric hyaluronidase. R. crassicauda venom and Rc1 (6,882 Da) demonstrated pro-inflammatory activities in vitro and a nociceptive response in vivo . Moreover, Rc1 toxin showed specificity for activating Na v 1.4, Na v 1.6, and BgNa v 1 voltage-gated ion channels. This study also represents a new perspective for the treatment of envenomings in Roraima, since the Brazilian scorpion and arachnid antivenoms were not able to recognize R. crassicauda venom and its fractions (with exception of hyaluronidase). Our work provides useful insights for the first understanding of the painful sting and pro-inflammatory effects associated with R. crassicauda envenomings., (Copyright © 2020 Abreu, Bordon, Cerni, Oliveira, Balenzuela, Alexandre-Silva, Zoccal, Reis, Wiezel, Peigneur, Pinheiro-Júnior, Tytgat, Cunha, Quinton, Faccioli, Arantes, Zottich and Pucca.)

This study aims to investigate active phytochemicals isolated from Pyrola incarnata Fisch. (P. incarnata) and their protection against neuroinflammation induced by LPS. Betulin, accompanied with other 9 compounds, were isolated from P. incarnata and elucidated by spectroscopic analysis ( 1 H-, 13 C NMR). ELISA kits and the measurement of NO production based on Griess reaction showed that betulin (5) (250 μg/mL) could suppress LPS-induced activation of microglial cell BV-2 better than others by inhibiting inflammatory cytokines (TNF-α, IL-6, IL-1β) expression and NO production. With the guidance of computer-aided drug design and the analysis of biological experiment, we demonstrated betulin could reduce LPS-induced iNOS expression, prevent JNKs pathways, and down-regulate the phosphorylation levels of NF-κB/p65. In conclusion, betulin isolated from P. incarnata possessed outstanding anti-neuroinflammation potential, presumably related to iNOS expression, JNKs and NF-κB/p65 pathways. Therefore, Pyrola incarnata may be a valuable natural resource and betulin is a potential drug for the treatment of neurodegenerative disorders by inhibiting inflammatory mediators., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Objectives: Feline infectious peritonitis (FIP) is a high mortality infectious disease. Single nucleotide polymorphisms (SNPs) in the genes encoding interferon gamma ( IFNG ), tumour necrosis factor alpha ( TNFA ) and dendritic cell-specific intercellular adhesion molecule-grabbing non-integrin (DC-SIGN; CD209 ) have been associated with increased and decreased risk of developing FIP. This study was designed to determine whether these associations were present in a UK population of pedigree cats using samples from cats euthanased with a confirmed diagnosis (FIP, n = 22; non-FIP, n = 10) or clinically healthy cats over 11 years of age (n = 3)., Methods: DNA was extracted from tissue (n = 32) or blood (n = 3) and PCR performed for regions of IFNG, TNFA and CD209 . PCR amplicons were sequenced, each SNP genotype was determined, and genotype/allele frequency for each SNP and FIP status were compared., Results: No significant association was found between the genotype and FIP status for any SNP analysed. There was a trend for the heterozygous CT genotype at both IFNG g.401 and IFNG g.408 to be associated with FIP ( P  = 0.13), but this genotype was also found in a substantial proportion of non-FIP cats. There was also a trend for the heterozygous CT genotype at IFNG g.428 to be associated with FIP ( P  = 0.06), although most cats with FIP had the CC genotype at this locus. No associations were found between any allele at TNFA g.-421, CD209 g.1900, CD209 g.2276, CD209 g.2392 and CD209 g.2713 and FIP., Conclusions and Relevance: The use of the IFNG, TNFA and CD209 SNPs described to predict the risk of FIP cannot currently be recommended.

Two 11,20-epoxybriaranes, including a known compound, juncenolide K ( 1 ), as well as a new metabolite, fragilide X ( 2 ), have been isolated from gorgonian Junceella fragilis collected off the waters of Taiwan. The absolute configuration of juncenolide K ( 1 ) was determined by single-crystal X-ray diffraction analysis for the first time in this study and the structure, including the absolute configuration of briarane 2 was established on the basis of spectroscopic analysis and compared with that of model compound 1 . One aspect of the stereochemistry of the known compound 1 was revised. Briarane 2 was found to enhance the generation of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) release from RAW 264.7 cells.

Introduction : Although the etiology of inflammatory bowel diseases (IBD) remains unknown, accumulating evidence suggests that the intestinal tissue damage in these disorders is due to a dynamic interplay between immune cells and non-immune cells, which is mediated by cytokines produced within the inflammatory microenvironment. Areas covered : We review the available data about the role of inflammatory cytokines in IBD pathophysiology and provide an overview of the therapeutic options to block the function of such molecules. Expert opinion : Genome studies, in vitro experiments with patients' samples and animal models of colitis, have largely advanced our understanding of how cytokines modulate the ongoing mucosal inflammation in IBD. However, not all the cytokines produced within the damaged gut seem to play a major role in the amplification and perpetuation of the IBD-associated inflammatory cascade. Indeed, while some of the anti-cytokine compounds are effective in some subgroups of IBD patients, others have no benefit. In this complex scenario, a major unmet need is the identification of biomarkers that can predict response to therapy and facilitate a personalized therapeutic approach, which maximizes the benefits and limits the adverse events.

Background: Acute pancreatitis (AP) usually has a mild course with a mortality rate below 1%. However, around 10% of patients develop severe AP (SAP) involving extra-pancreatic tissues and other organ systems. The mortality of SAP is around 42%. The outcome of SAP is closely related to the development of systemic inflammation and consecutive organ failures. Most current therapies including fluid resuscitation, antimicrobial therapy, drainage procedures, and endoscopic management of complications are symptomatic rather than causative approaches, except sphincterotomy for gallstone pancreatitis. Regarding the high mortality of SAP and its close association with systemic inflammation, extracorporeal removal of inflammatory mediators is an appealing approach. Several recent studies have demonstrated that the CytoSorb adsorber effectively eliminates inflammatory cytokines, such as IL-1ß, IL-6, IL-8, IL-10, and TNF-alpha. Some of these trials suggested that therapy with CytoSorb might improve outcome, including a reduction in the vasopressor dosage and reversal of shock.Therefore, it is the objective of this study to evaluate the effectiveness of 2 consecutive 24 h-treatments with CytoSorb on hemodynamics in patients with early SAP., Methods: This study includes patients with early SAP (APACHE-II ≥10) and transpulmonary thermodilution hemodynamic monitoring (PiCCO; EV-1000) within a maximum of seven days from the onset of pain. Eligible patients will be treated with 2 consecutive periods of CytoSorb. A 20%-improvement in the vasopressor dependency index (VDI) - which relates is derived from mean arterial pressure (MAP) and catecholamine dosage - is the primary outcome. In addition to this clinical outcome, there are several laboratory (cytokine levels) and translational endpoints (including multiplex-ELISAs of numerous anti- and pro-inflammatory cytokines/chemokines and DNA analyses). Primary outcome analysis will compare the incidence of the primary endpoint in 30 patients from the intervention group to 60 matched controls with advanced hemodynamic monitoring recruited from recent studies in SAP within the same setting and the same centers., Discussion: A potential improvement in hemodynamics and/or other outcomes by CytoSorb would provide a new therapeutic option in the early treatment of SAP with a pathophysiological rationale., Trial Registration: This study was registered on March 17, 2017 (ClinicalTrials.gov Identifier: NCT03082469). URL: https://clinicaltrials.gov/ct2/show/NCT03082469., Version: V_PACIFIC_1.0 September 30, 2018.

Acetobacter pasteurianus is an acetic acid-producing Gram-negative bacterium commonly found associated with plants and plant products and widely used in the production of fermented foods, such as kefir and vinegar. Due to the acid conditions of the bacterium living habitat, uncommon structural features composing its cell envelope are expected. In the present work we have investigated the A. pasteurianus CIP103108 lipopolysaccharide (LPS) structure and immunoactivity. The structure of the lipid A and of two different O-polysaccharides was assessed. Furthermore, immunological studies with human cells showed a low immunostimulant activity of the isolated LPS, in addition to a slight capability to lower the NF-kB activation upon stimulation by toxic LPS., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Septic cardiomyopathy represents cardiac impairment in sepsis and is a part of systemic involvement in sepsis. Cytokine storm is responsible for septic shock and for myocardial dysfunction of potentially reversible septic cardiomyopathy. Several case reports and case series demonstrated successful removal of circulating cytokines by combined blood purification techniques. In this way, septic shock and survival of septic patients improved. However, the evidences for reversal of myocardial dysfunction are rare., Case Presentation: We present a patient with a history of chemotherapy for coat cell lymphoma, splenectomy and autologous bone marrow transplantation, who suffered severe pneumococcal sepsis, septic shock and septic cardiomyopathy, resistant to pharmacological therapy. Combined blood purification techniques 36 h after the start of treatment successfully decreased Interleukin-6 level, lactacidosis, the need for vasopressors to maintain normotension, improved systolic function of the left ventricle and clinical outcome., Conclusions: Our case suggests that combined blood purification techniques initiated even 36 h after the start of treatment successfully removed inflammatory cytokines, reversed circulatory failure and improved left ventricular systolic function in pneumococcal sepsis.

Lonicera caerulea L. berry polyphenols (LCBP) are considered as major components for bioactivity. This study aimed to clarify the molecular mechanisms by monitoring inflammatory and antioxidant mediator actions in lipopolysaccharide (LPS)-induced mouse paw edema and macrophage cell model. LCBP significantly attenuated LPS-induced paw edema (3.0 ± 0.1 to 2.8 ± 0.1 mm, P < 0.05) and reduced (P < 0.05) serum levels of monocyte chemotactic protein-1 (MCP-1, 100.9 ± 2.3 to 58.3 ± 14.5 ng/mL), interleukin (IL)-10 (1596.1 ± 424.3 to 709.7 ± 65.7 pg/mL), macrophage inflammatory protein (MIP)-1α (1761.9 ± 208.3 to 1369.1 ± 56.4 pg/mL), IL-6 (1262.8 ± 71.7 to 499.0 ± 67.1 pg/mL), IL-4 (93.3 ± 25.7 to 50.7 ± 12.5 pg/mL), IL-12(p-70) (580.4 ± 132.0 to 315.2 ± 35.1 pg/mL), and tumor necrosis factor-α (TNF-α, 2045.5 ± 264.9 to 1270.7 ± 158.6 pg/mL). Cell signaling analysis revealed that LCBP inhibited transforming growth factor β activated kinase-1 (TAK1)-mediated mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) pathways, and enhanced the expression of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) and manganese-dependent superoxide dismutase (MnSOD) in earlier response. Moreover, cyanidin 3-glucoside (C3G) and (-)-epicatechin (EC), two major components of LCBP, directly bound to TAK1. These data demonstrated that LCBP might inhibit LPS-induced inflammation by modulating both inflammatory and antioxidant mediators.

Atherosclerosis is an important predictor of mortality in patients with chronic kidney disease (CKD) and is associated with a wide inflammatory response. The aim of this study is to evaluate in vitro how different membranes can remove mediators associated with this pathology in a closed loop dialysis model. We performed experimental hemofiltration in vitro using three different membrane materials. Human plasma was preliminarily incubated with various inflammatory mediators and filtered in a closed loop circulation model for 240 min. Respective concentrations of 17 different mediators were measured over time to study the removal mechanisms of each membrane, including associated removal time course. The experiment was repeated three times for the assay of tumor necrosis factor (TNF)-α to document the model variability. Means were compared using Mann-Whitney test. Most of the investigated mediators were effectively removed with the different dialysis membranes. Adsorption mechanism was mainly at the origin of the decrease in mediators circulating concentrations and was maximized in the region 10 000-20 000 Da. Especially, the HeprAN membrane showed fast removal capacities of mediators with elevated isoelectric point including complement factors and chemokines or having basic groups located in the protein periphery, plasminogen activator inhibitor (PAI-1), and TNF-α-like. The latter was further significantly removed with HeprAN and polymethylmethacrylate (PMMA) compared to polyethersulfone (PES) material (P < 0.01). We concluded that dialysis using ionic adsorptive membrane could have a beneficial impact for CKD patients with atherosclerosis and would deserve further clinical investigations., (© 2016 International Center for Artificial Organs and Transplantation and Wiley Periodicals, Inc.)

It has been speculated that extracorporeal blood purification therapies might improve the clinical outcome for patients with severe sepsis, with or without acute kidney injury, since the removal of inflammatory mediators and/or bacterial toxins from circulation could modulate the inflammatory responses that result in organ damage. Despite initial enthusiasm based on promising preliminary results, subsequent investigations did not show sustainable survival benefit. We review the principles and development of blood purification techniques for sepsis and septic acute kidney injury.

The venom of Micrurus lemniscatus, a coral snake of wide geographical distribution in South America, was fractionated by reverse-phase HPLC and the fractions screened for phospholipase A2 (PLA2) activity. The major component of the venom, a PLA2, here referred to as 'Lemnitoxin', was isolated and characterized biochemically and toxicologically. It induces myotoxicity upon intramuscular or intravenous injection into mice. The amino acid residues Arg15, Ala100, Asn108, and a hydrophobic residue at position 109, which are characteristic of myotoxic class I phospholipases A2, are present in Lemnitoxin. This PLA2 is antigenically related to M. nigrocinctus nigroxin, Notechis scutatus notexin, Pseudechis australis mulgotoxin, and Pseudonaja textilis textilotoxin, as demonstrated with monoclonal and polyclonal antibodies. Lemnitoxin is highly selective in its targeting of cells, being cytotoxic for differentiated myotubes in vitro and muscle fibers in vivo, but not for undifferentiated myoblasts or endothelial cells. Lemnitoxin is not lethal after intravenous injection at doses up to 2μg/g in mice, evidencing its lack of significant neurotoxicity. Lemnitoxin displays anticoagulant effect on human plasma and proinflammatory activity also, as it induces paw edema and mast cell degranulation. Thus, the results of this work demonstrate that Lemnitoxin is a potent myotoxic and proinflammatory class I PLA2., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

A novel lectin present in Dioclea reflexa seeds (DrfL) was discovered and described in this study. DrfL was purified in a single step by affinity chromatography in a Sephadex G-50 column. The lectin strongly agglutinated rabbit erythrocytes and was inhibited by α-methyl-D-mannoside, D-mannose, and D-glucose. The hemagglutinating activity of DrfL is optimum at pH 5.0-7.0, stable up to 50 °C, and dependent on divalent cations. Similar to other lectins of the subtribe Diocleinae, the analysis by mass spectrometry indicated that DrfL has three chains (α, β, and γ) with masses of 25,562, 12,874, and 12,706 Da, respectively, with no disulfide bonds or glycosylation. DrfL showed inflammatory activity in the paw edema model and exhibited low cytotoxicity against Artemia sp., (Copyright © 2015 John Wiley & Sons, Ltd.)

Objectives: Rapid diagnostic tests for bacteremia are important for early treatment to improve clinical outcome. We sought to identify plasma biomarkers that can identify patients with bacteremia using an untargeted global metabolomic analysis., Methods: Plasma metabolomic profiles were analyzed for 145 adult patients with (cases) and without (controls) bacteremia using ultra-high-performance liquid chromatography/quadrupole-time-of-flight mass spectrometry (UHPLC-Q-TOF-MS). All metabolites were compared between cases and controls using a 2-tier filtering approach, and each metabolite underwent receiver operating characteristic (ROC) curve analysis. Individual metabolites that distinguish between cases and controls were characterized. Subgroup analysis was performed to identify metabolites with prognostic significance., Results: After 2-tier filtering, 128 molecular features were identified to be potential biomarkers that could distinguish cases from controls. Five metabolites had an area under the ROC curve (AUC) of >0.8 in ROC curve analysis, including a sphingolipid, an acylcarnitine, a fatty acid ester, and 2 glycerophosphocholines. These metabolites could distinguish cases from controls in the unsupervised hierarchical clustering analysis. Subgroup analysis of bacteremic patients showed that the level of trans-2,3,4-trimethoxycinnamate was lower in fatal than non-fatal cases., Conclusions: Plasma lipid mediators of inflammation can distinguish bacteremia cases from non-bacteremia controls. These biomarkers may be used as targets for rapid test in clinical practice., (Copyright © 2015 The British Infection Association. Published by Elsevier Ltd. All rights reserved.)

Excessive consumption of horticultural fruit is a double-edged sword with both positive and negative effects. In Eastern countries, a large number of people have suffered from shang huo as a result of excessive consumption of "heating" foods, such as lychee, longan, mandarin orange, mango and civet durian. The present study adopted a step by step strategy screened the compositions with pro-inflammatory effect in satsuma fruits. The pro-inflammatory effects of all fractions were evaluated in RAW 264.7 cell lines by enzyme-linked immunosorbent assay (ELISA) and RT-PCR tests. The soluble water extract (SWE) from satsuma increased the production of prostaglandin E2 (PGE2) and promoted the expression level of cyclooxygenase-2 (COX-2) mRNA. SWE and high molecular weight molecules extracted from soluble water extract (HSWE) were respectively fractionated by dialysis bags and gel filtration chromatography. The macromolecular fraction named F1 was further obtained from HSWE, and could increase the production of inflammatory mediators. Finally F1 was resolved by SDS-PAGE and six proteins were identified by mass spectrometry. Compared with other detected proteins, polygalacturonase inhibitor (PGIP) and chitinase were the most likely candidate pro-inflammatory proteins according to molecular mass, and both of them were Citrus unshiu species. cDNA sequences of PGIP and chitinase were cloned and their functions were predicted as defensive proteins by SMART analysis. Excessive intake of these defensive proteins may result in adverse food reactions in human beings, such as shang huo and other immune responses.

Background: Plasmapheresis (PP) has been used in the treatment of various immunologic disorders, and its efficacy has mainly been attributed to the removal of humoral factors and autoantibodies. Besides these effects, PP may induce modifications of the cellular immunologic status, contributing to the restoration of impaired immunologic function. The effect of PP on lymphocyte subpopulations, plasma neopterin, and cytokines in renal transplant recipients was investigated in this study., Methods: We compared pre-PP and post-PP lymphocyte subpopulations and plasma neopterin in 37, and cytokine plasma levels in 30, potential renal transplant recipients. Plasma neopterin and cytokines were measured by enzyme-linked immunosorbent assay kits, lymphocyte subsets were determined using four-color fluorescence flow cytometry., Results: Lymphocyte subpopulation counts and ratios including CD45:μL (P=0.005), CD3:μL (P=0.02), CD4DR:μL (P=0.002), CD8:μL (P=0.01), and CD8DR:μL (P=0.005) T cells; CD4DR:CD4 (P=0.009) and CD8DR:CD8 (P=0.0004) ratios; DR cells:μL (P=0.003); CD19 B lymphocytes:μL (P=0.001); and plasma levels of neopterin (P<;0.0001), soluble interleukin-1 receptor antagonist (P<;0.0001), IL-8 (P=0.0001), and tumor necrosis factor-α (P=0.008) were significantly decreased after PP as compared with before PP. The results indicate a decrease of activated DR, CD4, and CD8 T lymphocytes and B lymphocytes, and a decrease of monocyte and macrophage activation as a result of PP., Conclusion: Based on these results, we conclude that PP not only removes antibodies from the plasma but, in addition, modulates T-lymphocyte activation and the inflammatory response by decreasing plasma proinflammatory cytokines.

IL12RB1 is essential for human resistance to multiple intracellular pathogens, including Mycobacterium tuberculosis. In its absence, the proinflammatory effects of the extracellular cytokines IL-12 and IL-23 fail to occur, and intracellular bacterial growth goes unchecked. Given the recent observation that mouse leukocytes express more than one isoform from il12rb1, we examined whether primary human leukocytes similarly express more than one isoform from IL12RB1. We observed that human leukocytes express as many as 13 distinct isoforms, the relative levels of each being driven by inflammatory stimuli both in vitro and in vivo. Surprisingly, the most abundant isoform present before stimulation is a heretofore uncharacterized intracellular form of the IL-12R (termed "isoform 2") that presumably has limited contact with extracellular cytokine. After stimulation, primary PBMCs, including the CD4(+), CD8(+), and CD56(+) lineages contained therein, alter the splicing of IL12RB1 RNA to increase the relative abundance of isoform 1, which confers IL-12/IL-23 responsiveness. These data demonstrate both a posttranscriptional mechanism by which cells regulate their IL-12/IL-23 responsiveness, and that leukocytes primarily express IL12RB1 in an intracellular form located away from extracellular cytokine.

Type 1 diabetes is characterized by the autoimmune destruction of pancreatic β-cells. Recognition of major histocompatibility complex (MHC)-bound peptides is critical for both the initiation and progression of disease. In this study, MHC peptide complexes were purified from NIT-1 β-cells, interferon-γ (IFN-γ)-treated NIT-1 cells, splenic and thymic tissue of 12-week-old NOD mice, and peptides identified by mass spectrometry. In addition to global liquid chromatography-tandem mass spectrometry analysis, the targeted approach of multiple-reaction monitoring was used to quantitate the immunodominant K(d)-restricted T-cell epitope islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP)₂₀₆₋₂₁₄. We identified >2,000 MHC-bound peptides; 1,100 of these presented by β-cells grown under normal conditions or after exposure to IFN-γ. These include sequences from a number of known autoantigens. Quantitation of IGRP₂₀₆₋₂₁₄ revealed low-level presentation by K(d) (~25 complexes/cell) on NIT-1 cells after IFN-γ treatment compared with the simultaneous presentation of the endogenously processed K(d)-restricted peptide Janus kinase-1₃₅₅₋₃₆₃ (~15,000 copies/cell). We have successfully sequenced peptides from NIT-1 β-cells under basal and inflammatory conditions. We have shown the feasibility of quantitating disease-associated peptides and provide the first direct demonstration of the disparity between presentation of a known autoantigenic epitope and a common endogenously presented peptide.

Ultrafiltration with a hemoconcentrator may remove excess fluid load and alleviate tissue edema and has been universally adopted in extracorporeal circulation protocols during pediatric cardiac surgery. Balanced ultrafiltration is advocated to remove inflammatory mediators generated during surgery. However, whether balanced ultrafiltration can remove all or a portion of the inflammatory mediator load remains unclear. The inflammatory mediator removal capacity of zero-balanced ultrafiltration was measured during pediatric extracorporeal circulation in vitro. Extracorporeal circulation was composed of cardiotomy reservoir, D902 Lilliput 2 membrane oxygenator, and Capiox AF02 pediatric arterial line filter. The Hemoconcentrator BC 20 plus was placed between arterial purge line and oxygenator venous reservoir. Fresh donor human whole blood was added into the circuit and mixed with Ringer's solution to obtain a final hematocrit of 24-28%. After 2 h of extracorporeal circulation, zero-balanced ultrafiltration was initiated and arterial line pressure was maintained at approximately 100 mmHg with Hoffman clamp. The rate of ultrafiltration (12 mL/min) was controlled by ultrafiltrate outlet pressure. Identical volume of plasmaslyte A was dripped into the circuit to maintain stable hematocrit during the 45 min of the experiment. Plasma and ultrafiltrate samples were drawn every 5 min, and concentrations of inflammatory mediators including interleukin-1β (IL-1β), IL-6, IL-10, neutrophil elastase (NE), and tumor necrosis factor-α (TNF-α) were measured. All assayed inflammatory mediators were detected in the ultrafiltrate, demonstrating that the ultrafiltrator may remove inflammatory mediators. However, dynamic observations suggested that the concentration of NE was highest among the five inflammatory mediators in both plasma and ultrafiltrate (P < 0.001). IL-1β had the lowest concentration in plasma, whereas the concentration of TNF-α was the lowest in ultrafiltrate (P < 0.001). Concentrations of all inflammatory mediators in the ultrafiltrate did not increase linearly compared with those in plasma. The respective ultrafiltrate to plasma concentration and amount ratios indicated that the total removal effect of hemoconcentrator on the inflammatory mediators was 4.17 ± 2.68% for IL-1β, 0.64 ± 0.69% for IL-6, 0.24 ± 0.18% for IL-10, 2.84 ± 1.65% for NE, and 0.51 ± 0.81% for TNF-α, respectively. Balanced ultrafiltration may selectively remove inflammatory mediators from serum. Respective ratios of inflammatory mediators in ultrafiltrate compared with plasma, as well as total amount of inflammatory mediators in the ultrafiltrate suggest that balanced ultrafiltration removes a limited portion of the total inflammatory mediator load., (© 2012, Copyright the Authors. Artificial Organs © 2012, International Center for Artificial Organs and Transplantation and Wiley Periodicals, Inc.)

The innate immune system plays an important role in rheumatoid arthritis (RA) pathogenesis. Previous studies support the role of TLR2 and 4 in RA and experimental arthritis models; however, the regulation and pathogenic effect of TLR5 is undefined in RA. In this study, we show that TLR5 is elevated in RA and osteoarthritis ST lining and sublining macrophages and endothelial cells compared with normal individuals. Furthermore, expression of TLR5 is elevated in RA synovial fluid macrophages and RA peripheral blood monocytes compared with RA and normal peripheral blood in vitro-differentiated macrophages. We also found that TLR5 on RA monocytes is an important modulator of TNF-α in RA synovial fluid and that TLR5 expression on these cells strongly correlates with RA disease activity and TNF-α levels. Interestingly, TNF-α has a feedback regulation with TLR5 expression in RA monocytes, whereas expression of this receptor is regulated by IL-17 and IL-8 in RA macrophages and fibroblasts. We show that RA monocytes and macrophages are more responsive to TLR5 ligation compared with fibroblasts despite the proinflammatory response being mediated through the same signaling pathways in macrophages and fibroblasts. In conclusion, we document the potential role of TLR5 ligation in modulating transcription of TNF-α from RA synovial fluid and the strong correlation of TLR5 and TNF-α with each other and with disease activity score in RA monocytes. Our results suggest that expression of TLR5 may be a predictor for RA disease progression and that targeting TLR5 may suppress RA.

Eicosanoids are key mediators and regulators of inflammation and oxidative stress often used as biomarkers for diseases and pathological conditions such as cardiovascular and pulmonary diseases and cancer. Analytically, comprehensive and robust quantification of different eicosanoid species in a multi-method approach is problematic because most of these compounds are relatively unstable and may differ in their chemical properties. Here we describe a novel ultra-performance liquid chromatography-selected reaction monitoring mass spectroscopy (UPLC-SRM/MS) method for simultaneous quantification of key urinary eicosanoids, including the prostaglandins (PG) tetranor PGE-M, 8-iso-, and 2,3-dinor-8-iso-PGF(2α); the thromboxanes (TXs) 11-dehydro- and 2,3-dinor-TXB₂; leukotriene E₄; and 12-hydroxyeicosatetraenoic acid. In contrast to previous methods, which used time-consuming and complex solid phase extraction, we prepared samples with a simple liquid/liquid extraction procedure. Because collision-induced dissociation produced characteristic product ions for all analytes, no derivatization step for SRM/MS analysis was necessary. Analytes were separated with a short UPLC reversed-phase column (1.7 µm particles), allowing shorter run times than conventional HPLC columns. The method was validated and applied to human urine samples showing excellent precision, accuracy, detection limits, and robustness. In summary, the developed method allows robust and sensitive profiling of urinary eicosanoid species, making it a useful and valuable tool for biomarker profiling in clinical/toxicological studies.

Mammalian host response to pathogens is associated with fluctuations in high abundant proteins in body fluids as well as in regulation of proteins expressed in relatively low copy numbers like cytokines secreted from immune cells and endothelium. Hence, efficient monitoring of proteins associated with host response to pathogens remains a challenging task. In this paper, we present a targeted proteome analysis of a panel of 20 proteins that are widely believed to be key players and indicators of bovine host response to mastitis pathogens. Stable isotope-labeled variants of two concordant proteotypic peptides from each of these 20 proteins were obtained through the QconCAT method. We present the quantotypic properties of these 40 proteotypic peptides and discuss their application to research in host-pathogen interactions. Our results clearly demonstrate a robust monitoring of 17 targeted host-response proteins. Twelve of these were readily quantified in a simple extraction of mammary gland tissues, while the expression levels of the remaining proteins were too low for direct and stable quantification; hence, their accurate quantification requires further fractionation of mammary gland tissues.

Background: Human immunodeficiency virus (HIV) infected children have an eleven-fold risk of acute lower respiratory tract infection. This places HIV-infected children at risk of airway destruction and bronchiectasis., Objective: To study predisposing factors for the development of bronchiectasis in a developing world setting., Methods: Children with HIV-related bronchiectasis aged 6-14 years were enrolled. Data were collected on demographics, induced sputum for tuberculosis, respiratory viruses (respiratory syncytial virus), influenza A and B, parainfluenza 1-3, adenovirus and cytomegalovirus), bacteriology and cytokines. Spirometry was performed. Blood samples were obtained for HIV staging, immunoglobulins, immunoCAP®-specific immunoglobulin E (IgE) for common foods and aeroallergens and cytokines., Results: In all, 35 patients were enrolled in the study. Of 161 sputum samples, the predominant organisms cultured were Haemophilus influenzae and parainfluenzae (49%). The median forced expiratory volume in 1 second of all patients was 53%. Interleukin-8 was the predominant cytokine in sputum and serum. The median IgE level was 770 kU/l; however, this did not seem to be related to atopy; 36% were exposed to environmental tobacco smoke, with no correlation between exposure and CD4 count., Conclusion: Children with HIV-related bronchiectasis are diagnosed after the age of 6 years and suffer significant morbidity. Immune stimulation mechanisms in these children are intact despite the level of immunosuppression.

Abdominal aortic aneurysms (AAA) are characterized by pathological remodeling of the aortic extracellular matrix (ECM). However, besides the well-characterized elastolysis and collagenolysis little is known about changes in other ECM proteins. Previous proteomics studies on AAA focused on cellular changes without emphasis on the ECM. In the present study, ECM proteins and their degradation products were selectively extracted from aneurysmal and control aortas using a solubility-based subfractionation methodology and analyzed by gel-liquid chromatography-tandem MS and label-free quantitation. The proteomics analysis revealed novel changes in the ECM of AAA, including increased expression as well as degradation of collagen XII, thrombospondin 2, aortic carboxypeptidase-like protein, periostin, fibronectin and tenascin. Proteomics also confirmed the accumulation of macrophage metalloelastase (MMP-12). Incubation of control aortic tissue with recombinant MMP-12 resulted in the extensive fragmentation of these glycoproteins, most of which are novel substrates of MMP-12. In conclusion, our proteomics methodology allowed the first detailed analysis of the ECM in AAA and identified markers of pathological ECM remodeling related to MMP-12 activity.

Purpose of Review: To provide a narrative synthesis of evidence on interferon-gamma release assays (IGRAs) for the diagnosis of latent tuberculosis infection (LTBI) in individuals with immune-mediated inflammatory disorders (IMIDs)., Recent Findings: Only a few studies have evaluated IGRAs in IMIDs, and most were small and varied considerably with respect to the use of immunosuppressive medications and types of IMIDs. Current evidence does not clearly suggest that IGRAs are better than tuberculin skin test (TST) in identifying individuals with IMID who could benefit from LTBI treatment. To date, no studies have been done on the predictive value of IGRAs in IMID patients. Important questions remain unanswered as to the impact of immunosuppressive medications and the impact of type of IMID on IGRA performance., Summary: Despite the lack of clear evidence, there is an increasing tendency for guidelines to prefer IGRA over TST in IMIDs or to recommend both TST and IGRA to enhance sensitivity. We believe the use of either test is acceptable for LTBI screening. Clinicians could consider starting with IGRAs in individuals with a history of Bacille Calmette-Guérin (BCG) vaccination after infancy or with repeated BCG vaccinations. When the index of suspicion for LTBI is high, both IGRA and TST could be performed, especially prior to initiating TNF-α inhibitor therapy. Regardless of the test used, it is important to remember that in the face of immune-suppression, both IGRA and TST can be falsely negative and are thus only diagnostic aids - they will need to be interpreted with other clinical and risk factor data.

DwL, a lectin extracted from the seeds of Dioclea wilsonii, is a metalloprotein with strong agglutinating activity against rabbit and ABO erythrocytes, inhibited by glucose and mannose. DwL was purified by affinity chromatography on a Sephadex G-50 column and ion exchange chromatography on a HiTrap SP XL column. SDS-PAGE revealed three electrophoretic bands corresponding to the α (25,634 ± 2 Da), β (12,873 ± 2 Da) and γ (12,779 ± 2 Da) chains. Protein sequencing was done by Tandem Mass Spectrometry. The primary sequence featured 237 amino acids and was highly homologous to other reported Diocleinae lectins. A complete X-ray dataset was collected at 2.0 Å for X-Man-complexed DWL crystals produced by the vapor diffusion method. The crystals were orthorhombic and belonged to the space group I222, with the unit-cell parameters a = 59.6, b = 67.9 and c = 109.0 Å. DWL differed in potency from other ConA-like lectins and was found to induce neutrophil migration in rats, making it particularly useful in structural/functional studies of this class of proteins.

The petroleum ether extract of the red alga Laurencia obtusa afforded three new C(15) acetogenins (cyclic ether enyne): (12Z)-cis-maneonene-D (1), (12E)-cis-maneonene-E (2), and (12Z)-trans-maneonene-C (3), along with one known cis-maneonene-A (4). Blood neutrophils were prepared, cultured, and incubated for 24, 48, and 72 h in medium with and without isolated compounds. Blood neutrophils were prepared, cultured, and incubated for 24, 48 and 72 h in medium with and without the isolated compounds. Both morphology and DNA fragmentation methods assessed the percentage of neutrophils apoptosis in each culture. In the present study, several observations have been made concerning the apoptosis-inducing or inhibiting effect of 1 and 2. Both compounds had no inhibition of apoptosis but apoptosis was enhanced significantly by aging. However, 1 stimulated apoptosis of normal only at the initial 24 h. After that there was no significant difference in apoptosis with or without compound 1, while 2 stimulated apoptosis at all the times. The apoptosis induced by these two compounds was demonstrated by DNA fragmentation assay and microscopic observation. These observations suggest that compounds 1 and 2 may be involved in regulation of programmed death in the initiation and propagation of inflammatory responses.

The anti-inflammatory drugs possess many serious side effects at doses commonly prescribed. It is really important to discover novel regulators of inflammation from natural sources with minimal adverse effects. Schinus areira L. is a plant native from South America and is used in folk medicine as an anti-inflammatory herb. For this study, the activity of aqueous extracts on inflammation and the effect on superoxide anion production in mice macrophages were assayed. Aqueous extracts were prepared by soaking herbs in cold water (cold extract), boiling water (infusion), and simmering water (decoction). Cold extract possess an anti-inflammatory activity. Decoction and infusion showed pro-inflammatory activity. Cold extract increased the production of superoxide anion. It has been proposed to use diverse methods to obtain extracts of S. areira L. with different effects. Cold extract, decoction, and infusion could be utilized as extracts or as pharmacological preparations for topical application.

Hedranthera barteri, anti-inflammatory activity, anti-nociceptive activity, anti-histaminic activityL. (HB) is used in the treatment of painful conditions and oedema amongst its folkloric use. The hexane and ethyl acetate fractions of the root of H. barteri were investigated for anti-nociceptive and antiinflammatory properties and probable mechanism of action. Hot plate, tail flick, formalin-induced oedema and acetic acidinduced writhing tests were employed to investigate the anti-nociceptive activity while the anti-inflammatory activity was investigated using carrageenan-induced paw oedema. Anti-histaminic potential of HB root extracts on the rat peritoneal mast cells (RPMCs) was explored through pectrofluorometric method. The root was screened for its phytochemical components. The HB root contains alkaloids,cardenolides and saponins. HXHBR exhibited higher anti-inflammatory potentials (P <0.001). HXHBR dose-dependently (P <0.01) reduced the histamine release from the rat peritoneal mast cells which is comparable with a standard anti-histaminic drug, ketotifen. These results showed that EAHBR and HXHBR possess anti-nociceptive and anti-inflammatory activities, and suggested its mechanism of action through the inhibition of histamine, an inflammatory mediator, usually released during the early phase of allergic responses and chronic phase of inflammatory pain. Flavonoids, alkaloids and/or saponins present in HB root may be responsible for its anti-nociceptive and anti-inflammatory properties.

Aim of Study: Andrographis paniculata has been known to possess widespread traditional application in the treatment of allergy and inflammatory diseases. In the current study, we sought to examine the effects of an extract of Andrographis paniculata leaves on inhibition of lipopolysaccharide (LPS) induced [nitric oxide (NO), prostaglandin E(2) (PGE(2)), interleukin-1beta (IL-1 beta), and interleukin-6 (IL-6)] and calcimycin (A23187) induced [leukotriene B(4) (LTB(4)), thromboxane B(2) (TXB(2)) and histamine] mediators in diverse cell based models., Materials and Methods: Effect of an extract of Andrographis paniculata leaves (AP) was studied on inhibition of LPS induced NO, PGE(2), IL-1 beta and IL-6 in J774A.1 murine macrophages; A23187 induced LTB(4) and TXB(2) in HL-60 promyelocytic leukemic cells and histamine in RBL-2H3 rat basophilic leukemia cells., Results and Conclusion: AP illustrated significant alleviation of pro-inflammatory, inflammatory, and allergic mediators. However, no inhibition was observed against histamine release. This outcome has been summed up to deduce that AP is fairly potent in attenuating the inflammation by inhibiting pro-inflammatory (NO, IL-1 beta and IL-6), inflammatory (PGE(2) and TXB(2)) and allergic (LTB(4)) mediators., (Copyright 2010 Elsevier Ireland Ltd. All rights reserved.)

Sepsis is one of the main causes of death in critically ill patients. The pathophysiology of sepsis is complex and not completely understood. The proinflammatory and anti-inflammatory response leads to cell and organ dysfunction and, in many cases, death. Thus, the goal of the intervention is to restore the homeostasis of circulating mediators rather than to inhibit selectively the proinflammatory or anti-inflammatory mediators. Blood purification has been reported to remove a wide array of inflammatory mediators. The effects are broad-spectrum and auto-regulating. Blood purification has also been demonstrated to restore immune function through improving antigen-presenting capability, adjusting leukocyte recruitment, oxidative burst and phagocytosis, and improving leukocyte responsiveness. A great deal of work has to be done in order to find and optimize the best extracorporeal blood purification therapy for sepsis. New devices specifically target the pathophysiological mechanisms involved in these conditions. High-volume hemofiltration, hemoadsorption, coupled plasma filtration adsorption, and high cutoff membrane are now being tested in septic patients. Preliminary data indicate the feasibility of these modified techniques in sepsis. Their impact on patient prognosis, however, still needs proof by large randomized clinical trials. Finally, the emerging paradigm of sepsis-induced immune suppression provides additional rationale for the development of extracorporeal blood purification therapy for sepsis., (2010 S. Karger AG, Basel.)

Recently, 'super high-flux' (SHF) or 'high cut-off' (HCO) membranes have been developed to increase the clearance of inflammatory mediators. In the experimental and clinical settings, SHF/HCO membranes appear to achieve greater clearance of inflammatory cytokines than conventional high-flux membranes. SHF/HCO membranes also restore immune cell function, attenuate hemodynamic instability and decrease plasma IL-6 levels. Moreover, SHF/HCO membranes can eliminate larger late-phase inflammatory mediators such as HMGB-1. Although albumin sieving coefficients with SHF/HCO membranes are greater than with conventional high-flux membranes, the daily amount lost is limited and can be replaced. Hemodialysis with SHF/HCO membranes can also achieve similar cytokine removal to hemofiltration with acceptable albumin losses. When strategies for sepsis or systemic inflammation treatment target middle molecular mediators, both SHF/HCO hemofiltration and hemodialysis appear feasible and safe and require further clinical investigation., (Copyright 2010 S. Karger AG, Basel.)

Introduction: A pro-apoptotic effect of circulating mediators on renal tubular epithelial cells has been involved in the pathogenesis of sepsis-associated acute kidney injury (AKI). Adsorption techniques have been showed to efficiently remove inflammatory cytokines from plasma. The aim of this study was to evaluate the efficiency of the hydrophobic resin Amberchrom CG161 M to adsorb from septic plasma soluble mediators involved in tubular injury., Methods: We enrolled in the study 10 critically ill patients with sepsis-associated AKI and we evaluated the effects of their plasma on granulocyte adhesion, apoptosis and functional alterations of cultured human kidney tubular epithelial cells. We established an in vitro model of plasma adsorption and we studied the protective effect of unselective removal of soluble mediators by the Amberchrom CG161 M resin on septic plasma-induced tubular cell injury., Results: Plasma from septic patients induced granulocyte adhesion, apoptosis and altered polarity in tubular cells. Plasma adsorption significantly decreased these effects and abated the concentrations of several soluble mediators. The inhibition of granulocyte adhesion to tubular cells was associated with the down-regulation of ICAM-1 and CD40. Resin adsorption inhibited tubular cell apoptosis induced by septic plasma by down-regulating the activation of caspase-3, 8, 9 and of Fas/death receptor-mediated signalling pathways. The alteration of cell polarity, morphogenesis, protein reabsorption and the down-regulation of the tight junction molecule ZO-1, of the sodium transporter NHE3, of the glucose transporter GLUT-2 and of the endocytic receptor megalin all induced by septic plasma were significantly reduced by resin adsorption., Conclusions: Septic plasma induced a direct injury of tubular cells by favouring granulocyte adhesion, by inducing cell apoptosis and by altering cell polarity and function. All these biological effects are related to the presence of circulating inflammatory mediators that can be efficiently removed by resin adsorption with a consequent limitation of tubular cell injury.

The current study explored if alpha-iso-cubebene, a novel cubebene sesquiterpene compound purified from Schisandra chinensis, could attenuate the activities of adhesion molecules in tumor necrosis factor-alpha (TNF-alpha)-stimulated human umbilical vein endothelial cells (HUVECs). The study was performed on HUVECs that were pretreated with 25 microg/ml of alpha-iso-cubebene before TNF-alpha treatment. Treatment of HUVECs with alpha-iso-cubebene for 6 h significantly inhibited TNF-alpha-induced reactive oxygen species (ROS) formation. HUVECs treated with alpha-iso-cubebene showed markedly suppressed TNF-alpha-induced mRNA expression of VCAM-1 and E-selectin, but little alteration in ICAM-1 mRNA expression. alpha-iso-Cubebene treatment also significantly decreased the TNF-alpha-induced cell surface and total protein expression of VCAM-1 and E-selectin without affecting ICAM-1 expression. In addition, treatment of HUVECs with alpha-iso-cubebene markedly reduced U937 monocyte adhesion to TNF-alpha-stimulated HUVECs. alpha-iso-Cubebene treatment did not affect translocation of NF-kappaB transcription factor from the cytosol into the nucleus. However, alpha-iso-cubebene significantly inhibited NF-kappaB transcription factor activation in TNF-alpha-stimulated HUVECs. The new anti-inflammatory agent alpha-iso-cubebene attenuates TNF-alpha-stimulated endothelial adhesion to monocytes by inhibiting intracellular ROS production, the activation of redox-sensitive NF-kappaB transcription factor and expression of VCAM-1 and E-selectin. Based on these findings, alpha-iso-cubebene is proposed as an effective new anti-inflammatory agent that may have a potential therapeutic use for the prevention and treatment of vascular diseases.

Background: High-volume haemofiltration (HVHF) has been suggested as an adjuvant treatment of septic shock due to its capacities to remove from blood both pro- and anti-inflammatory mediators involved in the sepsis syndrome. Adsorption properties of some haemofiltration membranes are also interesting with this indication because inflammatory mediators are caught in the membrane itself. The aim of this study was to determine the haemodynamic and immunological effects of a new haemofiltration membrane, which has enhanced adsorption properties due to a special surface treatment, allowing the adsorption of endotoxins., Methods: We compared this membrane to a standard haemofiltration membrane both in vitro and in 20 sepsis-induced pigs, randomized in two groups. One group was haemofiltered with the treated membrane and the other with the standard haemofiltration membrane during 6-h HVHF sessions., Results: At the end of the experiment, mean +/- SD crystalloids requirements (5937 +/- 1588 versus 7587 +/- 1456 ml, P = 0.026), colloids requirements (1437 +/- 320 versus 1912 +/- 538 ml, P = 0.027), lactic acidosis (pH = 7.20 +/- 0.11 versus 7.10 +/- 0.07, P = 0.026) and pulmonary arterial hypertension (MPAP = 24 +/- 7 versus 34 +/- 8 mmHg, P = 0.008) were less pronounced when HVHF was performed with the treated membrane. In addition, mean +/- SD endotoxins levels were lower in the treated membrane group after 1 hour of HVHF (1.91 +/- 1.19 versus 11.07 +/- 10.64 EU/ml, P = 0.035). Cytokines levels were not different between groups except for IL-1beta, which was slightly lower in the treated membrane group., Conclusions: The use of a membrane with enhanced adsorption properties during a 6-h HVHF session in septic pigs improves haemodynamics compared to a standard haemofiltration membrane. These results are probably due to an efficient endotoxins and cytokines adsorption. A human study using this membrane is now necessary to confirm these results.

The skin is constantly exposed to commensal microflora and pathogenic microbes. The stratum corneum of the outermost skin layer employs distinct tools such as harsh growth conditions and numerous antimicrobial peptides (AMPs) to discriminate between beneficial cutaneous microflora and harmful bacteria. How the skin deals with microbes that have gained access to the live part of the skin as a result of microinjuries is ill defined. In this study, we report that the chemokine CXCL14 is a broad-spectrum AMP with killing activity for cutaneous gram-positive bacteria and Candida albicans as well as the gram-negative enterobacterium Escherichia coli. Based on two separate bacteria-killing assays, CXCL14 compares favorably with other tested AMPs, including human beta-defensin and the chemokine CCL20. Increased salt concentrations and skin-typical pH conditions did not abrogate its AMP function. This novel AMP is highly abundant in the epidermis and dermis of healthy human skin but is down-modulated under conditions of inflammation and disease. We propose that CXCL14 fights bacteria at the earliest stage of infection, well before the establishment of inflammation, and thus fulfills a unique role in antimicrobial immunity.

Almost 10 years ago, standard hemofiltration (HF) was carried out at an ultrafiltration (UF) rate of not more than 2 1/hour and only a predilution variant was usually used. However, after Ronco published the results of his study early in the 20th century the situation changed as this study provided evidence that increasing an UF rate up to 35 ml/kg/hr had a positive effect in patients with acute renal failure (ARF). At the same time, there was a new standard for the determination of high-volume HF (HVHF) and new concepts generalized in this paper have emerged. Moreover, two different procedures of HVHF: (1) a continuous high-volume technique providing a rate of 50-70 ml/kg/hr during 24 hours; (2) an intermittent high-volume technique, the so-called HVHF with short courses of treatment, a very high volume at a rate of 100-120 ml/kg/hr during 4-8 hours. Later on this technique came to be called pulse HVHF. Both methods are designated by the abbreviation HVHF, but their concepts and results are slightly different, as will be discussed below. As for the higher doses of plasma replacement therapy, it is necessary to put the results of two recently published studies into routine clinical practice of continuous vein-venous HF (CVVHF) as soon as possible just in 2008. The performed studies suggest the most important role of adequate dosage of CVVHF and show that the use of a dose of 35 ml/kg/hr in critically ill patients with ARF increased survival by almost 20%. These two studies contain today's most vivid evidence, but one awaits the results of other studies that confirm (or do not confirm) these conclusions. In the world where more importance is attached to evidence-based medicine, two first-level studies will yield Stage A recommendations that may be guided by any intensive care specialist using CVVHF in expectation of the results of on-going investigations. Nevertheless, many difficulties are to be surmounted while introducing new procedures into routine intensive care practice. The most cardinal problems are those associated with requirements for blood flow via vascular access with pre- and postdilution procedures, with the type of employed membranes and substituting liquids, and with a possible need for concomitant dialysis. From the strictly practical standpoint, introduction of these changes into routine intensive care practice may be extremely difficult. This will undoubtedly require that physicians and nurses should make joint effects.

A variety of biologically active compounds with pharmacological applications has been reported to occur in marine sponges. The present study was undertaken to provide a set of data about an extract from Aplysina caissara, a Brazilian marine sponge. The antinociceptive and anti-inflammatory effects were investigated against different experimental models in mice. When evaluated against writhing test intraperitoneally (60 and 90 mg/kg), the extract significantly inhibited abdominal constriction by 33.7% and 41.4% respectively. In the formalin test (60 and 90 mg/kg), the extract of sponge inhibited 43.6% and 51.6% in the first phase and 98.2% and 97.2% in the second phase respectively. When evaluated against the hot plate test, both doses demonstrated activity. An increase in the hot plate latency was observed after 60 min. The anti-inflammatory effect was evaluated by formalin-induced mice paw edema. Extract from A. caissara (60 and 90 mg/kg) significantly reduced hind paw swelling. Mortality increased with increasing doses, with LD(50) of 212.2 mg/kg for intraperitoneal administration. These results demonstrated that the extract of the marine sponge A. caissara possesses antinociceptive and anti-edematogenic effects.

Background: Cannabinoid-induced analgesia was shown in animal studies of acute inflammatory and neuropathic pain. In humans, controlled clinical trials with Delta-tetrahydrocannabinol or other cannabinoids demonstrated analgesic efficacy in chronic pain syndromes, whereas the data in acute pain were less conclusive. Therefore, the aim of this study was to investigate the effects of oral cannabis extract in two different human models of acute inflammatory pain and hyperalgesia., Methods: The authors conducted a double-blind, crossover study in 18 healthy female volunteers. Capsules containing Delta-tetrahydrocannabinol-standardized cannabis extract or active placebo were orally administered. A circular sunburn spot was induced at one upper leg. Heat and electrical pain thresholds were determined at the erythema, the area of secondary hyperalgesia, and the contralateral leg. Intradermal capsaicin-evoked pain and areas of flare and secondary hyperalgesia were measured. Primary outcome parameters were heat pain thresholds in the sunburn erythema and the capsaicin-evoked area of secondary hyperalgesia. Secondary measures were electrical pain thresholds, sunburn-induced secondary hyperalgesia, and capsaicin-induced pain., Results: Cannabis extract did not affect heat pain thresholds in the sunburn model. Electrical thresholds (250 Hz) were significantly lower compared with baseline and placebo. In the capsaicin model, the area of secondary hyperalgesia, flare, and spontaneous pain were not altered., Conclusion: To conclude, no analgesic or antihyperalgesic activity of cannabis extract was found in the experiments. Moreover, the results even point to the development of a hyperalgesic state under cannabinoids. Together with previous data, the current results suggest that cannabinoids are not effective analgesics for the treatment of acute nociceptive pain in humans.

High mobility group box 1 protein (HMGB1), originally characterized as a nuclear DNA-binding protein, has also been described to have an extracellular role when it is involved in cellular activation and proinflammatory responses. In this study, FLAG-tagged HMGB1 was inducibly expressed in the presence of culture media with or without added IL-1beta, IFN-gamma, or TNF-alpha. HMGB1 purified from cells grown in culture media alone only minimally increased cytokine production by MH-S macrophages and had no effect on murine neutrophils. In contrast, HMGB1 isolated from cells cultured in the presence of IL-1beta, IFN-gamma, and TNF-alpha had enhanced proinflammatory activity, resulting in increased production of MIP-2 and TNF-alpha by exposed cells. IL-1beta was bound to HMGB1 isolated from cells cultured with this cytokine, and purified HMGB1 incubated with recombinant IL-1beta acquired proinflammatory activity. Addition of anti-IL-1beta Abs or the IL-1 receptor antagonist to cell cultures blocked the proinflammatory activity of HMGB1 purified from IL-1beta-exposed cells, indicating that such activity was dependent on interaction with the IL-1 receptor. These results demonstrate that HMGB1 acquires proinflammatory activity through binding to proinflammatory mediators, such as IL-1beta.

A dialyzable low molecular weight proinflammatory factor (X (2)) from rat spleen lymphocytes was isolated through a combination of gel filtration and high voltage paper electrophoresis (HVE) and then partially characterized. It was able to potentiate the formation of carrageenin induced edema on the rat paw. Its amino acid analysis revealed Glu, Cys and Gly (1:1:1), but gammaGlu as N-terminal residue, initially suggesting oxidized glutathione (GSSG), since it showed exactly the same HV electrophoretic mobility as GSSG at pH 6.5. However, neither GSSG nor a synthetic homologue showed any proinflammatory activity. On basis of its infrared spectrum, HVE mobility and presence of a gamma-Glu-Cys-Gly (GSH) moiety, the hypothesis of identity of X (2) with leukotriene C(4) (LTC(4)) was raised. Once again it was not confirmed, since LTC(4) did not show any proinflammatory activity too, leading us to infer that, even excluding LTC(4), our data are consistent with a structure bearing a GSH moiety conjugated with a hydroxylated insaturated fatty acid chain which contributes a -COO(-) group, thus providing a final net charge of -2 at pH 6.5 and an Mr = 600-650.

Background: Understanding the pathophysiology of chemokine secretion in endometriosis may offer a novel area of therapeutic intervention. This study aimed to identify chemokines differentially expressed in epithelial glands in eutopic endometrium from normal women and those with endometriosis, and to establish the expression profiles of key chemokines in endometriotic lesions., Methods: Laser capture microdissection isolated epithelial glands from endometrial eutopic tissue from women with and without endometriosis in the mid-secretory phase of their menstrual cycles. Gene profiling of the excised glands used a human chemokine and receptor cDNA array. Selected chemokines were further examined using real-time PCR and immunohistochemistry., Results: 22 chemokine/receptor genes were upregulated and two downregulated in pooled endometrial epithelium of women with endometriosis compared with controls. CCL16 and CCL21 mRNA was confirmed as elevated in some women with endometriosis compared to controls on individual samples. Immunoreactive CCL16 and CCL21 were predominantly confined to glands in eutopic and ectopic endometrium: leukocytes also stained. Immunoreactive CCL16 was overall higher in glands in ectopic vs. eutopic endometrium from the same woman (P < 0.05). Staining for CCL16 and CCL21 was highly correlated in individual tissues., Conclusion: This study provides novel candidate molecules and suggests a potential local role for CCL16 and CCL21 as mediators contributing to the inflammatory events associated with endometriosis.

This paper reports the cloning and sequencing of two pro-inflammatory cytokines, interleukin (IL)-1beta and IL-8, in haddock (Melanogrammus aeglefinus) by homology cloning. The complete transcript of the haddock IL-1beta was sequenced and contained 1043 bp, including a 762 bp open reading frame. The 3' end of the gene includes a polyadenylation signal 13 bp upstream of the poly(A) tail, along with 10 instability motifs. The predicted protein of 253aa revealed the presence of the IL-1 family signature and the absence of an ICE cut site. The cDNA of the chemokine IL-8 was sequenced in haddock and contained 903 bp of which 306 bp are the open reading frame. Interestingly, the predicted protein sequence of 101aa, contains an ELR motif preceding the CXC signature, common in all vertebrate IL-8 molecules but absent in all teleost genes sequenced to date. The expression of both haddock cytokines was studied in four different tissues: head kidney, spleen, liver and gill. Tissues were obtained from both healthy fish and fish stimulated in vivo with four commercial serotypes of LPS, namely Escherichia coli 026:B6, 055:B5, 0111:B4 and 0127:B8 and PMA. Haddock IL-1beta was not constitutively expressed and expression was only observed following stimulation. However, this expression was stimulant dependent and only PMA and LPS 026:B6 induced high levels of expression in the head kidney. The haddock IL-8 gene on the other hand, showed a constitutive expression, that could be up or down-regulated depending on the immunostimulant used, although to a lesser extent than IL-1beta.

Macrophage migration inhibitory factor (MIF) is a pro-inflammatory mediator with the ability to induce various immunomodulatory responses and override glucocorticoid-driven immunosuppression. Some of these functions have been linked to the unusual enzymatic properties of the protein, namely tautomerase and oxidoreductase activities. However, there are conflicting reports regarding the functional role of these enzymatic properties in normal physiological homeostasis and disease progression. Therefore, we have produced a highly pure, virtually endotoxin-free recombinant MIF preparation and fully characterized this using a variety of biochemical and biophysical approaches. The recombinant protein, with demonstrable enzymatic activity, was then used to systematically examine the biological activity of MIF. Surprisingly, treatment with MIF alone failed to induce cytokine expression, with the exception of IL-8. However, co-treatment of lipopolysaccharide (LPS) in conjunction with MIF produced synergistic secretion of tumor necrosis factor-alpha, interleukin (IL)-1, and IL-8 compared with LPS alone. The potentiating effect of MIF was seen at physiologically relevant concentrations. These data suggest that MIF has no conventional cytokine activity but, rather, acts to modulate and amplify the response to LPS.

Cell-derived microparticles (MP) are membrane fragments shed by virtually all eukaryotic cells upon activation or during apoptosis that play a significant role in physiologically relevant processes, including coagulation and inflammation. We investigated whether MP derived from monocytes/macrophages have the potential to modulate human airway epithelial cell activation. Monocytes/macrophages were isolated from the buffy coats of blood donors by Ficoll gradient centrifugation, followed by overnight culture of the mononuclear cell fraction. Adherent cells were washed and incubated with the calcium ionophore, A23187, or with histamine. The MP-containing supernatant was incubated with cells of the human bronchial epithelial line BEAS-2B and of the human alveolar line A549. IL-8, MCP-1, and ICAM-1 production was assessed by ELISA and by RT-PCR. In some experiments, monocytes/macrophages were stained with the fluorescent lipid intercalating dye PKH67, and the supernatant was analyzed by FACS. Stimulation of monocytes/macrophages with A23187 caused the release of particles that retain their fluorescent lipid intercalating label, indicating that they are derived from cell membranes. Incubation with A549 and BEAS-2B cells up-regulate IL-8 synthesis. Ultrafiltration and ultracentrifugation of the material abolished the effect, indicating that particulate matter, rather than soluble molecules, is responsible for it. Up-regulation of MCP-1 and ICAM-1 was also demonstrated in A549 cells. Similar results were obtained with histamine. Our data show that human monocytes/macrophages release MP that have the potential to sustain the innate immunity of the airway epithelium, as well as to contribute to the pathogenesis of inflammatory diseases of the lungs through up-regulation of proinflammatory mediators.