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ABSTRACTThis study aimed to assess movements occurring during goal scoring situations in a female professional league.Data from all the goals on the Women’s Super League 2018/2019 were collected through time-motion analysis using a modified version of the Bloomfield Movement Classification with differences analysed through chi-square. Analysis was performed on players (assistant, scorer [attackers], defender of assistant and defender of scorer [defenders]), movements, intensities and directions.Linear advancing motion (walking, jogging, running or sprint) (total percentage [95% CI] 37% attackers and 32.7% defenders) was the most common action preceding a goal, followed by deceleration (21.5% attackers; 18.4% defenders) and turn (19.2% attackers; 17.6% defenders). Other movements involved but with lower percentages were change in angle run (cut and arc run), ball blocking, lateral advancing motion (crossover and shuffle) and jumps. Players displayed similar tendencies but presented variations based on the role, with attackers performing more linear actions, subtle turns and cuts and defenders more ball blocking actions, lateral movements and high intensity linear actions and decelerations. Assistant performed the less percentage of involvements with at least 1 high intensity action (67.4%), scorer and defender of assistant showed similar values (86.3% and 87.1%), while defender of scorer had the highest percentage (97.3%).This study shows the importance of linear actions with other movements also being of high significance but with differentiated characteristics based on the role. This study could help practitioners design drills for the enhancement of physical capabilities related to movements occurring in goal scoring situations.

Human neutrophil elastase (HNE) plays an essential role in host defense against bacteria but is also involved in several respiratory diseases. Recent reports suggest that compounds exhibiting a combination of HNE inhibitory activity with antiradical properties may be therapeutically beneficial for the treatment of respiratory diseases involving inflammation and oxidative stress. We report here the synthesis and biological evaluation of novel ebselen analogues exhibiting HNE inhibitory and antiradical activities. HNE inhibition was evaluated in an enzymatic system using human HNE, whereas antiradical activity was evaluated in a cell-based assay system using phorbol 12-myristate 13-acetate (PMA)-stimulated murine bone marrow leukocytes as the source of reactive oxygen species (ROS). HNE inhibition was due to the N–CO group targeting Ser195-OH at position 2 of the scaffold, while antiradical activity was due to the presence of the selenium atom. The most active compounds 4d, 4f, and 4jexhibited a good balance between anti-HNE (IC50= 0.9–1.4 μM) and antiradical activity (IC50= 0.05–0.7 μM). Additionally, the solid-state structure of 4dwas determined and compared to that of the similar compound N-propionyl-1,2-benzisoselenazol-3(2H)-one.

ABSTRACTData were collected through time-motion analysis from soccer players participating in the English Premier League using a modified version of the Bloomfield Movement Classification with differences analysed through chi-square.The most common individual movement preceding a goal was a linear advancing motion (32.4 ± 1%), followed by deceleration (20.2 ± 0.9%) and turn (19.8 ± 0.9%). Actions also involved were change in angle run (cut and arc run), ball blocking, lateral advancing motion (crossover and shuffle) and jumps. Although players followed similar trends, there were dissimilarities based on the role, with attackers (assistant and scorer) performing more linear actions, subtle turns and cuts and defenders (defender of assistant and defender of scorer) more ball blockings, lateral movements and arc runs. In 82.9 ± 1.5% of player involvements, there was at least one high intensity (HI) movement with assistant showing the lowest percentage and defender of scorer the highest.This study shows the multidirectional nature and context specificity of soccer during goal scoring situations, with linear actions such as sprints being the most common movements, followed by decelerations and turns. Moreover, it highlights the recurrent application of these at HI, and so, training strategies should prioritize the development of player’s explosiveness.

As time goes by, J2ME is the buzzword that's appearing in more and more magazines and talked about beside more office coffee machines than any other. And slowly, the big […]

ROBERT CUMBERFORD in the May issue featuring your 2018 All-Stars opines regarding the Volvo V90 T6, 'This is the one that I can imagine buying and keeping in use for [...]

Formyl peptide receptor 2 (FPR2) agonists can boost the resolution of inflammation and can offer alternative approaches for the treatment of pathologies with underlying chronic neuroinflammation, including neurodegenerative disorders. Starting from the FPR2 agonist 2previously identified in our laboratory and through fine-tuning of FPR2 potency and metabolic stability, we have identified a new series of ureidopropanamide derivatives endowed with a balanced combination of such properties. Computational studies provided insights into the key interactions of the new compounds for FPR2 activation. In mouse microglial N9 cells and in rat primary microglial cells stimulated with lipopolysaccharide, selected compounds inhibited the production of pro-inflammatory cytokines, counterbalanced the changes in mitochondrial function, and inhibited caspase-3 activity. Among the new agonists, (S)-11lstands out also for the ability to permeate the blood–brain barrier and to accumulate in the mouse brain in vivo, thus representing a valuable pharmacological tool for studies in vivo.

ABSTRACTResearch into the physical demands of Gaelic football is limited with no research into the agility or change of direction (CoD) demands of the sport. This study examined the CoD demands of Gaelic football via a time-motion analysis of senior inter-county match play. The Bloomfield movement classification (BMC) was adapted for application to Gaelic football. A new “descriptor” was used in an effort to account for the decision-making component of agility by isolating actions that occurred during active engagement with play. Of 1,899 changes of direction (CoDs) identified, 1,035 occurred during active engagement in play. The left/right split for CoDs during active engagement in play was 47.1/49.9%, indicating no preference for completing actions to one side over the other. Whilst the most common CoDs were ≤90° (74.9%), 80% of CoDs greater than 270° took place during active engagement in play. CoD actions are very common in Gaelic football and may be more common than in other field and court sports. It is important that athletes are physically prepared to cope with the demands of very acute CoDs during meaningful periods of match play.

I didn't subscribe to your magazine to feed my passion for cars; I have a subscription to Car and Driver for that. I welcome and respect the new direction, so keep it up. - Doug Pahl Jr. Monona, WI I'm old enough to remember when Car and Driver was a car magazine. I've loved it as Car and Driver since I bought my first C/D subscription from David E. Davis for $4 at a folding card table in the parking lot during the 12 Hours of Sebring in the '60s. So, in the September issue we get not one but two annoying social-justice articles, four pages with essentially a single Bronco photo but zero content, an article on what first-time car owners in New York City will do with their cars, a full-page virtual car-warranty spam call from Ezra Dyer, and, in a supreme display of irony, a column on "reading the crowd" from your esteemed Ed. herself. [Extracted from the article]

Calculate using the Mustang's actual displacement of2264 cc and the Supra's 1998 cc, carry the one, and you get our correct percentage - Ed. - John Gatliff Mustang, OK R RATED In the long-term test of the 2019 Honda Civic Type R [November 2020], the service timeline shows that you replaced the left front tire at 17,731 miles and the right front tire only two weeks and 1000 miles later. - Ed Tinsley Austin, TX You, sir, are refined-Ed. Burke's never-ending behind-the-scenes efforts got us through deadline-week power outages, stories being killed in the 11th hour, and the ridiculousness that perpetually derails this place-Ed. [Extracted from the article]

Most injuries to the ulnar collateral ligament of the thumb, including some complete tears, will heal with immobilization, but some complete tears require surgery. In cases where the aponeurosis of the adductor pollicis muscle becomes pressed between the ligament ends, surgery is the only way to prevent chronic instability and weakness. This Stener's lesion can be determined by stress testing and the presence of a large mass on the ulnar side of the thumb.

The modulation of phagocyte responses is essential for successful performance of biomaterials in order to prevent negative outcomes associated with inflammation. Herein, we developed electrospun poly(ε-caprolactone) (PCL) scaffolds doped with the novel potent c-Jun N-terminal kinase (JNK) inhibitors 11H-indeno[1,2-b]quinoxalin-11-one oxime (IQ-1) and 11H-indeno[1,2-b]quinoxalin-11-one O-(O-ethylcarboxymethyl) oxime(IQ-1E) as a promising approach for modulating phagocyte activation. Optimized electrospinning parameters allowed us to produce microfiber composite materials with suitable mechanical properties. We found that embedded compounds were bound to the polymer matrix via hydrophobic interactions and released in two steps, with release mostly controlled by Fickian diffusion. The fabricated scaffolds doped with active compounds IQ-1and IQ-1Eeffectively inhibited phagocyte inflammatory responses. For example, they suppressed human neutrophil activation by the biomaterials, as indicated by decreased neutrophil reactive oxygen species (ROS) production and Ca2+mobilization. In addition, they inhibited lipopolysaccharide (LPS)-induced NF-κB/AP-1 reporter activity in THP-1Blue cells and interleukin (IL)-6 production in MonoMac-6 cells without affecting cell viability. These effects were attributed to the released compounds rather than cell–surface interactions. Therefore, our study demonstrates that doping tissue engineering scaffolds with novel JNK inhibitors represents a powerful tool for preventing adverse immune responses to biomaterials as well as serves as a platform for drug delivery.

To compare disease activity and disability over 2 years in early rheumatoid arthritis (RA) before and after implementation of treat‐to‐target therapy and identify predictors of adverse outcome. The Yorkshire Early Arthritis Register (YEAR) recruited 725 patients with early RAbetween 2002 and 2009, treated with a step‐up approach. The Inflammatory Arthritis Continuum study (IACON) recruited cases between 2010 and 2014 and treated to target. A total of 384 IACONcases met 2010 American College of Rheumatology/European League Against Rheumatism criteria. Latent growth curves of change in Disease Activity Score in 28 joints (DAS28) and the Health Assessment Questionnaire (HAQ) were compared between YEARand IACON. Latent class growth analysis identified trajectories of change. Baseline predictors of trajectories were identified using logistic regression. The mean DAS28 over 2 years was lower in IACONthan in YEAR. Latent trajectories of HAQchange in YEARwere high stable (21% of cohort), moderate reducing (35%), and low reducing (44%). Only moderate reducing (66%) and low reducing (34%) were seen in IACON. In both cohorts, female sex and fatigue predicted adverse HAQtrajectories (high stable and moderate reducing). Odds ratios (ORs) for moderate reducing compared to low reducing for women were 2.58 (95% confidence interval [95% CI] 1.69, 4.49) in YEARand 5.81 (95% CI2.44, 14.29) in IACON. ORs per centimeter fatigue visual analog score were 1.13 (95% CI1.07, 1.20) in YEARand 1.16 (95% CI1.12, 1.20) in IACON. Treat‐to‐target therapy gave more favorable trajectories of change in DAS28 and HAQ, but adverse HAQtrajectory was more likely in women with greater fatigue, suggesting such patients would benefit from interventions to improve function as well as reduce inflammation.

A provincial judicial inquiry has been struck to determine how and why hundreds of breast cancer patients received inaccurate hormone receptor tests at Newfoundland and Labrador's main medical laboratory. Meanwhile, [...]

Activation of neutrophils by Ferula iliensisessential oils enriched in (E/Z)-propenyl sec-butyl disulfides is mediated by activation of transient receptor potential V1 channels. Essential oil extracts from Ferula iliensishave been used traditionally in Kazakhstan for treatment of inflammation and other illnesses. Because little is known about the biologic activity of these essential oils that contributes to their therapeutic properties, we analyzed their chemical composition and evaluated their phagocyte immunomodulatory activity. The main components of the extracted essential oils were (E)-propenyl sec-butyl disulfide (15.7–39.4%) and (Z)-propenyl sec-butyl disulfide (23.4–45.0%). Ferulaessential oils stimulated [Ca2+]imobilization in human neutrophils and activated ROS production in human neutrophils and murine bone marrow phagocytes. Activation of human neutrophil [Ca2+]iflux by Ferulaessential oils was dose-dependently inhibited by capsazepine, a TRPV1 channel antagonist, indicating that TRPV1 channels mediate this response. Furthermore, Ferulaessential oils stimulated Ca2+influx in TRPV1 channel–transfected HEK293 cells and desensitized the capsaicin-induced response in these cells. Additional molecular modeling with known TRPV1 channel agonists suggested that the active component is likely to be (Z)-propenyl sec-butyl disulfide. Our results provide a cellular and molecular basis to explain at least part of the beneficial therapeutic properties of FEOs.

There is a paucity of information comparing competitive rugby league match play dynamics between northern and southern hemispheres. Notably, differences in the match demands of games played in an intensive period have not previously been reported. This study is the first to assess such demands, quantified using GPS/accelerometer technology, during a competitive three game period that comprised two games in England, interspersed with one game in Australia. The three games were completed over a period of 23 days. In-game data from fifteen elite level rugby league players were collected. The focus was to assess differences in activity profiles undertaken in each game. There were significant increases in the total number of high speed sprints, distance covered at high speed and acceleration/deceleration efforts undertaken in Australia when compared to England. No significant differences in other key performance indicators were observed. The current findings demonstrate minimal differences in the activity profiles of game play in elite professional rugby league, with the exception of high-speed activity and acceleration/deceleration efforts. The European team were defeated in their game in Australia, with clearly higher levels of total high speed sprints, acceleration and deceleration efforts being observed in that game when compared with games undertaken in England. Such findings emphasise the continuing use of GPS/accelerometer technologies in determining in-game performance characteristics associated with likely success, though the milieu of factors contributing to success must be considered in entirety.

The coherent amplifying network laser is based on an array of thousands of active laser fibers coherently combined to generate high peak-power pulses at a high repetition rate. To achieve such a massive network, new combination architectures are presented here. They are based on implementing a spherical array of amplifying fibers, thus removing the need for transport fibers from the initial scheme. These designs present an advantage in terms of scalability leading to significant reduction of the temporal fluctuations compared to those of a conventional high peak-power laser. Noise evolution with fiber number is calculated using a perturbative analysis of each channel parameters (phase, signal intensity, beam profile).

This paper provides a systematic and critical review of dynamic experimental rigs used for insect wing aerodynamics research. The goal is to facilitate meaningful comparison of data from existing rigs and provide insights for designers of new rigs. The scope extends from simple one degree of freedom rotary rigs to multi degrees of freedom rigs allowing various rotation and translation motions. Experimental methods are characterized using a consistent set of parameters that allows objective comparison of different approaches. A comprehensive catalogue is presented for the tested flow conditions (assessed through Reynolds number, Rossby number and advance ratio), wing morphologies (assessed through aspect ratio, planform shape and thickness to mean chord ratio) and kinematics (assessed through motion degrees of freedom). Links are made between the type of aerodynamic characteristics being studied and the type of experimental set-up used. Rig mechanical design considerations are assessed, and the aerodynamic measurements obtained from these rigs are discussed.

PVL‐mediated priming of PMNs enhances the host innate immune response. CA‐MRSA infections are often caused by strains encoding PVL, which can cause lysis of PMNs and other myeloid cells in vitro, a function considered widely as the primary means by which PVL might contribute to disease. However, at sublytic concentrations, PVL can function as a PMN agonist. To better understand this phenomenon, we investigated the ability of PVL to alter human PMN function. PMNs exposed to PVL had enhanced capacity to produce O2−in response to fMLF, but unlike priming by LPS, this response did not require TLR signal transduction. On the other hand, there was subcellular redistribution of NADPH oxidase components in PMNs following exposure of these cells to PVL—a finding consistent with priming. Importantly, PMNs primed with PVL had an enhanced ability to bind/ingest and kill Staphylococcus aureus. Priming of PMNs with other agonists, such as IL‐8 or GM‐CSF, altered the ability of PVL to cause formation of pores in the plasma membranes of these cells. Microarray analysis revealed significant changes in the human PMN transcriptome following exposure to PVL, including up‐regulation of molecules that regulate the inflammatory response. Consistent with the microarray data, mediators of the inflammatory response were released from PMNs after stimulation with PVL. We conclude that exposure of human PMNs to sublytic concentrations of PVL elicits a proinflammatory response that is regulated in part at the level of gene expression. We propose that PVL‐mediated priming of PMNs enhances the host innate immune response.

We tested the hypothesis that reactive oxygen species (ROS) contributed to renal hypoxia in C57BL/6 mice with ⅚ surgical reduction of renal mass (RRM). ROS can activate the mitochondrial uncoupling protein 2 (UCP-2) and increase O2usage. However, UCP-2 can be inactivated by glutathionylation. Mice were fed normal (NS)- or high-salt (HS) diets, and HS mice received the antioxidant drug tempol or vehicle for 3 mo. Since salt intake did not affect the tubular Na+transport per O2consumed (TNa/QO2), further studies were confined to HS mice. RRM mice had increased excretion of 8-isoprostane F2αand H2O2, renal expression of UCP-2 and renal O2extraction, and reduced TNa/QO2(sham: 20 ± 2 vs. RRM: 10 ± 1 μmol/μmol; P< 0.05) and cortical Po2(sham: 43 ± 2, RRM: 29 ± 2 mmHg; P< 0.02). Tempol normalized all these parameters while further increasing compensatory renal growth and glomerular volume. RRM mice had preserved blood pressure, glomeruli, and patchy tubulointerstitial fibrosis. The patterns of protein expression in the renal cortex suggested that RRM kidneys had increased ROS from upregulated p22phox, NOX-2, and -4 and that ROS-dependent increases in UCP-2 led to hypoxia that activated transforming growth factor-β whereas erythroid-related factor 2 (Nrf-2), glutathione peroxidase-1, and glutathione-S-transferase mu-1 were upregulated independently of ROS. We conclude that RRM activated distinct processes: a ROS-dependent activation of UCP-2 leading to inefficient renal O2usage and cortical hypoxia that was offset by Nrf-2-dependent glutathionylation. Thus hypoxia in RRM may be the outcome of NADPH oxidase-initiated ROS generation, leading to mitochondrial uncoupling counteracted by defense pathways coordinated by Nrf-2.

In efforts to identify novel small molecules with anti-inflammatory properties, we discovered a unique series of tetracyclic indenoquinoxaline derivatives that inhibited lipopolysaccharide (LPS)-induced nuclear factor-κB/activating protein 1 activation. Compound IQ-1(11H-indeno[1,2-b]quinoxalin-11-one oxime) was found to be a potent, noncytotoxic inhibitor of pro-inflammatory cytokine [interleukin (IL)-1α, IL-1β, IL-6, IL-10, tumor necrosis factor (TNF)-α, interferon-γ, and granulocyte-macrophage colony-stimulating factor] and nitric oxide production by human and murine monocyte/macrophages. Three additional potent inhibitors of cytokine production were identified through further screening of IQ-1analogs. The sodium salt of IQ-1inhibited LPS-induced TNF-α and IL-6 production in MonoMac-6 cells with IC50values of 0.25 and 0.61 μM, respectively. Screening of 131 protein kinases revealed that derivative IQ-3[11H-indeno[1,2-b]quinoxalin-11-one-O-(2-furoyl)oxime]was a specific inhibitor of the c-Jun N-terminal kinase (JNK) family, with preference for JNK3. This compound, as well as IQ-1and three additional oxime indenoquinoxalines, were found to be high-affinity JNK inhibitors with nanomolar binding affinity and ability to inhibit c-Jun phosphorylation. Furthermore, docking studies showed that hydrogen bonding interactions of the active indenoquinoxalines with Asn152, Gln155, and Met149 of JNK3 played an important role in enzyme binding activity. Finally, we showed that the sodium salt of IQ-1had favorable pharmacokinetics and inhibited the ovalbumin-induced CD4+T-cell immune response in a murine delayed-type hypersensitivity model in vivo. We conclude that compounds with an indenoquinoxaline nucleus can serve as specific small-molecule modulators for mechanistic studies of JNKs as well as a potential leads for the development of anti-inflammatory drugs.

N-Formyl peptide receptors (FPRs) are G protein-coupled receptors (GPCRs) involved in host defense and sensing cellular dysfunction. Thus, FPRs represent important therapeutic targets. In the present studies, we screened 32 ligands (agonists and antagonists) of unrelated GPCRs for their ability to induce intracellular Ca2+mobilization in human neutrophils and HL-60 cells transfected with human FPR1, FPR2, or FPR3. Screening of these compounds demonstrated that antagonists of gastrin-releasing peptide/neuromedin B receptors (BB1/BB2) PD168368 [(S)-a-methyl-a-[[[(4-nitrophenyl)amino]carbonyl]amino]-N-[[1-(2-pyridinyl) cyclohexyl]methyl]-1H-indole-3-propanamide] and PD176252 [(S)-N-[[1-(5-methoxy-2-pyridinyl)cyclohexyl]methyl]-a-methyl-a-[[-(4-nitrophenyl)amino]carbonyl]amino-1H-indole-3-propanamide] were potent mixed FPR1/FPR2 agonists, with nanomolar EC50values. Cholecystokinin-1 receptor agonist A-71623 [Boc-Trp-Lys(-N-2-methylphenylaminocarbonyl)-Asp-(N-methyl)-Phe-NH2] was also a mixed FPR1/FPR2 agonist, but with a micromolar EC50. Screening of 56 Trp- and Phe-based PD176252/PD168368 analogs and 41 related nonpeptide/nonpeptoid analogs revealed 22 additional FPR agonists. Most were potent mixed FPR1/FPR2/FPR3 agonists with nanomolar EC50values for FPR2, making them among the most potent nonpeptide FPR2 agonists reported to date. In addition, these agonists were also potent chemoattractants for murine and human neutrophils and activated reactive oxygen species production in human neutrophils. Molecular modeling of the selected agonists using field point methods allowed us to modify our previously reported pharmacophore model for the FPR2 ligand binding site. This model suggests the existence of three hydrophobic/aromatic subpockets and several binding poses of FPR2 agonists in the transmembrane region of this receptor. These studies demonstrate that FPR agonists could include ligands of unrelated GPCR and that analysis of such compounds can enhance our understanding of pharmacological effects of these ligands.

N-formyl peptide receptor (FPR1) and N-formyl peptide receptor-like 1 (FPRL1, now known as FPR2) are G protein-coupled receptors involved in host defense and sensing cellular dysfunction. Because of the potential for FPR1/FPR2 as a therapeutic target, our recent high-throughput screening efforts have focused on the identification of unique nonpeptide agonists of FPR1/FPR2. In the present studies, we screened a chemolibrary of drug-like molecules for their ability to induce intracellular calcium mobilization in RBL-2H3 cells transfected with human FPR1 or FPR2. Screening of these compounds resulted in the identification of novel and potent agonists that activated both FPR1 and FPR2, as well as compounds that were specific for either FPR1 or FPR2 with EC50values in the low micromolar range. Specificity of the compounds was supported by analysis of calcium mobilization in HL-60 cells transfected with human FPR1 and FPR2. In addition, all but one agonist activated intracellular calcium flux and chemotaxis in human neutrophils, irrespective of agonist specificity for FPR1 or FPR2. Molecular modeling of the group of FPR1 and FPR2 agonists using field point methodology allowed us to create pharmacophore models for ligand binding sites and formulate requirements for these specific N-formyl peptide receptor agonists. These studies further demonstrate that agonists of FPR1/FPR2 include compounds with wide chemical diversity and that analysis of such compounds can enhance our understanding of their ligand/receptor interaction.

The chemokine receptors CXCR1/2 are involved in a variety of inflammatory diseases, including chronic obstructive pulmonary disease. Several classes of allosteric small-molecule CXCR1/2 antagonists have been developed. The data presented here describe the cellular pharmacology of the acid and ester forms of the nicotinamide glycolate pharmacophore, a potent antagonist of CXCR2 signaling by the chemokines CXCL1 and CXCL8. Ester forms of the nicotinamide glycolate antagonized CXCL1-stimulated chemotaxis (IC(50) = 42 nM) and calcium flux (IC(50) = 48 nM) in human neutrophils, but they were inactive in cell-free assays of (125)I-CXCL8/CXCR2 binding and CXCL1-stimulated guanosine 5'-O-(3-[(35)S]thio)triphosphate ([(35)S]GTPgammaS) exchange. Acid forms of the nicotinamide glycolate were inactive in whole-cell assays of chemotaxis and calcium flux, but they inhibited (125)I-CXCL8/CXCR2 binding and CXCL1-stimulated [(35)S]GTPgammaS exchange. The (3)H ester was internalized by neutrophils and rapidly converted to the (3)H acid in a concentrative process. The (3)H acid was not internalized by neutrophils but was sufficient alone to inhibit CXCL1-stimulated calcium flux in neutrophils that were permeabilized by electroporation to permit its direct access to the cell interior. Neutrophil efflux of the acid was probenecid-sensitive, consistent with an organic acid transporter. These data support a mechanism wherein the nicotinamide glycolate ester serves as a lipophilic precursor that efficiently translocates into the intracellular neutrophil space to liberate the active acid form of the pharmacophore, which then acts at an intracellular site. Rapid inactivation by plasma esterases precluded use in vivo, but the mechanism elucidated provided insight for new nicotinamide pharmacophore classes with therapeutic potential.

AbstractMacrophages play essential roles in a wide variety of physiological and pathological processes. One of the unique features of these phagocytic leukocytes is their ability to fuse, forming multinucleated giant cells. Multinucleated giant cells are important mediators of tissue remodeling and repair and are also responsible for removal or sequestration of foreign material, intracellular bacteria and non-phagocytosable pathogens, such as parasites and fungi. Depending on the tissue where fusion occurs and the inflammatory insult, multinucleated giant cells assume distinctly different phenotypes. Nevertheless, the ultimate outcome is the formation of large cells that can resorb bone tissue (osteoclasts) or foreign material and pathogens (giant cells) extracellularly. While progress has been made in recent years, the mechanisms and factors involved in macrophage fusion are still not fully understood. In addition to cytokines and a number of adhesion proteins and receptors, it is becoming increasingly clear that NADPH oxidase-generated reactive oxygen species (ROS) also play an important role in macrophage fusion. In this review, we provide an overview of macrophage multinucleation, with a specific focus on the role of NADPH oxidases and ROS in macrophage fusion and in the function of multinucleated giant cells. In addition, we provide an updated overview of the role of these cells in inflammation and various autoimmune diseases.Copyright © 2009 S. Karger AG, Basel

ObjectiveTo compare work disability and job loss in early rheumatoid arthritis RA patients receiving adalimumab plus methotrexate adalimumab MTX versus MTX alone.MethodsIn this multicenter, randomized, controlled trial, patients with RA for <2 years who had never taken MTX and who selfreported work impairment were randomized to adalimumab MTX or placebo MTX for 56 weeks. Primary outcome was job loss of any cause andor imminent job loss at or after week 16. Secondary outcomes included disease activity, function Health Assessment Questionnaire HAQ score, and RA quality of life RAQoL questionnaire score. Work was evaluated with work diaries and the RA Work Instability Scale.ResultsAlthough job loss during the 56week study was significantly lower with adalimumab MTX 14 of 75 patients compared with MTX alone 29 of 73 patients; P 0.005, the primary end point was not met 12 of 75 versus 20 of 73 patients; P 0.092, likely owing to early drop out in the MTX group. There were significant improvements in American College of Rheumatology 20 response criteria, 28joint Disease Activity Score, ΔHAQ, ΔRAQoL, and working time lost in the adalimumab MTX group. Twentyfour serious adverse events were reported in 17 participants, with no differences between groups.ConclusionAdalimumab MTX reduced job loss and improved productivity in early RA when compared with MTX alone, which supports the early use of anti–tumor necrosis factor therapy and suggests its cost efficacy.

Development of immunomodulatory agents that enhance innate immune responses represents a promising strategy for combating infectious diseases. In the present studies, we screened a series of 71 arylcarboxylic acid hydrazide derivatives for their ability to induce macrophage tumor necrosis factor α (TNF-α) production and identified six such compounds, including one compound previously shown to be a formyl peptide receptor (FPR/FPRL1) agonist. The two most potent compounds [compound 1, nicotinic acid [5-(3-bromophenyl)-2-furyl]methylene-hydrazide; compound 2, 4-fluoro-benzoic acid [5-(3-trifluoromethyl-phenyl)-2-furyl]-methylene-hydrazide] were selected for further analysis. These compounds induced de novo production of TNF-α in a dose- and time-dependent manner in human and murine monocyte/macrophage cell lines and in primary macrophages. These compounds also induced mobilization of intracellular Ca2+, production of reactive oxygen species, and chemotaxis in human and murine phagocytes. Induction of macrophage TNF-α production was pertussis toxin-sensitive, and analysis of the cellular target of these compounds showed that they were FPRL1-specific agonists and that this response was blocked by FPR/FPRL1 and FPRL1-specific antagonists. In addition, pharmacophore modeling showed a high degree of similarity for low-energy conformations of these two compounds to the current pharmacophore model for FPR ligands ( Mol Pharmacol 68: 1301-1310, 2005 ). Overall, these compounds represent novel FPRL1 agonists that induce TNF-α, a response distinct from those induced by other known FPR and FPRL1 agonists.

Recruitment of phagocytes to inflammatory sites involves the coordinated action of several chemoattractants, including the anaphylatoxin C5a. While the C5a receptor (C5aR) has been well characterized in humans and rodents, little is known about the bovine C5aR. Here, we report cloning of bovine C5R1, the gene encoding bovine C5aR. We also analyzed genomic sequence upstream of the C5R1translation start site. Although the bovine C5aR amino acid sequence was well conserved among species, significant differences in conserved features were found, including major differences in the N terminus, intracellular loop 3, and transmembrane domain VII. Analysis of C5aR expression by flow cytometry and confocal microscopy demonstrated high levels of C5aR on all bovine neutrophils and a subset of bovine monocytes. C5aR was not expressed on resting or activated bovine lymphocytes, although C5aR message was present in these cells. C5aR was also expressed on a small subset of bovine mammary epithelial cells. Pharmacological analysis of bovine C5aR‐mediated responses showed that bovine C5a and C5adesArgboth induced dose‐dependent calcium fluxes and chemotaxis in bovine neutrophils, with similar efficacy for both agonists. Treatment of bovine neutrophils with C5a or C5adesArgresulted in homologous desensitization of bovine C5aR and cross‐desensitization to interleukin 8 (IL‐8) and platelet‐activating factor (PAF); whereas, treatment with IL‐8 or PAF did not cross‐desensitize the cells to C5a or C5adesArg. Overall, these studies provide important information regarding distinct structural and functional features that may contribute to the unique pharmacological properties of bovine C5aR.

No abstract.

Macrophages play an important role in the pathogenesis of chronic inflammatory disease. Activation of these phagocytes induces the production of proinflammatory cytokines, such as IL‐1 and TNF‐α and the generation of reactive oxygen species (ROS), such as superoxide anion (O2•–). Recently, we found that TNF‐α treatment of human monocytic cells (MonoMac1) and isolated human monocytes resulted in up‐regulation of the NADPH oxidase gene, neutrophil cytosolic factor 2 (NCF2). These results suggested that TNF‐α, produced by activated macrophages, could serve as an autocrine/paracrine regulator of the oxidase, resulting in increased and/or prolonged production of O2•–. To gain a better understanding of the mechanisms involved in NADPH oxidase regulation by TNF‐α, we evaluated transcriptional regulation of oxidase genes in MonoMac1 cells and human monocytes. We show that TNF‐α‐treated cells have increased levels of mRNA and up‐regulated expression of NADPH oxidase subunits p47phox, p67phox, and gp91phox, as well as increased oxidase activity. Pharmacological inhibitors of NF‐κB activation blocked TNF‐α‐induced up‐regulation of NCF1, NCF2, and CYBBmessage, which correlated with a reduction in expression of the corresponding oxidase proteins and decreased O2•–production. These data demonstrate that the increase in and/or maintenance of O2•–production in TNF‐α‐treated MonoMac1 cells and monocytes are a result, in part, of transcriptional up‐regulation of three essential NADPH oxidase genes via the NF‐κB pathway. This novel finding supports a model, whereby TNF‐α‐dependent activation of NF‐κB up‐regulates phagocyte NADPH oxidase activity, leading to enhanced ROS production and further NF‐κB activation, potentially contributing to sustained oxidant production in chronic inflammation.

The role of alternative tumor necrosis factor (TNF) antagonist therapies in the context of failure of initial TNF antagonist therapy in patients with rheumatoid arthritis (RA) has yet to be clearly defined. The goal of this study was to determine the efficacy of etanercept in patients who failed to respond to infliximab.Ninety‐five patients with RA who failed to respond to infliximab and methotrexate were treated with etanercept (with continuation of concomitant methotrexate). Thirty‐four patients never achieved a response to infliximab (primary nonresponse), 38 had an initial response to infliximab but relapsed (secondary nonresponse), and 23 demonstrated toxicity. Disease Activity Score in 28 joints (DAS28), European League Against Rheumatism (EULAR) response, and American College of Rheumatology (ACR) response were determined after 12 weeks of etanercept.After 12 weeks of etanercept, 38% of patients achieved an ACR 20% response (ACR20) on etanercept. Of these, 24% and 15% achieved ACR50 and ACR70 responses, respectively. In the primary infliximab nonresponse group, 42%, 30%, and 15% achieved ACR20, ACR50, and ACR70 responses, respectively; the percentages for the secondary nonresponse group were 34%, 21%, and 14%, respectively. Significant DAS28 reductions were observed in the entire cohort and nonresponse subtype groups. Sixty‐one percent of the cohort achieved either a moderate or good EULAR score (67% of primary and 56% of secondary infliximab failures). No toxicity was observed in patients who stopped infliximab due to intolerance; 19 of 23 continued etanercept after week 12.This study confirms that etanercept is effective in patients who fail to respond to infliximab and suggests a higher response in patients who have never had a response to infliximab.

We screened a chemolibrary of drug-like molecules for their ability to activate reactive oxygen species (ROS) production in murine phagocytes, and we identified 26 novel compounds with potent neutrophil activating properties. We used substructure screening, fragment-focusing, and structure-activity relationship analyses to further probe the parent library and defined at least two groups of activators of ROS production in murine neutrophils: t-butyl benzene and thiophene-2-amide-3-carboxylic ester derivatives. Further studies of the active compounds revealed 11 compounds that activated ROS production in human neutrophils, and six of these compounds also activated intercellular Ca2+mobilization and chemotaxis in human neutrophils. Of the latter compounds, compound 14 (1,3-benzodioxolane-5-carboxylic acid 4'-benzyloxy-3'-methoxybenzylidene-hydrazide) activated neutrophils at nanomolar concentrations, and Ca2+mobilization was inhibited by pertussis toxin and N-t-butoxycarbonyl-Phe-Leu-Phe-Leu-Phe (Boc-2), an antagonist of formyl peptide receptors (FPR/FPRL1). Likewise, activation by compound 14 was desensitized after N-formyl-Met-Leu-Phe pretreatment. Similar biological activities were found for compound 104 (1,3-benzodioxolane-5-carboxylic acid 3'-bromo-5'-ethoxy-4'-hydroxybenzylidene-hydrazide), an analog of compound 14. Furthermore, conformational analysis of the activators of chemotaxis and Ca2+mobilization showed a high degree of similarity in distances between pharmacophore points of compounds 14 and 104 with a model of FPR published by Edwards et al. (Mol Pharmacol 68:1301–1310, 2005), indicating that conformational features of the agonists identified here are structurally compatible with steric constraints of the ligand-binding pocket of the receptor. Based on these results, we conclude that compounds 14 and 104 represent novel small-molecule agonists of FPR. These studies enhance our understanding of FPR ligand/receptor interactions and structure/activity relationships of phagocyte agonists.

Anthrax lethal factor (LF) is a key virulence factor of anthrax lethal toxin. We screened a chemolibrary of 10 000 drug-like molecules for their ability to inhibit LF and identified 18 novel small molecules with potent LF inhibitory activity. Three additional LF inhibitors were identified through further structure−activity relationship (SAR) analysis. All 21 compounds inhibited LF with an IC50range of 0.8 to 11 M, utilizing mixed-mode competitive inhibition. An evaluation of inhibitory activity against a range of unrelated proteases showed relatively high specificity for LF. Furthermore, pharmacophore modeling of these compounds showed a high degree of similarity to the model published by Panchal et al. (Nat. Struct. Mol. Biol.2004, 11, 67−72), indicating that the conformational features of these inhibitors are structurally compatible with the steric constraints of the substrate-binding pocket. These novel LF inhibitors and the structural scaffolds identified as important for inhibitory activity represent promising leads to pursue for further LF inhibitor development.

The NADPH oxidase was originally identified as a key component of human innate host defence. In phagocytes, this enzyme complex is activated to produce superoxide anion and other secondarily derived ROS (reactive oxygen species), which promote killing of invading micro-organisms. However, it is now well-established that NADPH oxidase and related enzymes also participate in important cellular processes not directly related to host defence, including signal transduction, cell proliferation and apoptosis. These enzymes are present in essentially every organ system in the body and contribute to a multitude of physiological events. Although essential for human health, excess NADPH-oxidase-generated ROS can promote numerous pathological conditions. Herein, we summarize our current understanding of NADPH oxidases and provide an overview of how they contribute to specific human diseases.

AbstractThis study examined the effectiveness of the Parents Plus Programme for families of pre-school children with significant behavioural problems, comparing those with and without developmental disabilities. Twenty-two parents of children with developmental disabilities and conduct problems (the disability group), and 17 parents of children with conduct problems, but without developmental disabilities (the conduct problems group), were assessed before and after participating in the Parents Plus Programme, and at 10 months follow-up. More than 70% of cases in both the disability and conduct problems groups showed clinically significant improvement on the Total Difficulties scale of the Strengths and Difficulties Questionnaire. During the follow-up period, parents in the disability group showed a deterioration in psychological adjustment (on the General Health Questionnaire-12), while parents in the conduct problems group showed an improvement. Parents in the disability group reported a higher level of goal attainment compared with parents in the conduct problem group. Parents in both the disability and conduct problems groups evaluated the Parents Plus Programme equally positively. The Parents Plus Programme requires refinement to become more effective for families of pre-school children with developmental disabilities.

The phagocyte nicotinamide adenine dinucleotide phosphate (NADPH) oxidase plays an instrumental role in host defense and contributes to microbicial killing by releasing highly reactive oxygen species. This multicomponent enzyme is composed of membrane and cytosolic components that assemble in the plasma membrane or phagolysosome. While the guanosine S′-triphosphatase (GTPase) Rac2 has been shown to be a critical regulator of NADPH oxidase activity and assembly, the role of its effector, p21-activated kinase (Pak), in oxidase function has not been well defined. Using HIV-1 Tat-mediated protein transduction of Pak inhibitory domain, we show here that Pak activity is indeed required for efficient superoxide generation in intact neutrophils. Furthermore, we show that Pak translocates to the plasma membrane upon N-formyl-methionyl-leucyl-phenylalanine (fMLF) stimulation and colocalizes with translocated p47phoxand with p22phox,a subunit of flavocytochrome b558. Although activated Pak phosphorylated several essential serine residues in the C-terminus of p47phox,direct binding to p47phoxwas not observed. In contrast, active Pak bound directly to p22phox,suggesting flavocytochrome b was the oxidase-associated membrane target of this kinase and this association may facilitate further phosphorylation of p47phoxin the assembling NADPH oxidase complex.

The phagocyte reduced nicotinamide adenine dinucleotide phosphate oxidase is a multiprotein enzyme that catalyzes the production of microbicidal oxidants. Although oxidase assembly involves association of several membrane and cytosolic oxidase proteins, one of the cytosolic cofactors, p67phox, appears to play a more prominent role in final activation of the enzyme complex. Based on the importance of p67phox, we investigated transcriptional regulation of the p67phoxgene [neutrophil cytosolic factor 2 (NCF2)] and demonstrated previously that activator protein‐1 (AP‐1) was essential for basal transcriptional activity. As p67phoxcan be up‐regulated by tumor necrosis factor α (TNF‐α), which activates AP‐1, we hypothesized that TNF‐α might regulate NCF2transcription via AP‐1. In support of this hypothesis, we show here that NCF2promoter‐reporter constructs are up‐regulated by TNF‐α but only when AP‐1 factors were coexpressed. Consistent with this observation, we also demonstrate that NCF2mRNA and p67phoxprotein are up‐regulated by TNF‐α in various myeloid cell lines as well as in human monocytes. It was surprising that mutagenesis of the AP‐1 site in NCF2promoter constructs did not eliminate TNF‐α induction, suggesting additional elements were involved in this response and that AP‐1 might play a more indirect role. Indeed, we used NCF2promoter‐deletion constructs to map a novel TNF‐α‐responsive region (TRR) located between −56 and −16 bp upstream of the translational start site and demonstrated its importance in vivo using transcription factor decoy analysis. Furthermore, DNase footprinting verified specific binding of factor(s) to the TRR with AP‐1 binding indirectly to this region. Thus, we have identified a novel NCF2promoter/enhancer domain, which is essential for TNF‐α‐induced up‐regulation of p67phox.

We previously demonstrated prolonged, profound CD4+ T-lymphopenia in rheumatoid arthritis (RA) patients following lymphocyte-depleting therapy. Poor reconstitution could result either from reduced de novo T-cell production through the thymus or from poor peripheral expansion of residual T-cells. Interleukin-7 (IL-7) is known to stimulate the thymus to produce new T-cells and to allow circulating mature T-cells to expand, thereby playing a critical role in T-cell homeostasis. In the present study we demonstrated reduced levels of circulating IL-7 in a cross-section of RA patients. IL-7 production by bone marrow stromal cell cultures was also compromised in RA. To investigate whether such an IL-7 deficiency could account for the prolonged lymphopenia observed in RA following therapeutic lymphodepletion, we compared RA patients and patients with solid cancers treated with high-dose chemotherapy and autologous progenitor cell rescue. Chemotherapy rendered all patients similarly lymphopenic, but this was sustained in RA patients at 12 months, as compared with the reconstitution that occurred in cancer patients by 3–4 months. Both cohorts produced naïve T-cells containing T-cell receptor excision circles. The main distinguishing feature between the groups was a failure to expand peripheral T-cells in RA, particularly memory cells during the first 3 months after treatment. Most importantly, there was no increase in serum IL-7 levels in RA, as compared with a fourfold rise in non-RA control individuals at the time of lymphopenia. Our data therefore suggest that RA patients are relatively IL-7 deficient and that this deficiency is likely to be an important contributing factor to poor early T-cell reconstitution in RA following therapeutic lymphodepletion. Furthermore, in RA patients with stable, well controlled disease, IL-7 levels were positively correlated with the T-cell receptor excision circle content of CD4+ T-cells, demonstrating a direct effect of IL-7 on thymic activity in this cohort.