Environmental drivers of breeding sites in blackfly species of medical and veterinary importance in eastern Spain.

Geographical distribution and abundance of the pupae of six blackfly species of medical and veterinary concern were studied in eastern Spain according to three different sets of explanatory variables including in-stream variables, both (i) abiotic (i.e., physicochemical) and (ii) biotic (i.e., richness and abundance of either taxonomically or ecologically close related taxa), as well as (iii) meteorological and landscape variables. The results showed specific habitat requirements for pupation in Simulium (Boophthora) erythrocephalum (De Geer, 1776) and Simulium (Wilhelmia) equinum (Linnaeus, 1758), two of the six species studied regarding elevation and temperature. While the rest of the species showed a certain degree of ecological overlap, co-occurrence was in general low, which suggested that antagonistic biotic factors may be important in structuring blackfly assemblages. In effect, biotic predictors explained a high proportion (50%-70%) of the variability in the abundance of the pupae of the most generalist blackfly species, although further studies are needed to disentangle the sign of interspecific interactions. At the landscape level, S. (W.) equinum and S. (W.) pseudequinum Séguy, 1921 breeding habitats were associated with the presence of pig farms, and S. (Simulium) reptans (Linnaeus, 1758) and S. (B.) erythrocephalum with the presence of cattle.

Mosquito Magnet® traps as a potential means of monitoring blackflies of medical and veterinary importance.

Mosquito Magnet® traps, deployed in widespread parts of England as part of nationwide mosquito surveillance projects, also caught blackflies. As many as 1242 blackflies were caught in a trapping session lasting 4 days. Principal among the species caught were Simulium equinum, Simulium lineatum and Simulium ornatum s.l. As S. ornatum s.l. is a vector that transmits Onchocerca linealis to cattle and S. equinum is responsible for dermatitis ('sweet itch') in cattle and horses, it is suggested that Mosquito Magnet® traps could be used to monitor and partially control these pests, as well as nuisance anthropophilic blackflies such as Simulium posticatum that can cause simuliidosis in southern England.

MAG-EPA resolves lung inflammation in an allergic model of asthma.

BACKGROUND: Asthma is a chronic disease characterized by airways hyperresponsiveness, inflammation and airways remodelling involving reversible bronchial obstruction. Omega-3 fatty acids and their derivatives are known to reduce inflammation in several tissues including lung. OBJECTIVES: The effects of eicosapentaenoic acid monoacylglyceride (MAG-EPA), a newly synthesized EPA derivative, were determined on the resolution of lung inflammation and airway hyperresponsiveness in an in vivo model of allergic asthma. METHODS: Ovalbumin (OVA)-sensitized guinea-pigs were treated or not with MAG-EPA administered per os. Isometric tension measurements, histological analyses, homogenate preparation for Western blot experiments or total RNA extraction for RT-PCR were performed to assess the effect of MAG-EPA treatments. RESULTS: Mechanical tension measurements revealed that oral MAG-EPA treatments reduced methacholine (MCh)-induced bronchial hyperresponsiveness in OVA-sensitized guinea-pigs. Moreover, MAG-EPA treatments also decreased Ca(2+) hypersensitivity of bronchial smooth muscle. Histological analyses and leucocyte counts in bronchoalveolar lavages revealed that oral MAG-EPA treatments led to less inflammatory cell recruitment in the lung of OVA-sensitized guinea-pigs when compared with lungs from control animals. Results also revealed a reduction in mucin production and MUC5AC expression level in OVA-sensitized animals treated with MAG-EPA. Following MAG-EPA treatments, the transcript levels of pro-inflammatory markers such as IL-5, eotaxin, IL-13 and IL-4 were markedly reduced. Moreover, per os MAG-EPA administrations reduced COX2 over-expression in OVA-sensitized animals. CONCLUSION AND CLINICAL RELEVANCE: We demonstrate that MAG-EPA reduces airway hyperresponsiveness and lung inflammation in OVA-sensitized animals, a finding consistent with a decrease in IL-4, IL-5, IL-13, COX-2 and MUC5AC expression levels in the lung. The present data suggest that MAG-EPA represents a new potential therapeutic strategy for resolving inflammation in allergic asthma.

Severe anaphylaxis caused by orally administered vancomycin to a patient with Clostridium difficile infection.

We report the first case of anaphylaxis to oral vancomycin in a cystic fibrosis patient with severe and relapsing Clostridium difficile infection (CDI) refractory to metronidazole. The patient's colitis has been successfully treated with a combination of intravenous metronidazole and tigecycline.

Cigarette smoke-induced proteostasis imbalance in obstructive lung diseases.

The airway and alveolar surface is exposed daily to 8,000 L of air containing oxygen, particles, bacteria, allergens and pollutants, all of which have the potential to induce oxidative stress within cells. If one is also a cigarette smoker, then the exposure to reactive oxidants increases exponentially. More than any other tissue, the lung is at risk of undergoing oxidative changes in protein expression, structure and function. The oxidant burden of chronic cigarette smoke exposure can overwhelm the lung cells' capacity to maintain proteostasis, a process of regulated protein synthesis, folding and turnover. Somewhat surprisingly, most chronic cigarette smokers do not develop chronic obstructive pulmonary disease (COPD), likely because cells initiate a highly effective unfolded protein response (UPR) in the presence of oxidant-derived endoplasmic reticulum (ER) stress that allows cells to survive. The UPR initiates several signaling pathways that decrease protein translation, limit cell cycle progression, increase protein degradation and chaperone-mediated protein folding, and activate the transcription factor Nrf2 that induces antioxidant gene expression. Each of these actions decreases ER stress in a process of "healthy proteostasis". If these responses are insufficient, apoptosis ensues. In this article, we review the mechanisms of healthy and dysfunctional proteostasis related to cigarette smoke exposure and COPD.

Neutrophil-mediated epithelial injury during transmigration: role of elastase.

Neutrophil-mediated injury to gut epithelium may lead to disruption of the epithelial barrier function with consequent organ dysfunction, but the mechanisms of this are incompletely characterized. Because the epithelial apical junctional complex, comprised of tight and adherens junctions, is responsible in part for this barrier function, we investigated the effects of neutrophil transmigration on these structures. Using a colonic epithelial cell line, we observed that neutrophils migrating across cell monolayers formed clusters that were associated with focal epithelial cell loss and the creation of circular defects within the monolayer. The loss of epithelial cells was partly attributable to neutrophil-derived proteases, likely elastase, because it was prevented by elastase inhibitors. Spatially delimited disruption of epithelial junctional complexes with focal loss of E-cadherin, beta-catenin, and zonula occludens 1 was observed adjacent to clusters of transmigrating neutrophils. During neutrophil transmigration, fragments of E-cadherin were released into the apical supernatant, and inhibitors of neutrophil elastase prevented this proteolytic degradation. Addition of purified leukocyte elastase also resulted in release of E-cadherin fragments, but only after opening of tight junctions. Taken together, these data demonstrate that neutrophil-derived proteases can mediate spatially delimited disruption of epithelial apical junctions during transmigration. These processes may contribute to epithelial loss and disruption of epithelial barrier function in inflammatory diseases.

In vitro generation of peroxynitrite by 2- and 4-hydroxyestrogens in the presence of nitric oxide.

Estrogen metabolism is altered in most, if not all, breast cancer tumors. These alterations primarily lead to the formation of the catechol estrogen metabolites, 2- and 4-hydroxyestrogens, which can generate superoxide anion radicals (O(2)(*)(-)) through the redox cycling of semiquinone/quinone derivatives. In breast cancer cells, the activity of nitric oxide synthase is also frequently elevated, resulting in an increased level of exposure to nitric oxide ((*)NO). Since (*)NO rapidly reacts with O(2)(*)(-) to produce the peroxynitrite anion (ONOO(-)), this study was undertaken to determine whether ONOO(-) can be generated when 2- and 4-hydroxyestrogens are incubated in vitro with (*)NO donor compounds. Using dihydrorhodamine 123 as a specific probe for ONOO(-) formation, a ratio of 100 microM dipropylenetriamine NONOate (DPTA/NO) to 10 microM 4-hydroxyestradiol (4-OHE(2)) gave an optimal ONOO(-) production of 11.9 +/- 1.9 microM (mean +/- SD). Quantification of ONOO(-) was not modified by mannitol, supporting the idea that the hydroxyl radical was not involved. This production of ONOO(-) required the presence of the catechol structure of estrogen metabolites since all methoxyestrogens that were tested were inactive. Hydroxyestrogen metabolites derived from estradiol showed the same efficiency in producing ONOO(-) as those originating from estrone. With DPTA/NO, the 4-hydroxyestrogens generated 30-40% more ONOO(-) than the 2-hydroxyestrogens. Optimal production of ONOO(-) was assessed with DPTA/NO and diethylenetriamine NONOate (initial (*)NO generation rates of 0.76 and 0.08 microM min(-1), respectively). With faster (*)NO-releasing compounds, such as diethylamine NONOate and spermine NONOate, lower levels of ONOO(-) were detected. These data suggest that once the optimal concentration of (*)NO was obtained, the reaction between (*)NO and 4-OHE(2) was saturated. The excess of (*)NO would probably react with aqueous oxygen to form nitrite (NO(2)(-)). Since the third-order reaction rate for the reaction between 2(*)NO and O(2) is 2 x 10(6) M(-2) s(-1), it can therefore be suggested that the reaction between (*)NO and 4-OHE(2) occurs at a faster rate.

Albumin-mediated regulation of cellular glutathione and nuclear factor kappa B activation.

Human serum albumin (HSA) is a cystine-rich serum protein taken up by many cells through receptor-mediated and fluid-phase endocytosis. We hypothesized that HSA may play a role in modulating cellular antioxidant redox signaling. Lung epithelial cells (A549), fibroblasts (HFL1), and blood lymphocytes had increased glutathione (GSH) levels after 8 h incubation with HSA. Similar GSH increases were observed with either plasma-derived or recombinant HSA. Serum depleted of HSA had no effect on cellular GSH. The GSH increase was also observed in normal murine lungs upon in vivo airway instillation of HSA. GSH enhancement was not related to the redox state of the free cysteine residue (Cys-34) on HSA, however, reduction of disulfide bonds in HSA inhibited the increase in cellular GSH. In addition, the albumin-mediated increase in GSH was inhibited by the vacuolar (H(+))-ATPase inhibitors, bafilomycin A(1) and concanamycin, as well as by the membrane pH-disrupting ionophore monensin, but not by 20 mM NH(4)Cl. The degree to which albumin increased GSH levels was sufficient to protect cells against H(2)O(2)-mediated cytotoxicity and to decrease TNF-alpha-mediated NF-kappaB activation. We conclude that albumin specifically modulates cellular GSH levels, an effect sufficient to protect cells against oxidant injury and regulate NF-kappaB activation.

Synthesis and biological evaluation of new analogues of the active fungal metabolites N-(2-methyl-3-oxodecanoyl)-2-pyrroline and N-(2-methyl-3-oxodec-8-enoyl)-2-pyrroline (II).

New analogues of the bioactive enamides isolated from P. brevicompactum (2 and 3) have been synthesized to improve the biological activities. Two different structural modifications have been introduced: substitution of the aliphatic side chain present in the natural products (1-4) by other groups frequently found in other active compounds and use of other nitrogen-containing five-membered rings with different degrees of oxidation. In this way, the insecticidal and fungicidal activities have been improved. Thus, compound 9, which possess a 3-pyrroline ring, exhibited important insecticidal activity against third-instar nymphs of Oncopeltus fasciatus Dallas (100% mortality at 7.5 microg/cm(2)). Remarkable fungicidal activity was also found, and preliminary structure-activity relationships could be established.

Oestrogen metabolism in lymphangioleiomyomatosis: catechol-O-methyltransferase pathway is not involved.

BACKGROUND: Lymphangioleiomyomatosis (LAM) is an uncommon lung disease for which no effective method of treatment has been found. The predilection of LAM for premenopausal women has led to the assumption that hormonal factors play an important role in the pathogenesis of this disease. The aim of this study was to determine if women with LAM manifest alterations in the catechol-O-methyltransferase (COMT) pathway which is essential for preventing the generation of oestrogen derived reactive oxygen species (ROS). METHODS: Blood samples were collected from 15 women with LAM and compared with appropriate controls. The distribution of high and low activity alleles of COMT was determined with a PCR based RFLP assay. The enzymatic activity of COMT was measured in each sample and the potential presence of a circulating inhibitor of COMT was determined. Since an alteration in the COMT pathway could increase the oxidative stress, the plasma concentration of malondialdehyde (MDA), a secondary product generated from lipid peroxidation, has been used as an internal marker. RESULTS: The distribution of high and low activity alleles of COMT (named COMT(HH), COMT(LL), and COMT(HL)) was similar in the two groups with proportions of 40%, 7%, and 53%, respectively, in the women with LAM and 38%, 6%, and 56% in the control subjects. The mean (SD) COMT activity was 24.2 (12.3) pmol/min/mg protein in women with LAM and 24.1 (6.3) pmol/min/mg protein in the control group. Incubation of plasma from women in the two groups with a preparation of commercial COMT showed that no detectable COMT inhibitor was present. The plasma concentration of MDA in the women with LAM was also not significantly different from control subjects. CONCLUSIONS: This study shows that there are no significant alterations in the COMT pathway of women with LAM. It is therefore unlikely that alterations in oestrogen mediated cell signalling pathways are mediated by oxidants derived from an excess of catecholoestrogens in LAM.

Direct correlation of glutathione and ascorbate and their dependence on age and season in human lymphocytes.

BACKGROUND: Endogenous reactive oxygen species appear to contribute to aging and cancer and dietary antioxidants, present in fruit and vegetables, counteract these effects. OBJECTIVE: The objective was to examine the association between intracellular glutathione, ascorbate (vitamin C), and alpha-tocopherol (vitamin E) in human lymphocytes. DESIGN: The study group consisted of 240 healthy nonsmoking volunteers with an approximately equal number of male and female subjects subdivided into 3 age groups: 18-39, 40-59, and >/=60 y). Glutathione, glutathione disulfide, ascorbate, and alpha-tocopherol were measured in lymphocytes by HPLC. RESULTS: The average concentration of antioxidants in lymphocytes was 27 +/- 8 nmol/mg protein for glutathione, 21 +/- 8 nmol/mg protein for ascorbate, and 0.4 +/- 0.2 nmol/mg protein for alpha-tocopherol. There was a strong positive correlation between glutathione and ascorbate (r = 0.62, P < 0.001). No correlation was observed for glutathione and ascorbate with alpha-tocopherol. The concentration of glutathione in lymphocytes was inversely correlated with age (r = -0.19, P < 0.01), as was that of ascorbate (r = -0.22, P < 0.01), with 10-20% lower values in elderly than in young and elderly subjects. The concentrations of glutathione in lymphocytes were as much as 25% higher and those of ascorbate were as much as 38% higher during the summer than during the winter. The seasonal variation of ascorbate in lymphocytes was described by a linear function for age and a periodic sine function for season. CONCLUSION: Glutathione and ascorbate are directly correlated in human lymphocytes.

Insecticidal, anti-juvenile hormone, and fungicidal activities of organic extracts from different Penicillium species and their isolated active components.

Organic extracts from mycelium and culture broth of 21 Penicillium isolates have been tested for insecticidal, insect anti-juvenile hormone (anti-JH), and antifungal activities. Culture broth extracts were the most active, mainly against insects; nearly 25% of them have shown high entomotoxicity (100% mortality at 100 microg/cm(2)). A strong in vivo anti-JH activity against Oncopeltus fasciatus Dallas was detected in the culture broth extracts from P. brevicompactum P79 and P88 isolates. The two new natural products isolated from P79, N-(2-methyl-3-oxodec-8-enoyl)-2-pyrroline (1) and 2-hept-5-enyl-3-methyl-4-oxo-6,7,8,8a-tetrahydro-4H-pyrrolo[2,1-b]-1, 3-oxazine (2), possessed anti-JH and insecticidal activity, respectively, against O. fasciatus. Synthesized natural compound 1 has shown an ED(50) of 0.7 microg/nymph when assayed on newly molted fourth-instar nymphs of O. fasciatus. Promising biological activities have also been detected in the synthetic precursors.

Synthesis and biological evaluation of new analogues of the active fungal metabolites N-(2-methyl-3-oxodecanoyl)-2-pyrroline and N-(2-methyl-3-oxodec-8-enoyl)-2-pyrroline.

To evaluate the effect of simplifying the beta-ketoamide system present in active isolated metabolites from Penicillium brevicompactum (2 and 3) on the activity, new analogues with a monocarbonylic amide functionality have been obtained. In this way, the insecticidal and fungicidal activities have been improved in relation to the natural products taken as lead molecules. Thus, two of the synthetic analogues (5a and 5b) showed very important insecticidal activities against third-instar nymphs of Oncopeltus fasciatus Dallas, with acute LD(50) values of 3.0 and 1.5 microg/cm(2), respectively. Moreover, some analogues showed good levels of fungicidal activity against a wide range of commercially important and taxonomically diverse fungi; remarkably, compound 7c has proved to be highly active against Colletotrichum gloesporoides and Colletotrichum coccodes, with ED(50) values of 2.04 and 11.7 microg/mL, respectively.

Aerosolized prolastin suppresses bacterial proliferation in a model of chronic Pseudomonas aeruginosa lung infection.

High levels of active neutrophil elastase (HNE) are present in the respiratory secretions of patients with cystic fibrosis (CF). We hypothesized that aerosolized Prolastin (alpha(1)-protease inhibitor or alpha(1)PI, purified from human blood) could suppress airway neutrophil inflammation and accelerate bacterial clearance from the lung in a model of chronic Pseudomonas aeruginosa lung infection. Because human alpha(1)PI effectively inhibits rat as well as human neutrophil elastase (NE) activity in vitro, we choose to test this hypothesis using a rat agar bead model of chronic P. aeruginosa lung infection. In this model, aerosolized Prolastin significantly decreased elastase activity (p < 0.01), lung neutrophil counts (p < 0.01), and bacterial colony counts (p < 0.01). Prolastin had no direct bactericidal effect on P. aeruginosa in vitro. Lung tissue histopathology revealed a marked decrease in lung inflammation in animals treated with Prolastin. These studies indicate that Prolastin can significantly decrease the elastase burden in the chronically infected lung. In addition, not only does Prolastin suppress lung inflammation, but it also markedly decreases P. aeruginosa density in a rat model of chronic P. aeruginosa lung infection. These data suggest that aerosolized alpha(1)PI may represent a useful nonantibiotic adjunct in the treatment and control of infection and inflammation associated with CF lung disease.

[Psychological distress and preoperative fear in surgical patients]. / Estudio del malestar psicológico y del miedo preoperatorio en pacientes quirúrgicos.

OBJECTIVE: To study the prevalence of psychological disorder, cognitive deterioration and anxiety in patients undergoing surgical procedures with general anesthesia. PATIENTS AND METHODS: A representative sample (n = 450) of surgical patients at a tertiary hospital was selected, excluding patients with a history of mental illness or drug use, and those with cancer. After admission, the day before surgery, we collected demographic, medical and surgical data and administered the Spanish versions of Folstein's Mini Cognitive Examination (MCE) and Goldberg's General Health Questionnaire (GHQ). The patients were also asked if they felt anxiety about the surgical procedure and what they feared. RESULTS: The prevalence of cognitive deterioration (MCE) was 8.7% and the prevalence of psychological disorder (GHQ 28) was 29.8% (higher for women). Combining the two instruments, 38.5% showed relevant psychological disorder. Some type of anxiety was expressed by 60.9%, with the fear of "not waking up" being the most common (26%). CONCLUSIONS: The prevalence of psychological disorder is somewhat lower than that reported by other authors for presurgical patients, probably because our study enrolled patients with no history of mental illness related to other causes. The prevalence of anxiety found is similar to that reported in the literature.

Glucose stimulates phagocytosis of unopsonized Pseudomonas aeruginosa by cultivated human alveolar macrophages.

Glucose has previously been shown to increase the in vitro phagocytosis of unopsonized Pseudomonas aeruginosa by freshly explanted murine peritoneal macrophages (PM) and cultivated alveolar macrophages (AM). This study examined the effect of glucose on the same phagocytosis process in human AM in order to determine whether this phenomenon is conserved among species. Freshly explanted human AM phagocytosed unopsonized P. aeruginosa at a low level (2 bacteria/macrophage/30 min), whereas mouse AM ingested a negligible number of P. aeruginosa (0.01 bacterium/macrophage/30 min). Glucose had no effect on this or other phagocytic processes in freshly explanted mouse or human AM. However, following in vitro cultivation for 72 h, human AM phagocytosed three to four times more unopsonized P. aeruginosa than did freshly explanted cells, but only in the presence of glucose. This glucose-inducible phagocytic response had also been observed in cultivated murine AM. Although similar increases were also detected for the phagocytosis of latex particles and complement-coated sheep erythrocytes by cultivated human AM, these processes were not glucose dependent. The lack of response to glucose in freshly explanted mouse AM was attributed to insufficient glucose transport; however, freshly explanted human AM exhibited significant facilitative glucose transport activity that was inhibitable by cytochalasin B and phloretin. Taken together, these results suggest that the process of glucose-inducible phagocytosis of unopsonized P. aeruginosa is conserved among macrophages from different species, including humans, and that AM, but not PM, required cultivation for this glucose effect to occur. Glucose transport by AM appears to be necessary but not sufficient for phagocytosis of unopsonized P. aeruginosa.

Proteolytic cleavage of ICAM-1 by human neutrophil elastase.

Human leukocyte elastase (HLE) participates in tissue destruction in a number of inflammatory disorders, including rheumatoid arthritis and cystic fibrosis. Since HLE has been shown to bind to Mac-1, and ICAM-1 plays a key role during the recruitment and the activation of leukocytes at inflamed sites, we investigated the capacity of HLE to cleave ICAM-1. Flow-cytometric analyses showed a dose-dependent cleavage of ICAM-1 by HLE on different human cell lines. The cleavage was completely inhibited by alpha1-antitrypsin, a natural HLE protease inhibitor. The ability of HLE to degrade ICAM-1 was further confirmed by electrophoretic analysis using a soluble form of ICAM-1 (D1-D5). Enzymatic removal of N-linked glycosylation did not significantly modulate ICAM-1 cleavage by HLE, while removal of sialic acid residues partially reduced the sensitivity of ICAM-1 to HLE. We further showed that sputum of cystic fibrosis patients contains high levels of HLE activity capable of cleavage of cell surface ICAM-1. The cleavage induced by incubation of cells with the sputum sample was totally inhibited by alpha1-antitrypsin and the specific peptidic HLE inhibitor N-methoxysuccinyl-Ala-Ala-Pro-Val-chloromethylketone. Moreover, the cleavage of ICAM-1 was concomitant to that of CD4 at the surface of the same cell, at the same amplitude, and at all HLE concentrations. The capacity of HLE to modulate the expression of ICAM-1 on the surface of leukocytes by proteolytic cleavage brings support to the hypothesis that overproduction of HLE can cause severe immunologic lung disorders by affecting intercellular adhesion.

DNase I acutely increases cystic fibrosis sputum elastase activity and its potential to induce lung hemorrhage in mice.

The potential of DNase I to increase cystic fibrosis sputum elastase activity and lung damage was evaluated. Sputum from CF patients induced little lung hemorrhage when instilled intranasally in C57BL/6 mice. However, sputum treated in vitro by the addition of 1 mg/ml bovine DNase I showed increased neutrophil elastase activity (7.97 +/- 1.56 versus 3.91 +/- 0.62 microM, p < 0.01) and induced marked lung hemorrhage in mice (bronchoalveolar lavage fluid hemoglobin = 192.8 +/- 40.7 versus 44.5 +/- 12.0 microg/ml, p < 0.01). These effects were not observed with DNase I alone in phosphate buffer and were suppressed by the human neutrophil elastase inhibitor methoxysuccinyl-alanyl-alanyl-prolyl-valine-chloromethylketone (MeOSAAPV-CMK). In vivo administration of 2.5 mg aerosolized recombinant human DNase I to patients with CF resulted in a 2.2-fold increase of sputum elastase activity within 1 h of treatment. Elastase levels returned to pre-rhDNase therapy levels 24 h after aerosol treatment. Sputum collected 1 h after rhDNase on 4 separate days from two of six patients in which elastase levels were highest, induced lung hemorrhage when instilled intranasally in mice. We conclude that DNase I therapy of patients with cystic fibrosis can acutely increase the elastase activity of sputum and also its potential to induce hemorrhage in the murine lung.