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Phagocytes generate superoxide (O2*-) by an enzyme complex known as reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Its catalytic component, responsible for the NADPH-driven reduction of oxygen to O2*-, is flavocytochrome b559, located in the membrane and consisting of gp91phox and p22phox subunits. NADPH oxidase activation is initiated by the translocation to the membrane of the cytosolic components p47phox, p67phox, and the GTPase Rac. Cytochrome b559 is converted to an active form by the interaction of gp91phox with p67phox, leading to a conformational change in gp91phox and the induction of electron flow. We designed a new family of NADPH oxidase activators, represented by chimeras comprising various segments of p67phox and Rac1. The prototype chimera p67phox (1-212)-Rac1 (1-192) is a potent activator in a cell-free system, also containing membrane p47phox and an anionic amphiphile. Chimeras behave like bona fide GTPases and can be prenylated, and prenylated (p67phox -Rac1) chimeras activate the oxidase in the absence of p47phox and amphiphile. Experiments involving truncations, mutagenesis, and supplementation with Rac1 demonstrated that the presence of intrachimeric bonds between the p67phox and Rac1 moieties is an absolute requirement for the ability to activate the oxidase. The presence or absence of intrachimeric bonds has a major impact on the conformation of the chimeras, as demonstrated by fluorescence resonance energy transfer, small angle X-ray scattering, and gel filtration. Based on this, a "propagated wave" model of NADPH oxidase activation is proposed in which a conformational change initiated in Rac is propagated to p67phox and from p67phox to gp91phox.

Public transport in Delhi was amended by the Supreme Court of India to use Compressed Natural Gas (CNG) instead of diesel or petrol. After the implementation of CNG since April 2001, Delhi has the highest fraction of CNG-run public vehicles in the world and most of them were introduced within 20 months. In the present study, the concentrations of various criteria air pollutants (SPM, PM(10), CO, SO(2) and NO(x)) and organic pollutants such as benzene, toluene, xylene (BTX) and polycyclic aromatic hydrocarbons (PAHs) were assessed before and after the implementation of CNG. A decreasing trend was found for PAHs, SO(2) and CO concentrations, while the NO(x) level was increased in comparison to those before the implementation of CNG. Further, SPM, PM(10), and BTX concentrations showed no significant change after the implementation of CNG. However, the BTX concentration demonstrated a clear relation with the benzene content of gasoline. In addition to the impact of the introduction of CNG the daily variation in PAHs levels was also studied and the PAHs concentrations were observed to be relatively high between 10 pm to 6 am, which gives a proof of a relation with the limited day entry and movement of heavy vehicles in Delhi.

Landfilling is one of the most common ways of municipal solid waste disposal. Degradation of organic waste produces CH(4) and other landfill gases that significantly contribute to global warming. However, before entering the atmosphere, part of the produced CH(4) can be oxidised while passing through the landfill cover. In the present study, the oxidation rate of CH(4) was studied with various types of compost as possible landfill cover. The influence of incubation time, moisture content and temperature on the CH(4) oxidation capacity of different types of compost was examined. It was observed that the influence of moisture content and temperature on methane oxidation is time-dependent. Maximum oxidation rates were observed at moisture contents ranging from 45% to 110% (dry weight basis), while the optimum temperature ranged from 15 to 30 degrees C.

MdfA is an Escherichia coli multidrug transporter of the major facilitator superfamily (MFS) of secondary transporters. Although several aspects of multidrug recognition by MdfA have been characterized, better understanding the detailed mechanism of its function requires structural information. Previous studies have modeled the 3D structures of MFS proteins, based on the X-ray structure of LacY and GlpT. However, because of poor sequence homology, between LacY, GlpT, and MdfA additional constraints were required for a reliable homology modeling. Using an algorithm that predicts the angular orientation of each transmembrane helix (TM) (kPROT), we obtained a remarkably similar pattern for the 12 TMs of MdfA and those of GlpT and LacY, suggesting that they all have similar helix packing. Consequently, a 3D model was constructed for MdfA by structural alignment with LacY and GlpT, using the kPROT results as an additional constraint. Further refinement and a preliminary evaluation of the model were achieved by correlated mutation analysis and the available experimental data. Surprisingly, in addition to the previously characterized membrane-embedded glutamate at position 26, the model suggests that Asp34 and Arg112 are located within the membrane, on the same face of the cavity as Glu26. Importantly, Arg112 is evolutionarily conserved in secondary drug transporters, and here we show that a positive charge at this position is absolutely essential for multidrug transport by MdfA.

Activation of the phagocyte NADPH oxidase is the consequence of the assembly of membranal cytochrome b559 with the cytosolic components p47phox, p67phox, and the GTPase Rac and is mimicked by a cell-free system comprising these components and an activator. We designed a variant of this system, consisting of membranes, p67phox) prenylated Rac1-GDP, and the Rac-specific guanine nucleotide exchange factor (GEF) Trio, in which oxidase activation is induced in the absence of an activator and p47phox. We now show that: 1) Trio and another Rac GEF (Tiam1) act by inducing GDP to GTP exchange on prenylated Rac1-GDP and that our earlier assertion that activation is GTP-independent is explained by contamination of p67phox preparations with GTP and/or ATP. 2) Oxidase activation by Rac GEFs is supported not only by GTP but also by ATP. 3) Non-hydrolysable GTP analogs are active, whereas ATP analogs, incapable of gamma-phosphoryl transfer, are inactive. 4) The ability of ATP to support GEF-induced oxidase activation is explained by ATP serving as a gamma-phosphoryl donor for a membrane-localized nucleoside diphosphate kinase (NDPK), converting GDP to GTP. 5) The existence of a NDPK in macrophage membranes is proven by functional, enzymatic, and immunologic criteria. 6) NDPK acts on free GDP, and the newly formed GTP is bound again to Rac. 7) Free GDP is derived exclusively by dissociation from prenylated Rac1-GDP, mediated by GEF. NDPK and GEF appear to be functionally linked in the sense that the availability of GDP, serving as substrate for NDPK, is dependent on the level of activity of GEF.

NADPH oxidase activation involves the assembly of membrane-localized cytochrome b559 with the cytosolic components p47phox, p67phox, and the small GTPase Rac. Assembly is mimicked by a cell-free system consisting of membranes and cytosolic components, activated by an anionic amphiphile. We reported that a chimeric construct, consisting of residues 1-212 of p67phox and full-length Rac1, activates the oxidase in vitro in an amphiphile-dependent manner, and when prenylated, in the absence of amphiphile and p47phox. We subjected chimera p67phox-(1-212)-Rac1 to mutational analysis and found that: 1) replacement of a single basic residue at the C terminus of the Rac1 moiety by glutamine is sufficient for loss of activity by the non-prenylated chimera; replacement of all six basic residues by glutamines is required for loss of activity by the prenylated chimera. 2) A V204A mutation in the activation domain of the p67phox moiety leads to a reduction in activity. 3) Mutating residues, known to participate in the interaction between free p67phox and Rac1, in the p67phox-(R102E) or Rac1 (A27K, G30S) moieties of the chimera, leads to a marked decrease in activity, indicating a requirement for intrachimeric bonds, in addition to the engineered fusion. 4) Chimeras, inactive because of mutations A27K or G30S in the Rac1 moiety, are reactivated by supplementation with exogenous Rac1-GTP but not with exogenous p67phox. This demonstrates that Rac has a dual role in the assembly of NADPH oxidase. One is to tether p67phox to the membrane; the other is to induce an "activating" conformational change in p67phox.

Genotypes 1 and 2 of Cryptosporidium parvum are the primary types associated with infections in humans, with type 1 being by far the predominant genotype. The frequency of mixed infection with both genotypes in humans is relatively rare, while type 1, which experimentally infects other mammals, has been found to naturally infect almost exclusively humans. One possible explanation for the absence of type 1 in other mammals and the low frequency of mixed infections in humans is the inability of type 1 to compete with type 2 in nature when both occur simultaneously. To investigate this, we challenged gnotobiotic piglets with equal number of oocysts of type 1 and type 2, given either simultaneously or with type 2 given 24 or 48 h after type 1. The genotype of the oocysts excreted in feces and the relative distribution of each of the genotypes throughout the gut at necropsy were determined. Regardless of the time interval between challenges with the two genotypes, type 2 invariably displaced type 1. The rate of displacement was rapid when both genotypes were given simultaneously, after which no traces of type 1 were detected in the feces or in gut sections by PCR. Infection with type 1 24 or 48 h before challenge with type 2, while permitting type 1 to become established, was still rapidly eliminated within 3 days after challenge with type 2. These observations have major implications regarding the relative perpetuation and survival of these two genotypes in mammals.

Spatial patterns of various criteria air pollutants, like SO2, NO2, O3, and TSP were studied at Shahdara National Ambient Air Quality Monitoring station in Delhi (India) in July 1999. The minimum pollutant concentrations were observed during morning hours, whereas the highest concentrations were found during the late night hours, which seem to be related with the vehicular emission. Pre-monsoon daily ambient air quality spatial pattern was compared with the spatial pattern during initial and subsequent rain shower of monsoon. These spatial patterns were found to be essentially the same before and during rain, however a significant decrease in SO2, NO2 and TSP concentrations (40-45%) was observed after initial and subsequent rains of the monsoon, demonstrating the importance of rainfall in the scavenging of these criteria air pollutants.

The effect of fireworks on air quality was assessed from the ambient concentrations of various air pollutants (SO2, NO2, PM10 and TSP) during Diwali festival in Hisar city (India), in November 1999. The extensive use of fireworks was found to be related to short-term variation in air quality. During the festival the concentration of SO2 was observed to be increased approximately 10-fold at few sites, whereas the concentrations of NO2, PM10 and TSP increased 2-3 times, compared to the data collected on a typical winter day in December 1999. The maximum NO2 concentration was observed a day after the festival. The diurnal pattern of the above pollutants showed a slight increase in the night. The levels of these pollutants observed during Diwali were found to be moderately high, which can be associated with serious health impacts.

This study was designed to determine the prognostic significance of multidrug resistance, mediated by P-glycoprotein (Pgp) expression, in Ewing sarcoma. The clinical and laboratory features, treatment protocol, and outcome of 75 patients with Ewing sarcoma or peripheral neuroectodermal tumor treated between 1972 and 1997 were reviewed. Pgp expression was tested with the monoclonal antibody JSB-1. Thirty-four (64%) of the 53 tissue samples from untreated patients stained positive for Pgp. Progression-free and overall survival were 44 and 59%, respectively, in patients with negative findings, and 28 and 41% in those with positive findings; neither difference was significant. Of the 12 relapsed patients, 6 (50%) expressed more Pgp after chemotherapy than at diagnosis and 4 (33%) expressed less. Within these subgroups, 5 out of 6 and 3 out of 4 died from the disease. No correlation was found between Pgp and known prognostic factors of Ewing tumors. Pgp expression is probably an intrinsic factor of Ewing tumors but has no correlation to prognosis.

The effect of histamine on the production of cytokines by subpopulations of mononuclear cells was studied. A 3.5-fold increase in the number of myeloid colony-forming units (CFU-C) was observed when bone marrow cells were cultured in the presence of conditioned medium prepared from nonadherent mononuclear cells cultured with 10(-4) M histamine (CM-histamine) compared with phosphate-buffered saline (CM-PBS). Using ELISA and radioimmunoassay kits, histamine was found to enhance the production of GM-CSF (9.6-fold) and IL-6 (8.2-fold) by mononuclear cells but not by nonadherent cells or large granular lymphocytes. Anti-GM-CSF and anti-IL-6 antibodies markedly blocked cytokine activity in CM-PBS, whereas the blocking effect in CM-histamine was moderate, indicating enhanced GM-CSF and IL-6 activity in CM-histamine. No GM-CSF or IL-6 levels could be detected in CM-histamine or CM-PBS prepared from CD3+, CD4+, or CD8+ lymphocytes. Preincubation of CM-histamine with H1 and H2 receptor antagonists resulted in complete blocking of the histamine-enhanced colony-stimulating activity. We conclude that histamine is able to activate human mononuclear cells to generate cytokines such as GM-CSF and IL-6 via H1 and H2 receptors.

We compared the effect of haematopoietic growth factors granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin (IL)-1, IL-3, and IL-5 on the functional activation of human eosinophils and neutrophils from the same donor. All four colony-stimulating factors (CSF) enhanced the phagocytosis of Candida albicans by eosinophils and increased staphylococcal, but not Candida, killing. GM-CSF and IL-5 had a profound stimulating effect on eosinophil staphylocidal activity. GM-CSF and IL-3 enhanced the generation of leukotriene C4 (LTC4) induced by calcium ionophore A23187 and the release of arylsulphatase and beta-glucuronidase from specific and small granules of eosinophils. In contrast, IL-1 and IL-5 had no effect on degranulation. GM-CSF and IL-1 enhanced phagocytosis of C. albicans by neutrophils, and GM-CSF stimulated degranulation and the release of the enzymes beta-glucuronidase and arylsulphatase from neutrophils while IL-1 stimulated the release of arylsulphatase only. This study indicates that the eosinophil-active colony-stimulating factors can markedly enhance the host defence function of the eosinophil and even make it the equal of the neutrophil in staphylocidal activity. The CSF-activated eosinophil, however, may cause inappropriate inflammation and normal tissue damage.

Four experiments investigated the perception of tonal structure in polytonal music. The experiments used musical excerpts in which the upper stave of the music suggested a different key than the lower stave. In Experiment 1, listeners rated the goodness of fit of probe tones following an excerpt from Dubois's Circus. Results suggested that listeners were sensitive to two keys, and weighted them according to their perceived importance within the excerpt. Experiment 2 confirmed that music within each stave reliably conveyed key structure on its own. In Experiment 3, listeners rated probe tones following an excerpt from Milhaud's Sonata No. 1 for Piano, in which different keys were conveyed in widely separate pitch registers. Ratings were collected across three octaves. Listeners did not associate each key with a specific register. Rather, ratings for all three octave registers reflected only the key associated with the upper stave. Experiment 4 confirmed that the music within each stave reliably conveyed key structure on its own. It is suggested that when one key predominates in a polytonal context, other keys may not contribute to the overall perceived tonal structure. The influence of long-term knowledge and immediate context on the perception of tonal structure in polytonal music is discussed.

Contact sensitivity (CS), anti-DNP (dinitrophenyl) IgG and IgE type antibodies were induced by skin applications of the contact sensitizer dinitrofluorobenzene. CS response was suppressed by treatment with the monovalent hapten dinitrobenzene sulfonate (DNBS) but was not affected by treatment with the DNP-conjugates (DNP22-Ficoll, DNP17-polyethylene glycol and DNP2-polyvinylpyrrolidone). On the other hand, IgE type response was depressed by treatment with the various DNP conjugates but was not affected by DNBS. The IgG type response was not appreciably affected by any of the treatments.

Mice of the randomly bred strain ICR and of the inbred strain BALB/c could be successfully sensitized to chromate by repeated topical applications of a solution of potassium bichromate in dimethyl sulfoxide. The specificity of chromium sensitization was ascertained by ear tests, specific desensitization by intravenous injection of potassium bichromate and by histological ear sections showing marked infiltrates of leukocytes following ear application of chromium-dimethyl sulfoxide in already sensitized animals. It is concluded that mice may provide a useful experimental model for study of contact sensitivity to chromium.

The frequency of serum alkaline phospatase phenotypes and secretor trait is determined for Israeli Jews originating from three distinct geographical regions: Eastern and Central Europe (group I), North Africa (group II) and the Middle East (group III). The frequency of the Pp 2 serum alkaline phosphatase phenotype in each of the three groups were 33.56% (group I), 29.79% (group II) and 24.07% (group III). The average frequency for the three groups combined was 30.58%. The corresponding frequencies of the se gene in the three groups were 0.4765, 0.5149 and 0.5468, respectively, and the average frequency for the total number examined was 0.4978.

The Nesbit operation gave excellent results in 85% of 22 men with erectile deformities of the penis due to various congenital causes. The dermal graft procedure has been abandoned for the treatment of Peyronie's disease as a result of an unacceptably high incidence of post-operative erectile problems. The Nesbit technique successfully corrected the erectile deformity in 77% of 62 men with Peyronie's disease. Those patients with Peyronie's disease and erectile impairment due to cavernous fibrosis are best treated by the insertion of a penile prosthesis.