RNA polymerase II CTD S2P is dispensable for embryogenesis but mediates exit from developmental diapause in C. elegans.

Serine 2 phosphorylation (S2P) within the CTD of RNA polymerase II is considered a Cdk9/Cdk12-dependent mark required for 3'-end processing. However, the relevance of CTD S2P in metazoan development is unknown. We show that cdk-12 lesions or a full-length CTD S2A substitution results in an identical phenotype in Caenorhabditis elegans Embryogenesis occurs in the complete absence of S2P, but the hatched larvae arrest development, mimicking the diapause induced when hatching occurs in the absence of food. Genome-wide analyses indicate that when CTD S2P is inhibited, only a subset of growth-related genes is not properly expressed. These genes correspond to SL2 trans-spliced mRNAs located in position 2 and over within operons. We show that CDK-12 is required for maximal occupancy of cleavage stimulatory factor necessary for SL2 trans-splicing. We propose that CTD S2P functions as a gene-specific signaling mark ensuring the nutritional control of the C. elegans developmental program.

Cortisol levels in scalp hair of patients with structural heart disease.

BACKGROUND: Stress is considered a modifiable risk factor for cardiovascular disease. Scalp hair analysis is a tool to assess long-term exposure to the stress hormone cortisol. We aimed to determine the association between hair cortisol concentrations (HCC) and clinical characteristics in patients with structural heart disease. Additionally, we investigated potential predictors for longitudinal change in HCC. METHODS: The study consisted of 261 patients with structural heart disease from a randomized controlled trial of mindfulness training. One sample of scalp hair was used to determine HCC both at baseline and at 12-week follow-up. In 151 patients, HCC was available (mean age: 41.3 years, range 18-65). We investigated the association between HCC at baseline and several physiological measures (BMI, blood pressure, heart rate, respiratory rate, 6-minute walk test), as well as psychological parameters (physical and mental component summary measures (SF-36), emotional distress (HADS), and perceived stress). Additionally, we used these clinical parameters to predict HCC change over time. RESULTS: The median HCC was 22.3 pg/mg of hair (23.5 interquartile range). In multivariable linear regression analyses, an association was observed between log-transformed HCC and BMI (ß 0.171, p=0.037), respiratory rate (ß 0.194, p=0.016), and the physical summary score (ß -0.163, p=0.054). Independent predictors of log-transformed HCC change after 12 weeks were mental summary score (ß -0.200, p=0.019) and diastolic blood pressure (ß -0.171, p=0.049). CONCLUSIONS: In patients with structural heart disease a positive association exists between HCC and BMI. Mental health status may predict a change in long-term cortisol over time.

Three-dimensional computed tomographic evaluation of Le Fort III distraction osteogenesis with an external device in syndromic craniosynostosis.

There is little anteroposterior growth of the midface in patients with syndromic craniosynostosis who are followed up over time without intervention. A Le Fort III with distraction osteogenesis can be done to correct this. This is a controlled way in which to achieve appreciable stable advancement of the midface without the need for bone grafting, but the vector of the movement is not always predictable. The purpose of this study was to evaluate the 3-dimensional effect of Le Fort III distraction osteogenesis with an external frame. Ten patients (aged 7-19 years) who had the procedure were included in the study. The le Fort III procedure and the placement of the external frame were followed by an activation period and then a 3-month retention period. Computed tomographic (CT) images taken before and after operation were converted and loaded into 3-dimensional image rendering software and compared with the aid of a paired sample t test and a colour-coded qualitative analysis. Comparison of the CT data before and after distraction indicated that the amount of midface advancement was significant. Le Fort III distraction osteogenesis is an effective way to advance the midface. However, the movement during osteogenesis is not always exactly in the intended direction, and a secondary operation is often necessary. Three-dimensional evaluation over a longer period of time is necessary.

Effect of Test Parameters on the Friction Behaviour of Anodized Aluminium Alloy.

The tribological behaviour of anodic oxide layer formed on Al5754, used in automotive applications, was investigated against test parameters. The friction coefficient under different normal loads, sliding speeds, and oxide thicknesses was studied using a pin on disc tribometer. Results show that the increase of load and sliding speed increase the friction coefficient. The rise of contact pressure and temperature seems to cause changes in wear mechanism. Glow-discharge optical emission spectroscopy (GDOES) was used to investigate the chemical composition of the oxide layer. Morphology and composition of the wear tracks were analyzed by scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS). On the basis of these characterization techniques, a wear mechanism was proposed. The observed mechanical properties can be related to the morphology and the chemical composition of the layer.

Electrochemical removal of Cr(VI) from aqueous media using iron and aluminum as electrode materials: towards a better understanding of the involved phenomena.

In the present work, electrocoagulation process with iron and aluminum electrodes was investigated to deepen the understanding of the mechanism of hexavalent chromium (Cr(VI)) removal. Electrocoagulation treatment efficiency was studied with regards to the abatement of Cr(VI) and the resulting species-namely Cr(III), Fe(II) and/or Fe(III). Unlike iron, aluminum electrodes were found to be unsatisfactory for Cr(VI) removal. To elucidate the removal mechanism of hexavalent chromium, different anode/cathode materials and configurations (Fe/Fe, Pt Ti (platinized titanium)/Fe, Al/Al and Pt Ti/Al) were considered. At nearly neutral pH and considering aluminum electrodes, both electrochemical reduction (Cr(VI) to Cr(III)) at the cathode surface and adsorption on Al(OH)(3) floc mechanisms were responsible for Cr(VI) exhaustion. However, the contribution of the two mechanisms to Cr(VI) removal was not discriminated. On the other hand, in the case of iron electrodes, even though electrochemical reduction may contribute to chromium removal, its influence seemed to be minor since the effect was confined to less than 5% of the removal efficiency. Hence, there was essentially one real root for the reduction of Cr(VI) by electrocoagulation with iron electrodes, and it was proven to be the chemical reduction by Fe(II) anodically generated. Moreover, the resulting Cr(III) was quickly removed from solution, via efficient precipitation as Cr(OH)(3) hydroxides. Besides, the electrodissolved iron remained at low level owing to the precipitation of Fe(OH)(2) and/or Fe(OH)(3). Although chemical reduction by Fe(II) predominantly governed the removal of Cr(VI), acidic pH media was found to promote electrochemical reduction of hexavalent chromium at the cathode and accordingly to enhance Cr(VI) removal efficiency. In contrast, a delay of Cr(III) precipitation and a contamination of the electrolyte by electrodissolved iron were observed under acidic conditions. The effect of supporting electrolyte (Na(2)SO(4) and NaCl) on chromium removal was also studied. No conspicuous disparity in the treatment efficiency was noted under the electrocoagulation conditions used.

Behavior of aluminum electrodes in electrocoagulation process.

In the present work, electrocoagulation process with aluminum electrodes was investigated. Different operational conditions such as composition of Na(2)SO(4) based solutions, pH and current density were examined in a systematic manner. Their influence on (i) electrode polarization phenomena, (ii) pH evolution during electrolysis and (iii) the amount of Al released (coagulant) was investigated. For this purpose, potentiodynamic tests and electrolyses using different electrochemical cell configurations were conducted. It is mainly found that (i) a minimum Cl(-) concentration of the electrolyte of about 60ppm is required to breakdown the anodic passive film and considerably reduce the cell voltage during electrolysis; (ii) the anodic dissolution efficiency is unit; (iii) the global amount of coagulant (Al(3+)) generated has two origins: electrochemical oxidation of the anode and "chemical" attack of the cathode and (iv) electrolysis with Al electrodes acts as pH neutralization of the electrolytic medium. Taking into account advantage of the pH evolution observed during electrolysis, electrocoagulation tests were performed to treat a synthetic wastewater containing heavy metallic ions (Ni(2+), Cu(2+), Zn(2+)). Removal efficiencies over 98% were reached. Furthermore, our results displayed prominently that an increase of current density notably reduces the treatment duration without inducing a strong increase of the charge loading.

Expérience d'utilisation de la moustiquaire imprégnée d'insecticide en zone de haute transmission palustre de Kinshasa

PROBLEMATIQUE. En zone de transmission pérenne du paludisme où la lutte semble s'enliser, la moustiquaire imprégnée d'insecticide (MII) s'impose comme un dispositif efficace de lutte au niveau des ménages. Nos OBJECTIFS DE RECHERCHE consistait à évaluer l'impact sur la morbidité de l'enfant dans une zone de haute transmission du paludisme de Kinshasa, du taux d'utilisation ainsi que l'intégrité de la MII, d'une part, et d'autre part, l'influence de l'âge, des saisons et du standing familial. METHODOLOGIE. Une cohorte de 1.400 enfants d'âge préscolaire habitant dans une zone de haute transmission du paludisme de Kinshasa, a été mise sous MII avec l'accord éclairé des parents, et suivie durant 11 mois. RESULTATS. Le risque fébrile, pour ne prendre que ce paramètre, qui était de 13,5% au début de l'étude, a été divisé pratiquement par trois à la fin de la période d'observation (Rapport de chances = 0,29). Une ou deux nuits passées sans la MII (RC=1,81), de même que la présence d'un seul trou dans la MII (RC=2,2), ont pratiquement doublé ce risque. Ce risque a diminué graduellement avec l'âge. La présence de plus de 10 personnes à charge dans le ménage a augmenté ce risque de près de 20% (RC=1,19). Six décès, dont deux imputés à la rougeole, ont été enregistrés après 4 et 9 mois d'utilisation de la MII. CONCLUSIONS. Pour autant qu'elle soit régulièrement utilisée et que son intégrité soit assurée, la MII réduit significativement le risque morbide imputable au paludisme. L'hypothèse de la perte de la prémunition a été invoquée pour expliquer les décès de cause non définie, probablement due au paludisme, survenus en dépit de l'utilisation régulière supposée de la MII

Quantitative determination of hydrogen in solids by gas chromatography.

Processes such as electroplating or acid cleaning are notorious causes of post-processing failure through hydrogen embrittlement. So, the determination of amounts of hydrogen in metals is of great importance. An analysis method for investigation of H content in solids has been established based on hot extraction and gas chromatography system. Hot extraction in inert gas enables complete and/or partial removal of the hydrogen from the samples. A gas chromatography system is used to determine quantitatively the amount of thermally desorbed hydrogen. An investigation of the baking operating conditions is made of the hydrogen desorption rate of zinc-plated steel parts. Then, an analysis of the polarisation conditions upon chromium electroplating is given.

HU-GFP and DAPI co-localize on the Escherichia coli nucleoid.

The heterodimeric HU protein, one of the most abundant DNA binding proteins, plays a pleiotropic role in bacteria. Among others, HU was shown to contribute to the maintenance of DNA superhelical density in Escherichia coli. By its properties HU shares some traits with histones and HMG proteins. More recently, its specific binding to DNA recombination and repair intermediates suggests that HU should be considered as a DNA damage sensor. For all these reasons, it will be of interest to follow the localization of HU within the living bacterial cells. To this end, we constructed HU-GFP fusion proteins and compared by microscopy the GFP green fluorescence with images of the nucleoid after DAPI staining. We show that DAPI and HU-GFP colocalize on the E. coli nucleoid. HU, therefore, can be considered as a natural tracer of DNA in the living bacterial cell.

Rodent malaria in the natural host--irradiated sporozoites of Plasmodium berghei induce liver-stage specific immune responses in the natural host Grammomys surdaster and protect immunized Grammomys against P. berghei sporozoite challenge.

The choice of the host in studying host-parasite interactions is of crucial importance, and the use of a natural host is most appropriate in answering pertinent questions related to human malaria. The Grammomys surdaster is the natural host and reservoir of the rodent malaria parasite Plasmodium berghei. This natural host is difficult to protect by irradiated sporozoite immunization, a situation comparable to what has been observed in humans with P. falciparum. This is in contrast to the complete protection that can be induced in artificial hosts like inbred mice strains. The natural host is highly susceptible to P. berghei hepatic stage infections. Immunization with irradiated sporozoites in Grammomys generates blocked hepatic stage parasites and immunized Grammomys protected upon live sporozoite challenge generate antibody and T cell proliferative responses to these hepatic stages. Associated with proliferation, cytokines are secreted into culture supernatants constituted mainly of Interferon gamma, negligible amounts of TNF-alpha, and no IL-4. Natural host-parasite interactions of Grammomys surdaster-P. berghei can help define the effector mechanism(s) in the Plasmodium falciparum-human interaction.

Overproduction and improved strategies to purify the threenative forms of nuclease-free HU protein.

The heterodimeric HU protein was isolated from Escherichia coli as one of the most abundant DNA binding proteins associated with the bacterial nucleoid. HUalphabeta is composed of two very homologous subunits, but HU can also be present in E. coli under its two homodimeric forms, HUalpha(2) and HUbeta(2). This protein is conserved either in its heterodimeric form or in one of its homodimeric forms in all bacteria, in plant chloroplasts and in some viruses. HU can participate, like the histones, in the maintenance of DNA supercoiling and in DNA condensation. This protein which does not recognize any specific sequence on double-stranded DNA, has been shown to bind specifically to cruciform DNA as does the eukaryotic HMG1 protein and to a series of structures which are found as intermediates of DNA repair, e.g., nick, gap, 3'overhang, etc. The strong binding of HU to these diverse DNA structures could explain, in part at least, its pleiotropic role in the bacterial cell. To understand all the facets of its interactions with nucleic acids, it was necessary to develop a procedure which allowed the purification of the three forms of HU under their native form and without the nuclease activity strongly associated with the protein. We describe here such a procedure as well as demonstrating that the three histidine-tagged HUs we have produced, have conserved the binding characteristics of native HUs. Interestingly, by two complementation tests, we show that the histidine-tagged HUs are fully active in vivo.

Plasmodium berghei development in irradiated sporozoite-immunized C57BL6 mice.

The C57BL6 strain of mice is highly susceptible to Plasmodium berghei sporozoite infections and consequently requires repeated immunizations with irradiated sporozoites to obtain protective immunity. After a live sporozoite challenge in the immunized hosts, hepatic-stage parasites found in the liver after 48 h are of different sizes--small schizonts corresponding to blocked forms (derived from irradiated sporozoites), and schizonts of intermediate size (derived from live sporozoites). Large schizonts corresponding to mature hepatic forms are found only in unimmunized but challenged C57BL6 mice. Using monoclonal and polyclonal antibodies directed to liver-stage parasites, different patterns of binding reactivity to the above forms are observed. More than 20% of the irradiated sporozoites transform into blocked forms after immunization and persist in the liver. Upon sporozoite challenge in such immunized animals the rate of transformation of sporozoites into hepatic parasites is less than 2%. These observations shed light on the fate of live sporozoite development in irradiated sporozoite-immunized C57BL6 mice.

Irradiated sporozoites prime mice to produce high antibody titres upon viable Plasmodium berghei sporozoite challenge, which act upon liver-stage development.

C57BL6 mice were protected against Plasmodium berghei sporozoite challenge by immunization with live 12 krad dose-irradiated sporozoites, but not by 20 krad dose-irradiated sporozoites. Immunization with 12 krad irradiated sporozoites generated low levels of antibody reactive to liver-stage parasites (titres of 1/100). Inoculation of as few as 100 live P. berghei sporozoites induced complete host protection accompanied by a very quick and high boost of antibody titres up to 1/4000. This sporozoite challenge-drive antibody boost was absent in mice immunized by 20 krad sporozoites and in non-protected, and non-immunized mice. Antibody was mainly liver-stage (LS) specific and due to an increase of IgG2a and IgG2b. The in vitro effect of pre- and post-challenge sera upon either sporozoite invasion or LS development was assessed in Hep-G2 cultures. Both were found to have a strong effect upon LS development even at 1/2500 dilution, and conversely a low effect upon invasion. These results suggest that sporozoites irradiated at doses that induce protection are able to prime T-cells which, upon challenge by non-irradiated sporozoites, provide help to B-lymphocytes to trigger the production of high titres of anti-LS antibodies that can inhibit LS development in vitro.

Blocked hepatic-stage parasites and decreased susceptibility to Plasmodium berghei infections in BALB/c mice.

The BALB/c strain of mice is comparatively more resistant to sporozoite infections of Plasmodium berghei than the C57BL6 strain. Infection with live sporozoites results in the formation of small hepatic forms in the BALB/c liver that persist for as long as 6 days. Upon infection with small numbers of sporozoites, some of the parasites are destroyed in the liver whereas the rest persist as blocked forms. When larger numbers of sporozoites are injected the same process occurs but, in addition, a fraction of the liver-stage parasites complete full development and give rise to blood forms. Although blocked liver forms persist until day 6 post-infection they actually develop to only 24 h of maturity. The nature of these persistent forms is similar to those obtained from irradiated sporozoite immunization. There is a stronger cell proliferation to liver-stage antigens by spleen lymphocytes of irradiated sporozoite-immunized BALB/c mice in comparison to that of immunized C57BL6 mice suggesting that a stronger priming to liver-stage antigens, probably due to the presence of blocked hepatic forms in the liver for a longer period of time (as compared to C57BL6), occurs in the BALB/c mice. This could be a reason for the long-lasting protective memory observed in BALB/c mice.

Cytokine levels during mild and cerebral falciparum malaria in children living in a mesoendemic area.

Cell-mediated immunity and cytokines are probably involved in the pathogenesis of malaria. To investigate the role and the activity of different immune cells, we measured levels of tumour necrosis factor-(TNF-alpha), gamma interferon (IFN-gamma) and several interleukins (IL-2, IL-4, IL-6 and IL-10) in children with mild (MM) and cerebral (CM) Plasmodium falciparum malaria and compared them with those of healthy children from Guadalupe--Lobata District, St. Tomé Island, where malaria is mesoendemic. Both groups of patients had significantly higher levels of IL-6, IL-10 and TNF-alpha than controls. For IL-2, IL-4 and IFN-gamma we found no difference between the groups. However, 24 h after admission the levels of IL-10 and IL-6 were significantly higher in CM than in MM patients, although 7 days after treatment they returned to normal levels, similar to those found in control children. Therefore, TNF-alpha IL-6 and IL-10 increase during Plasmodium falciparum attacks in all children, not only in those with cerebral malaria. This finding suggests the activation of the monocyte/macrophage system during the early stage of clinical malaria.

Plasmodium falciparum chloroquine and quinine sensitivity in asymptomatic and symptomatic children in São Tomé Island.

Plasmodium falciparum sensitivity to quinine in São Tomé was determined by in vivo and in vitro tests in 56 children with mild or cerebral malaria. Chloroquine sensitivity was assessed by in vitro tests in 105 parasitaemic asymptomatic children from the same community as the cases. The WHO standard methodology was used. No resistance to quinine was found by in vivo or in vitro tests in either group of patients or in asymptomatic children, although some degree of chloroquine resistance was found with the in vitro test. This was more common in patients than in asymptomatic children. Chloroquine resistance may be explained by the recent history of malaria in São Tomé Island, which caused an important decrease of immunity among the population and consequently the emergence of resistant strains. Implications of the use of in vivo/in vitro tests for determining the antimalarial drug policy within the primary health care system are discussed.

Adult Schistosoma mansoni worms positively modulate soluble egg antigen-induced inflammatory hepatic granuloma formation in vivo. Stereological analysis and immunophenotyping of extracellular matrix proteins, adhesion molecules, and chemokines.

Synchronized liver granulomas were induced by injecting Sepharose beads to which SEA soluble egg antigen (SEA) or the concanavalin A binding fraction of SEA had been coupled into a mesenteric vein in naive, single-sex (35 days) and bisexually (28 days) Schistosoma mansoni-infected and Plasmodium berghei-immunized mice. Stereological analysis revealed that peak granuloma formation was already reached 8 days after injection in single-sex infected mice compared with 16 days in naive animals. No difference in granuloma formation between naive and P. berghei-immunized animals and between unisexually and bisexually S. mansoni-infected mice was observed. This suggests that the positive immunomodulatory effect on the granulomogenesis is worm specific and not likely to be due to arousal of the immune system by unrelated factors, nor is it influenced by the gender or degree of maturation of female worms. At all stages in time, the concanavalin A binding-fraction-induced granulomas reached only 65 to 70% of the volume of SEA-induced granulomas. Immunophenotyping of extracellular matrix proteins around deposited heads revealed that fibronectin was the dominant extracellular matrix protein and that also type I and IV collagen and laminin were deposited. Temporal analysis of the expression of the adhesion molecules ICAM-1, LFA-1, VLA-4, and VLA-6 was performed. Morphological evidence is presented for the role of adhesion molecules in the initiation and maintenance of hepatic granuloma formation. The chemokine monocyte chemoattractant protein-1 was expressed in the granuloma and in hepatic artery branches. From these data, it is concluded that adult S. mansoni worms positively modulate schistosomal hepatic granuloma formation in vivo. Adhesion molecules and chemokines play important roles in schistosomal granuloma formation.

Immunoblot characterization of IgG, IgM and IgE antibodies elicited during the course of fatal and non-fatal infections of Plasmodium berghei in DBA/2 and Balb/c mice.

When infected with the CRS line of Plasmodium berghei ANKA, DBA/2 mice died of a fulminating infection around day 20 post-infection whereas Balb/c mice had crises around day 15 then low or subpatent parasitaemias until approximately day 73, when sterile immunity is believed to have supervened. Immunoblots for parasite-specific immunoglobulins G, M and E were made from the sera taken during the course of infection in each mouse strain. Although both strains elicited antibodies to a 128-kDa antigen by day 8, this was solely of the IgG class in the DBA/2 but of both IgG and IgM in the Balb/c. Crisis and subpatency were associated with the appearance of IgG and IgM antibodies to 74- and 80-kDa antigens. The most vigorous immune response was observed in sera from the Balb/c mice on day 73, with antibodies to many antigens of 19-180 kDa; IgE antibody (to antigens of 38 and 45 kDa) was then evident for the first time.