Role of gastrin/pentagastrin in regulation of intestinal cytochrome P-450

1. In the absence of intraluminal inducers, low "basal" levels of cytochrome P-450 and its dependent MFO activities are detected in the rat intestinal mucosa, and may be regulated by endogenous hormones. 2. Rats were nutritionally maintained by either short term (48 hr) intravenous glucose infusion or chronic (8 days) intravenous hyperalimentation, and were treated with various doses of pentagastrin in the infusate. 3. Regardless of the dose (6-90 micrograms/kg/hr) or duration of infusion (2-8 days), pentagastrin had no effect on small intestinal cytochrome P-450, its dependent MFO activity, or the activity of delta-aminolevulinic acid synthetase. 4. The intestinal trophic peptide hormone, gastrin, apparently does not regulate the cytochrome P-450-dependent MFO system of the small intestine.     

Novel Virus Influenza A (H1N1sw) in South-Eastern France,April-August 2009

Background

In April 2009, the first cases of pandemic (H1N1)-2009 influenza [H1N1sw] virus were detected in France. Virological surveillance was undertaken in reference laboratories of the seven French Defence Zones.

Methodology/Principal Findings

We report results of virological analyses performed in the Public Hospitals of Marseille during the first months of the outbreak. (i) Nasal swabs were tested using rapid influenza diagnostic test (RIDT) and two RT-PCR assays. Epidemiological characteristics of the 99 first suspected cases were analyzed, including detection of influenza virus and 18 other respiratory viruses. During three months, a total of 1,815 patients were tested (including 236 patients infected H1N1sw virus) and distribution in age groups and results of RIDT were analyzed. (ii) 600 sera received before April 2009 and randomly selected from in-patients were tested by a standard hemagglutination inhibition assay for antibody to the novel H1N1sw virus. (iii) One early (May 2009) and one late (July 2009) viral isolates were characterized by sequencing the complete hemagglutinine and neuraminidase genes. (iiii) Epidemiological characteristics of a cluster of cases that occurred in July 2009 in a summer camp were analyzed.

Conclusions/Significance

This study presents new virological and epidemiological data regarding infection by the pandemic A/H1N1 virus in Europe. Distribution in age groups was found to be similar to that previously reported for seasonal H1N1. The first seroprevalence data made available for a European population suggest a previous exposure of individuals over 40 years old to influenza viruses antigenically related to the pandemic (H1N1)-2009 virus. Genomic analysis indicates that strains harbouring a new amino-acid pattern in the neuraminidase gene appeared secondarily and tended to supplant the first strains. Finally, in contrast with previous reports, our data support the use of RIDT for the detection of infection in children, especially in the context of the investigation of grouped cases.     

TGF-beta-associated enhanced susceptibility to leishmaniasis following intramuscular vaccination of mice with Leishmania amazonensis antigens

Leishmania amazonensis and Leishmania braziliensis are the main causal agents of anergic diffuse cutaneous leishmaniasis and hyperergic mucosal leishmaniasis in man, respectively. In this work we demonstrate that intramuscular vaccination of BALB/c mice with whole antigens of L. amazonensis (LaAg) but not L. braziliensis (LbAg) results in increased susceptibility to cutaneous leishmaniasis. LaAg vaccination resulted in an increased capacity of the draining lymph nodes to produce IL-10 and TGF-beta during antigen recall responses. In vitro cultivation with LaAg but not LbAg induced increased apoptosis of CD8+ T cells. Following infection with L. amazonensis, LaAg-vaccinated mice produced significantly more TGF-beta and a higher serum IgG1/IgG2a antibody ratio compared with LbAg-vaccinated and non-vaccinated animals. The association of TGF-beta with enhanced susceptibility to infection was confirmed in mice co-vaccinated with LaAg and neutralizing anti-TGF-beta antibodies. Upon parasite challenge, these animals developed much smaller lesion sizes and parasite burdens, comparable with non-vaccinated controls. The disease-promoting effect of LaAg vaccination is not a general event, as in contrast to BALB/c, the disease outcome in C57Bl/6 mice was unaltered. Together, these findings indicate that species-specific components of L. amazonensis activate overt TGF-beta production that predisposes more susceptible individuals to aggravated disease following vaccination.     

Solid state and solution studies of a vanadium(III)-L-cysteine compound and demonstration of its antimetastatic, antioxidant and inhibition of neutral endopeptidase activities

Reaction of one equivalent of vanadium(III) chloride with three equivalents of l-cysteine(H2Cys) in methyl alcohol affords a VIII-Cys compound that is formulated as [VIII(Hcys)3].2HCl.2.5H2O 1. The solid state characterization of 1 was performed by microanalysis, circular dichroism (CD) and infrared studies as well as room temperature magnetic susceptibility. These studies have shown coordination of each HCys- ligand to the VIII atom through an amine nitrogen and a carboxylate oxygen atoms. Solution studies of 1 were carried out in water and methanol by UV-visible, CD and electron paramagnetic resonance (EPR) spectroscopies. According to these studies, it was evident that despite the progressive oxidation of 1 to oxovanadium(IV) species, some V(III) species were also present in solution after several hours. Compound 1, VIVOSO4.5H2O and l-cysteine were examined for their total antioxidant capacity (TAC) and lag time. Compound 1 exhibited significantly greater total antioxidant capacity and lag time values than l-cysteine. VIVOSO4.5H2O did not show any total antioxidant capacity or lag time. The inhibition of neutral endopeptidase (NEP) activity caused by 1, VIVOSO4.5H2O and thiorphan was also measured. Compound 1, at a concentration of 10(-3) M, showed inhibition of NEP activity as potent as thiorphan at 10(-6) M, while VIVOSO4.5H2O in the same concentration exhibited less than 50% inhibitory activity than that of thiorphan at 10(-6) M. Moreover, the antimetastatic effects of compound 1, l-cysteine and VIVOSO4.5H2O were examined on Wistar rats, treated with 3,4-benzopyrene. The results revealed that 1 prevents significantly lung metastases (only 9.5% of animals treated with 1 showed metastases), whereas 47-52% of the rats of the control group and those treated with l-cysteine and VIVOSO4.5H2O exhibited metastases.     

Functional Expression of Sinorhizobium meliloti BetS, a High-Affinity Betaine Transporter, in Bradyrhizobium japonicum USDA110

Among the Rhizobiaceae, Bradyrhizobium japonicum strain USDA110 appears to be extremely salt sensitive, and the presence of glycine betaine cannot restore its growth in medium with an increased osmolarity (E. Boncompagni, M. Østerås, M. C. Poggi, and D. Le Rudulier, Appl. Environ. Microbiol. 65:2072-2077, 1999). In order to improve the salt tolerance of B. japonicum, cells were transformed with the betS gene of Sinorhizobium meliloti. This gene encodes a major glycine betaine/proline betaine transporter from the betaine choline carnitine transporter family and is required for early osmotic adjustment. Whereas betaine transport was absent in the USDA110 strain, such transformation induced glycine betaine and proline betaine uptake in an osmotically dependent manner. Salt-treated transformed cells accumulated large amounts of glycine betaine, which was not catabolized. However, the accumulation was reversed through rapid efflux during osmotic downshock. An increased tolerance of transformant cells to a moderate NaCl concentration (80 mM) was also observed in the presence of glycine betaine or proline betaine, whereas the growth of the wild-type strain was totally abolished at 80 mM NaCl. Surprisingly, the deleterious effect due to a higher salt concentration (100 mM) could not be overcome by glycine betaine, despite a significant accumulation of this compound. Cell viability was not significantly affected in the presence of 100 mM NaCl, whereas 75% cell death occurred at 150 mM NaCl. The absence of a potential gene encoding Na⁺/H⁺ antiporters in B. japonicum could explain its very high Na⁺ sensitivity.     

Direct targeting and rapid isolation of BAC clones spanning a defined chromosome region

To isolate genes of interest in plants, it is essential to construct bacterial artificial chromosome (BAC) libraries from specific genotypes. Construction and organisation of BAC libraries is laborious and costly, especially from organisms with large and complex genomes. In the present study, we developed the pooled BAC library strategy that allows rapid and low cost generation and screening of genomic libraries from any genotype of interest. The BAC library is constructed, directly organised into a few pools and screened for BAC clones of interest using PCR and hybridisation steps, without requiring organization into individual clones. As a proof of concept, a pooled BAC library of approximately 177,000 recombinant clones has been constructed from the barley cultivar Cebada Capa that carries the Rph7 leaf rust resistance gene. The library has an average insert size of 140 kb, a coverage of six barley genome equivalents and is organised in 138 pools of about 1,300 clones each. We rapidly established a single contig of six BAC clones spanning 230 kb at the Rph7 locus on chromosome 3HS. The described low-cost cloning strategy is fast and will greatly facilitate direct targeting of genes and large-scale intra- and inter-species comparative genome analysis.Edwige Isidore and Beatrice Scherrer contributed equally to the work.     

Urea potentiometric enzymatic biosensor based on charged biopolymers and electrodeposited polyaniline

A potentiometric biosensor based on urease was developed for the quantitative determination of urea concentration in aqueous solutions for biomedical applications. The urease was either physisorbed onto an electrodeposited polyaniline film (PANI), or immobilized on a layer-by-layer film (LbL) assembled over the PANI film, that was obtained by the alternate deposition of charged polysaccharides (carboxymethylpullulan (CMP) and chitosan (CHI)). In the latter case, the urease (Urs) enzyme was either physically adsorbed or covalently grafted to the LbL film using carbodiimide coupling reaction. Potentiometric responses of the enzymatic biosensors were measured as a function of the urea concentration in aqueous solutions (from 10(-6) to 10(-1) mol L(-1) urea). Very high sensitivity and short response time were observed for the present biosensor. Moreover, a stability study showed a higher stability over time for the potentiometric response of the sensor with the enzyme-grafted LbL film, testifying for the protective nature of the polysaccharide coating and the interest of covalent grafting.     

Tuberculosis Exacerbates HIV-1 Infection through IL-10/STAT3-Dependent Tunneling Nanotube Formation in Macrophages

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Heating and microwave assisted SPPS of C-terminal acid peptides on trityl resin: the truth behind the yield

Despite correct purity of crude peptides prepared on trityl resin by Fmoc/tBu microwave assisted solid phase peptide synthesis, surprisingly, lower yields than those expected were obtained while preparing C-terminal acid peptides. This could be explained by cyclization/cleavage through diketopiperazine formation during the second amino acid deprotection and third amino acid coupling. However, we provide here evidence that this is not the case and that this yield loss was due to high temperature promoted hydrolysis of the 2-chlorotrityl ester, yielding premature cleavage of the C-terminal acid peptides.     

Global Analysis of Apicomplexan Protein S‐Acyl Transferases Reveals an Enzyme Essential for Invasion

The advent of techniques to study palmitoylation on a whole proteome scale has revealed that it is an important reversible modification that plays a role in regulating multiple biological processes. Palmitoylation can control the affinity of a protein for lipid membranes, which allows it to impact protein trafficking, stability, folding, signalling and interactions. The publication of the palmitome of the schizont stage of Plasmodium falciparum implicated a role for palmitoylation in host cell invasion, protein export and organelle biogenesis. However, nothing is known so far about the repertoire of protein S‐acyl transferases (PATs) that catalyse this modification in Apicomplexa. We undertook a comprehensive analysis of the repertoire of Asp‐His‐His‐Cys cysteine‐rich domain (DHHC‐CRD) PAT family in Toxoplasma gondii and Plasmodium berghei by assessing their localization and essentiality. Unlike functional redundancies reported in other eukaryotes, some apicomplexan‐specific DHHCs are essential for parasite growth, and several are targeted to organelles unique to this phylum. Of particular interest is DHHC7, which localizes to rhoptry organelles in all parasites tested, including the major human pathogen P. falciparum. TgDHHC7 interferes with the localization of the rhoptry palmitoylated protein TgARO and affects the apical positioning of the rhoptry organelles. This PAT has a major impact on T. gondii host cell invasion, but not on the parasite's ability to egress.