Repeated inhalations of diesel exhaust particles and oxidatively damaged DNA in young oxoguanine DNA glycosylase (OGG1) deficient mice

DNA repair may prevent increased levels of oxidatively damaged DNA from prolonged oxidative stress induced by, e.g. exposure to diesel exhaust particles (DEP). We studied oxidative damage to DNA in broncho-alveolar lavage cells, lungs, and liver after 4 × 1.5 h inhalations of DEP (20 mg/m³) in Ogg1^- / -  and wild type (WT) mice with similar extent of inflammation. DEP exposure increased lung levels of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) in Ogg1^- / -  mice, whereas no effect on 8-oxodG or oxidized purines in terms of formamidopyrimidine DNA glycosylase (FPG) sites was observed in WT mice. In both unexposed and exposed Ogg1^- / -  mice the level of FPG sites in the lungs was 3-fold higher than in WT mice. The high basal level of FPG sites in Ogg1^- / -  mice probably saturated the assay and prevented detection of DEP-generated damage. In conclusion, Ogg1^- / -  mice have elevated pulmonary levels of FPG sites and accumulate genomic 8-oxodG after repeated inhalations of DEP.     

Pakistanis living in Oslo have lower serum 1,25-dihydroxyvitamin D levels but higher serum ionized calcium levels compared with ethnic Norwegians. The Oslo Health Study

Background

Clinical determination of mid-parental height is an important part of the assessment of a child's growth, however our clinical impression has been that parents cannot be relied upon to accurately report their own heights. Therefore, we conducted this study to assess the accuracy of parental height self-reporting and its effect on calculated mid-parental target height for children presenting to a pediatric endocrinology office.

Methods

All parents bringing their children for an initial evaluation to a pediatric endocrinology clinic over a period of nine months were questioned and then measured by a pediatric endocrinologist. Parents were blinded to the study. Mid-parental target heights, based on reported and actual height were compared.

Results

There were 241 families: 98 fathers and 217 mothers in our study. Mean measured paternal height was 173.2 cm, self reported 174.9 cm (p < 0.0001), partner reported 177 cm (p = 0.0004). Only 50% of fathers and 58% of mothers reported their height within ± 2 cm of their measured height, while 15% of fathers and 12% of mothers were inaccurate by more than 4 cm. Mean measured maternal height was 160.6 cm, self-reported 161.1 cm (NS), partner reported 161.7 cm (NS). Inaccuracy of height self-report had a small but significant effect on the mean MPTH (0.4 cm, p = 0.045). Analysis showed that only 70% of MPTH calculated by reported heights fell within ± 2 cm of MPTH calculated using measured heights, 24% being in ± 2–4 cm range, and 6% were inaccurate by more than 4 cm.

Conclusion

There is a significant difference in paternal measured versus reported heights with an overall trend for fathers to overestimate their own height. A large subset of parents makes a substantial error in their height self-report, which leads to erroneous MPTH. Inaccuracy is even greater when one parent reports the other parent's height. When a child's growth is in question, measured rather than reported parental heights should be obtained.     

The cell wall binding domain of Listeria bacteriophage endolysin PlyP35 recognizes terminal GlcNAc residues in cell wall teichoic acid

The cell wall binding domains (CBD) of bacteriophage endolysins target the enzymes to their substrate in the bacterial peptidoglycan with extraordinary specificity. Despite strong interest in these enzymes as novel antimicrobials, little is known regarding their interaction with the bacterial wall and their binding ligands. We investigated the interaction of Listeria phage endolysin PlyP35 with carbohydrate residues present in the teichoic acid polymers on the peptidoglycan. Biochemical and genetic analyses revealed that CBD of PlyP35 specifically recognizes the N-acetylglucosamine (GlcNAc) residue at position C4 of the polyribitol-phosphate subunits. Binding of CBDP35 could be prevented by removal of wall teichoic acid (WTA) polymers from cell walls, and inhibited by addition of purified WTAs or acetylated saccharides. We show that Listeria monocytogenes genes lmo2549 and lmo2550 are required for decoration of WTAs with GlcNAc. Inactivation of either gene resulted in a lack of GlcNAc glycosylation, and the mutants failed to bind CBDP35. We also report that the GlcNAc-deficient phenotype of L. monocytogenes strain WSLC 1442 is due to a small deletion in lmo2550, resulting in synthesis of a truncated gene product responsible for the glycosylation defect. Complementation with lmo2550 completely restored display of characteristic serovar 1/2 specific WTA and the wild-type phenotype.     

Short-term and long-term effects of increasing temperatures on the stability and the production of volatile sulfur compounds in full-scale thermophilic anaerobic digesters

This study compares the effect of a rapid increase of the digester temperature (from 54 degrees C to 58 degrees C in 2 weeks) with a slow increase (from 53.9 degrees C to 57.2 degrees C at a rate of 0.55 degrees C per month) on full-scale thermophilic anaerobic digestion at Hyperion Treatment Plant. The short-term test demonstrated that rapidly increasing the digester temperature caused elevated production of volatile sulfur compounds, most notably methyl mercaptan, but volatile solids destruction and methane production were not significantly affected. The increase of the volatile fatty acid to alkalinity ratio from 0.1 to over 0.3 indicated a transient change in digester biochemistry, which was reversed by lowering the temperature. In the long term-test, a slow increase of digester temperature, the production of hydrogen sulfide increased above temperatures of 56.1 degrees C, but was controlled by increased injection of ferrous chloride. Methyl mercaptan was detected in trace amounts at the highest temperature tested (57.2 degrees C). This test showed insignificant effects on other digestion parameters, although some temperature-independent changes were observed that could have been seasonal effects over the year that the long-term test lasted. Thus a slow temperature increase was preferable. This observation contrasts with previous results showing the desirability of a rapid temperature rise to first establish a thermophilic culture when converting from mesophilic operation. Further research is warranted on temperature limits and process changes to optimize thermophilic anaerobic digestion.     

Kinetics of in vivo elimination of suicide gene-expressing T cells affects engraftment, graft-versus-host disease, and graft-versus-leukemia after allogeneic bone marrow transplantation

Suicide gene therapy is one approach being evaluated for the control of graft-vs-host disease (GVHD) after allogeneic bone marrow transplantation (BMT). We recently constructed a novel chimeric suicide gene in which the entire coding region of HSV thymidine kinase (HSV-tk) was fused in-frame to the extracellular and transmembrane domains of human CD34 (DeltaCD34-tk). DeltaCD34-tk is an attractive candidate as a suicide gene in man because of the ensured expression of HSV-tk in all selected cells and the ability to rapidly and efficiently purify gene-modified cells using clinically approved CD34 immunoselection techniques. In this study we assessed the efficacy of the DeltaCD34-tk suicide gene in the absence of extended ex vivo manipulation by generating transgenic animals that express DeltaCD34-tk in the peripheral and thymic T cell compartments using the CD2 locus control region. We found that DeltaCD34-tk-expressing T cells could be purified to near homogeneity by CD34 immunoselection and selectively eliminated ex vivo and in vivo when exposed to low concentrations of GCV. The optimal time to administer GCV after allogeneic BMT with DeltaCD34-tk-expressing transgenic T cells was dependent on the intensity of the conditioning regimen, the leukemic status of the recipient, and the dose and timing of T cell infusion. Importantly, we used a controlled graft-vs-host reaction to promote alloengraftment in sublethally irradiated mice and provide a graft-vs-leukemia effect in recipients administered a delayed infusion of DeltaCD34-tk-expressing T cells. This murine model demonstrates the potential usefulness of DeltaCD34-tk-expressing T cells to control GVHD, promote alloengraftment, and provide a graft-vs-leukemia effect in man.     

Insights into type IV pilus biogenesis and dynamics from genetic analysis of a C-terminally tagged pilin: a role for O-linked glycosylation

Type IV pili are surface organelles essential for pathogenicity of many Gram-negative bacteria. In Neisseria gonorrhoeae, the major subunit of type IV pili, PilE, is a target of its general O-linked glycosylation system. This system modifies a diverse set of periplasmic and extracellular gonococcal proteins with a variable set of glycans. Here we show that expression of a particular hexa-histidine-tagged PilE was associated with growth arrest. By studying intra- and extragenic suppressors, we found that this phenotype was dependent on pilus assembly and retraction. Based on these results, we developed a sensitive tool to identify factors with subtle effects on pilus dynamics. Using this approach, we found that glycan chain length has differential effects on the growth arrest that appears to be mediated at the level of pilin subunit-subunit interactions and bidirectional remodelling of pilin between its membrane-associated and assembled states. Gonococcal pilin glycosylation thus plays both an intracellular role in pilus dynamics and potential extracellular roles mediated through type IV pili. In addition to demonstrating the effect of glycosylation on pilus dynamics, the study provides a new way of identifying factors with less dramatic effects on processes involved in type IV pilus biogenesis.     

Standardization of Misleading Immunoassay Based 25-Hydroxyvitamin D Levels with Liquid Chromatography Tandem-Mass Spectrometry in a Large Cohort Study

Background

The interest in vitamin D measurement has strongly increased in recent years. The best indicator for circulating vitamin D levels is 25-hydroxy-vitamin D (25(OH)D) which is often measured by different immunoassays. We demonstrate problems in comparability of measures by different immunoassays and the need for standardization in the context of a large population-based cohort study.

Methods

25(OH)D was measured with the immunoassays Diasorin Liaison in 2006 in 5,386 women and in the context of another project with IDS-iSYS in 4,199 men in 2009–2010 (when the Diasorin Liaison was no longer available in the version utilized in 2006). Standardization was performed by re-measuring of 25(OH)D levels in 97 men and 97 women with liquid chromatography tandem-mass spectrometry (LC-MS/MS) to obtain linear regression conversion equations.

Results

Applying a 30 nmol/L cut-off value for vitamin D deficiency would have resulted in 48.3% of women and 12.1% of men with vitamin D deficiency ahead of standardization. The large gender difference was strongly attenuated after standardization of the assays with only 15.7% of women and 14.3% of men with vitamin D deficiency. Standardization on average increased the 25(OH)D levels by 10.3 nmol/L in women and decreased 25(OH)D levels by 2.9 nmol/L in men.

Conclusion

The standardization with LC-MS/MS revealed that much of the observed gender difference was only assay-driven and the extremely high proportion of 48.3% vitamin D deficient women proved to be an exaggeration of the old version of the Diasorin-Liaison immunoassay. Standardization of 25(OH)D immunoassay results by LC-MS/MS is recommended to improve their accuracy and comparability, provided the LC-MS/MS method itself is adequately validated and standardized.     

MetaboLights: towards a new COSMOS of metabolomics data management

Exciting funding initiatives are emerging in Europe and the US for metabolomics data production, storage, dissemination and analysis. This is based on a rich ecosystem of resources around the world, which has been build during the past ten years, including but not limited to resources such as MassBank in Japan and the Human Metabolome Database in Canada. Now, the European Bioinformatics Institute has launched MetaboLights, a database for metabolomics experiments and the associated metadata (http://www.ebi.ac.uk/metabolights). It is the first comprehensive, cross-species, cross-platform metabolomics database maintained by one of the major open access data providers in molecular biology. In October, the European COSMOS consortium will start its work on Metabolomics data standardization, publication and dissemination workflows. The NIH in the US is establishing 6?C8 metabolomics services cores as well as a national metabolomics repository. This communication reports about MetaboLights as a new resource for Metabolomics research, summarises the related developments and outlines how they may consolidate the knowledge management in this third large omics field next to proteomics and genomics.     

Screening for antibacterial and antifungal activities in marine benthic invertebrates from northern Norway

Benthic marine invertebrates collected from sub-Arctic regions of northern Norway, were found to be a promising source of novel bioactive compounds against human and fish pathogenic bacteria and fungi. Lyophilized material from seven species of ascidians, six sponges and one soft alcyonid coral were extracted with 60% acidified acetonitrile (ACN). After separation into an ACN-rich phase (ACN-extract) and an aqueous phase, and subsequent solid-phase extraction of the aqueous phase, fractions differing in polarity were obtained and screened for antibacterial and antifungal activities, along with the more lipophilic ACN-extracts. Antimicrobial activity was determined against two Gram-negative, two Gram-positive bacteria, and two strains of fungi. Notably, all the invertebrate species in the study showed activity against all four strains of bacteria and the two strains of fungi. In general, the aqueous fractions displayed highest antimicrobial activity, and the most potent extracts were obtained from the colonial ascidian Synoicum pulmonaria which displayed activity against bacteria and fungi at a concentration of 0.02 mg/ml; the lowest concentration tested.