Biases and Power for Groups Comparison on Subjective Health Measurements

Subjective health measurements are increasingly used in clinical research, particularly for patient groups comparisons. Two main types of analytical strategies can be used for such data: so-called classical test theory (CTT), relying on observed scores and models coming from Item Response Theory (IRT) relying on a response model relating the items responses to a latent parameter, often called latent trait. Whether IRT or CTT would be the most appropriate method to compare two independent groups of patients on a patient reported outcomes measurement remains unknown and was investigated using simulations. For CTT-based analyses, groups comparison was performed using t-test on the scores. For IRT-based analyses, several methods were compared, according to whether the Rasch model was considered with random effects or with fixed effects, and the group effect was included as a covariate or not. Individual latent traits values were estimated using either a deterministic method or by stochastic approaches. Latent traits were then compared with a t-test. Finally, a two-steps method was performed to compare the latent trait distributions, and a Wald test was performed to test the group effect in the Rasch model including group covariates. The only unbiased IRT-based method was the group covariate Wald’s test, performed on the random effects Rasch model. This model displayed the highest observed power, which was similar to the power using the score t-test. These results need to be extended to the case frequently encountered in practice where data are missing and possibly informative.     

Blimp1 Prevents Methylation of Foxp3 and Loss of Regulatory T Cell Identity at Sites of Inflammation

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Enantiomers of 4-amino-3-fluorobutanoic acid as substrates for gamma-aminobutyric acid aminotransferase. Conformational probes for GABA binding

Gamma-aminobutyric acid aminotransferase (GABA-AT), a pyridoxal 5'-phosphate dependent enzyme, catalyzes the degradation of the inhibitory neurotransmitter gamma-aminobutyric acid (GABA) to succinic semialdehyde with concomitant conversion of pyridoxal 5'-phosphate (PLP) to pyridoxamine 5'-phosphate (PMP). The enzyme then catalyzes the conversion of alpha-ketoglutarate to the excitatory neurotransmitter L-glutamate. Racemic 4-amino-3-fluorobutanoic acid (3-F-GABA) was shown previously to act as a substrate for GABA-AT, not for transamination, but for HF elimination. Here we report studies of the reaction catalyzed by GABA-AT on (R)- and (S)-3-F-GABA. Neither enantiomer is a substrate for transamination. Very little elimination from the (S)-enantiomer was detected using a coupled enzyme assay; The rate of elimination of HF from the (R)-enantiomer is at least 10 times greater than that for the (S)-enantiomer. The (R)-enantiomer is about 20 times more efficient as a substrate for GABA-AT catalyzed HF elimination than GABA is a substrate for transamination. The (R)-enantiomer also inhibits the transamination of GABA 10 times more effectively than the (S)-enantiomer. Using a combination of computer modeling and the knowledge that vicinal C-F and C-NH3+ bonds have a strong preference to align gauche rather than anti to each other, it is concluded that on binding of free 3-F-GABA to GABA-AT the optimal conformation places the C-NH3+ and C-F bonds gauche in the (R)-enantiomer but anti in the (S)-enantiomer. Furthermore, the dynamic binding process and the bioactive conformation of GABA bound to GABA-AT have been inferred on the basis of the different biological behavior of the two enantiomers of 3-F-GABA when they bind to the enzyme. The present study suggests that the C-F bond can be utilized as a conformational probe to explore the dynamic binding process and provide insight into the bioactive conformation of substrates, which cannot be easily determined by other biophysical approaches.     

Analytical biochemistry of DNA–protein assemblies from crude cell extracts

Purification of specific DNA–protein complexes is a challenging task, as the involved interactions can be both electrostatic/H-bond and hydrophobic. The chromatographic stringency needed to obtain reasonable purifications uses salts and detergents. However, these components elicit the removal of proteins unspecifically bound to the chromatographic support itself, thus contaminating the purification products. In this work, a photocleavable linker connected the target oligonucleotidic sequence to the chromatographic beads so as to allow the irradiation-based release of the purified DNA–protein complexes off the beads. Our bioanalytical conditions were validated by purifying the tetracycline repressor protein onto a specific oligonucleotide. The purification factor was unprecedented, with a single contaminant. The robustness of our method was challenged by applying it to the purification of multiprotein assemblies forming onto DNA damage-mimicking oligonucleotides. The purified components were identified as well-known DNA repair proteins, and were shown to retain their enzymatic activities, as seen by monitoring DNA ligation products. Remarkably, kinase activities, also monitored, were found to be distinct on the beads and on the purified DNA–protein complexes, showing the benefits to uncouple the DNA–protein assemblies from the beads for a proper understanding of biochemical regulatory mechanisms involved in the DNA–protein assemblies.     

Analytical biochemistry of DNA--protein assemblies from crude cell extracts

Purification of specific DNA-protein complexes is a challenging task, as the involved interactions can be both electrostatic/H-bond and hydrophobic. The chromatographic stringency needed to obtain reasonable purifications uses salts and detergents. However, these components elicit the removal of proteins unspecifically bound to the chromatographic support itself, thus contaminating the purification products. In this work, a photocleavable linker connected the target oligonucleotidic sequence to the chromatographic beads so as to allow the irradiation-based release of the purified DNA-protein complexes off the beads. Our bioanalytical conditions were validated by purifying the tetracycline repressor protein onto a specific oligonucleotide. The purification factor was unprecedented, with a single contaminant. The robustness of our method was challenged by applying it to the purification of multiprotein assemblies forming onto DNA damage-mimicking oligonucleotides. The purified components were identified as well-known DNA repair proteins, and were shown to retain their enzymatic activities, as seen by monitoring DNA ligation products. Remarkably, kinase activities, also monitored, were found to be distinct on the beads and on the purified DNA-protein complexes, showing the benefits to uncouple the DNA-protein assemblies from the beads for a proper understanding of biochemical regulatory mechanisms involved in the DNA-protein assemblies.     

Ecdysteroids from the medicinal fern Microsorum scolopendria (Burm. f.)

Fronds of the fern Microsorum scolopendria are widely used in traditional medicine in the Society Islands. They were investigated for the presence of ecdysteroids, which might be responsible for at least some of their medicinal properties. M. scolopendria represents an excellent source of ecdysone (0.16% of dry weight) and 20‐hydroxyecdysone (0.20%), and also contains significant amounts (0.01–0.02%) of makisterones A and C, inokosterone and amarasterone A, together with lower amounts of poststerone and of a compound tentatively identified as 24,28‐diepi‐cyasterone. During this study, three new minor phytoecdysteroids, namely 20‐deoxymakisterone A, a 25(?)‐epimer of amarasterone A and 25‐deoxyecdysone 22‐glucoside were also isolated by a combination of normal‐ and reversed‐phase HPLC and subsequently identified by NMR. Copyright © 2007 John Wiley & Sons, Ltd.     

Distribution and ecology of the generalist lactic acid bacterium Carnobacterium maltaromaticum in different freshwater habitats: Metabolic and antagonistic abilities

We explored the distribution, metabolic and antagonistic activities of Carnobacterium maltaromaticum, isolated from freshwater locations in Denmark during winter or early spring. This species was widely distributed in such habitats although it was relatively rare in low pH locations. Isolates possessed a diverse metabolism, potentially enabling functional capacities independent of habitat. The intraspecies competition showed a relatively high degree of mostly low-intensity interactions, which overall were not correlated with phylogeny or location. Only a few isolates exhibited broad-spectrum inhibition activity, targeting species from other genera and families, including one isolate that exhibited a broad inhibitory activity due to H₂O₂ production. Bioinformatic analyses revealed that the frequency of bacteriocinogenic systems was low, and only one unmodified bacteriocin, piscicolin 126, correlated with phenotypic antagonistic activity. Furthermore, most potential bacteriocin gene complexes were not complete. Overall, this study showed C. maltaromaticum to be a generalist (nomadic) species with a constant presence in freshwater habitats, especially those with pH values >5. General metabolic properties did not suggest a strong degree of adaptation to the freshwater environment, and bacteriocin-mediated antagonistic activities appeared to play a minimal ecological role.     

Kite aerial photography: a low‐cost method for monitoring seabird colonies

Obtaining aerial high‐resolution images of bird nesting colonies using remote‐sensing technology such as satellite‐based remote sensing, manned aircraft, or Unmanned Aerial Vehicles might not be possible for many researchers due to financial constraints. Kite Aerial Photography (KAP) provides a possible low‐cost alternative. We collected digital images of ground‐nesting seabirds (i.e., cormorants and penguins) in two different ecosystems using a kite‐based platform equipped with consumer‐grade digital cameras with time‐lapse capability to obtain estimates of breeding population size. KAP proved to be an efficient method for acquiring high‐resolution aerial images. We obtained images of colonies of seabirds ranging in size from hundreds to several hundreds of thousands breeding pairs during flights lasting from a few minutes up to three hours, from flat to very steep areas, and in contrasted wind conditions (from 0.5 to 6 Beaufort force). KAP is an efficient low‐cost method for acquiring high‐resolution aerial images and an alternative to ground‐based censuses, especially useful in rugged areas.     

Dental Microwear Texture Analysis of Varswater Bovids and Early Pliocene Paleoenvironments of Langebaanweg,Western Cape Province,South Africa

The extensive early Pliocene mammalian assemblages at Langebaanweg hold the potential to provide important information about paleoenvironments of the southwestern tip of Africa, an area that today consititutes the Fynbos Biome. We here add to a growing body of literature on the paleoenviornments of the site with an examination of dental microwear textures of bovids from the Varswater Formation. Microwear texture analysis is a new, automated and repeatable approach that measures whole surfaces in three dimensions without observer error. A study of extant ruminants indicates that grazers have more anisotropic microwear surface textures, whereas browsers have more complex microwear surface textures. Fossil bovids recovered from the Muishond Fontein Pelletal Phosphorite Member vary in their microwear textures, with some taxa falling within the extant browser range, some closer to extant grazers, and others in between. These results are consistent with scenarios suggesting mosaic habitats including fynbos vegetation, some (probably C₃) grasses, and woodland elements when these fossils were accumulated.