Genetic studies of the leptin receptor gene in morbidly obese French Caucasian families

Family studies have shown that in some populations up to 75% of the variation of body mass index can be explained by genetic factors. However, in humans, no major obesity gene has been identified to date. In contrast, there are a number of genetically well defined animal models for obesity. In two of those models (ob/ob and db/db), defects in the same pathway are responsible for obesity. Recently, some evidence has been found for the OB gene also being involved in human obesity. In this study we investigated the potential role of the OB receptor (OBR) in the etiology of massive obesity in humans using familial linkage analyses and case-control association studies. The typing of two microsatellite markers (D1S198 and D1S209), flanking the OBR gene, in 256 sib pairs showed no evidence for linkage with obesity. In order to be able to detect small gene effects, association studies with a 3′-UTR insertion/deletion polymorphism were carried out. The results of these analyses remained non-significant (χ² = 3.442, P = 0.18). However, subjects heterozygous for the insertion/deletion polymorphism showed a slight trend towards lower insulin values 30 min after an oral glucose load compared to homozygous individuals (P = 0.02). In summary, our results do not support a major role of the human OBR gene in the development of morbid obesity in our population. Received: 4 December 1996 / Accepted: 25 June 1997     

Structural characterization of canine PYY

PYY was purified from canine colonic mucosa by sequential steps of reverse phase HPLC and ion-exchange FPLC. Microsequence, amino acid and mass spectral analyses of the purified peptide and its tryptic fragments were consistent with the structure: YPAKPEAPGEDASPEELSRYYASLRHYLNLVTRQRY-amide. Canine PYY(1-36) has the identical sequence as porcine and rat PYY but differs from human PYY at position 3, with Ala instead of Ile, and position 18, with Ser instead of Asn. A smaller form, PYY(3-36), was also purified and characterized. It may differ in its biological activity from the intact peptide and could act as a partial antagonist or agonist of PYY(1-36).     

Hyperosmosis enhances radiation and hydroxyurea resistance of Schizosaccharomyces pombe checkpoint mutants through the spindle checkpoint and delayed cytokinesis

The DNA damage and stress response pathways interact to regulate cellular responses to genotoxins and environmental stresses. How these pathways interact in Schizosaccharomyces pombe is not well understood. We demonstrate that osmotic stress suppresses the DNA damage sensitivity of checkpoint mutants, and that this occurs through three distinct cell cycle delays. A delay in G2/M is dependent on Srk1. Progression through mitosis is halted by the Mad2‐dependent spindle checkpoint. Finally, cytokinesis is impaired by modulating Cdc25 expression. These three delays, imposed by osmotic stress, together compensate for the loss of checkpoint signalling.     

Localization of a non-Mendelian gene in Chlamydomonas reinhardii

Summary Transfer of a non-Mendelian neamine-dependent (nd) mutant to an antibioticfree medium results in neamine-sensitive and neamine-resistent revertants. These reversions are caused by extranuclear mutations.The neamine-sensitive revertants are no more able to split offnd-cells after back-donation to neamine containing medium. Therefore they are different from the streptomycin-sensitive revertants of a streptomycin-dependent (sd) mutant. These mutants were capable ofsd-segregation though their potence ofsd-segregation diminished on antibiotic-free medium with increasing time of cultivation.The different behaviour can be explained by the fact that manysd-genes are present which have to be appointed to the mitochondria. On the other side, thend-gene exists only in few copies and is located therefore in the chloroplast.Several experiments with differing methods are discussed to localize the extranuclear genes.

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Vorgelegt durch G. Melchers     

New Approaches for Absolute Quantification of Stable‐Isotope‐Labeled Peptide Standards for Targeted Proteomics Based on a UV Active Tag

Targeted proteomics depends on the availability of stable isotope labeled (SIL) peptide standards, which for absolute protein quantification need to be absolutely quantified. In the present study, three new approaches for absolute quantification of SIL peptides are developed. All approaches rely on a quantification tag (Qtag) with a specific UV absorption. The Qtag is attached to the peptide during synthesis and is removed by tryptic digestion under standard proteomics workflow conditions. While one quantification method (method A) is designed to allow the fast and economic production of absolutely quantified SIL peptides, two other methods (methods B and C) are developed to enable the straightforward re‐quantification of SIL peptides after reconstitution to control and monitor known problems related to peptide solubility, precipitation, and adhesion to vials. All methods yield consistent results when compared to each other and when compared to quantification by amino acid analysis. The precise quantitation methods are used to characterize the in vivo specificity of the H3 specific histone methyltransferase EZH2.     

Aerial visual acuity in harbor seals (<Emphasis Type="Italic">Phoca vitulina</Emphasis>) as a function of luminance

In this study, we measured aerial visual acuity in harbor seals. As a first approach to the hypothesis that harbor seals can obtain acute aerial visual acuity mediated by the interaction of the vertical slit-shaped pupil and the corneal flattening although refractive measurements had revealed aerial myopia, visual acuity was tested as a function of luminance and pupil dilation. We analyzed aerial visual acuity (minimal resolvable stripe width) in three harbor seals in a two-alternative-forced-choice discrimination experiment. Our results further support the hypothesis that harbor seals possess an aerial visual acuity comparable to the acuity in clear waters if the vertical slit pupil does not exceed the zone of corneal flattening in bright light. When the pupil dilates with decreasing luminance, visual acuity decreases which might be due to deflected light from the stronger curved peripheral cornea.     

Separation of carvedilol enantiomers in very small volumes of human plasma by capillary electrophoresis with laser-induced fluorescence

A sensitive capillary electrophoretic method for the determination of carvedilol enantiomers in 100 μl of human plasma has been developed and validated. Carvedilol and the internal standard carazolol are isolated from plasma samples by liquid–liquid extraction using diethylether. A sensitive and selective detection is provided by helium–cadmium laser-induced fluorescence. The total analysis time is 17.5 min, about 30 min are needed for the sample preparation. The linearity of the assay ranges from 1.56 to 50 ng/ml per carvedilol enantiomer. The limits of quantification (LOQ) for the carvedilol enantiomers in 100 μl of human plasma are 1.56 ng/ml. The inter-day accuracy for R-carvedilol is between 95.8 and 103% (104% at LOQ) and for S-carvedilol between 97.1 and 103% (107% at LOQ); the inter-day precision values are between 3.81 and 8.64% (10.9% at LOQ) and between 5.47 and 7.86% (7.91% at LOQ) for R- and S-carvedilol, respectively. The small sample volume needed is especially advantageous for the application in clinical studies in pediatric patients. As an application of the assay concentration/time profiles of the carvedilol enantiomers in a 5-year-old patient receiving a test dose of 0.09 mg/kg carvedilol are reported.     

German Translation and Validation of the Cognitive Style Questionnaire Short Form (CSQ-SF-D)

Background

The Cognitive Style Questionnaire is a valuable tool for the assessment of hopeless cognitive styles in depression research, with predictive power in longitudinal studies. However, it is very burdensome to administer. Even the short form is still long, and neither this nor the original version exist in validated German translations.

Methods

The questionnaire was translated from English to German, back-translated and commented on by clinicians. The reliability, factor structure and external validity of an online form of the questionnaire were examined on 214 participants. External validity was measured on a subset of 90 subjects.

Results

The resulting CSQ-SF-D had good to excellent reliability, both across items and subscales, and similar external validity to the original English version. The internality subscale appeared less robust than other subscales. A detailed analysis of individual item performance suggests that stable results could be achieved with a very short form (CSQ-VSF-D) including only 27 of the 72 items.

Conclusions

The CSQ-SF-D is a validated and freely distributed translation of the CSQ-SF into German. This should make efficient assessment of cognitive style in German samples more accessible to researchers.