Nondestructive amino acid analysis at the picomole level of proline-containing peptides using aminopeptidase M and prolidase: application to peptides containing tyrosine sulfate.

A micromethod for nondestructive amino acid analysis was developed. Peptides were digested with amino-peptidase M followed by prolidase. The whole procedure was performed in a microvial for the autosampler of the analyzer. The resulting method has a working range from 10 to 600 pmol, at least, of sample taken into work. The usefulness of the method was demonstrated by its ability to determine the sulfation of tyrosine preceding proline in chicken gastrin.     

Ethnic Differences in Correlates of Obesity between Latin‐American and Black Women

Objective: To date, no studies have examined dietary intake, physical activity, and body image in a large sample of Latin‐American and black women recruited using the same methodology. The aim of this study was to examine three potential correlates of obesity (dietary intake, body image, and physical activity) in a large sample of Latin‐American and black women across the weight spectrum. Research Methods and Procedures: Participants were black (n = 271) and Latin‐American (n = 234) adult women who completed a 24‐hour dietary recall and physical activity and body image questionnaires. Results: After controlling for BMI, education, marital status, and number of children, black women consumed more kilocalories, dietary fat (grams), and percent calories from fat than Latin‐American women, who consumed more carbohydrates (grams) and dietary fiber (total and soluble). Black women engaged in more sedentary behavior than Latin‐American women. Although Latin‐American women weighed less than black women, they perceived their current body image as heavier and reported greater body image dissatisfaction than black women. Black women also reported a higher ideal body image than Latin‐American women. Discussion: The combined effect of a diet higher in calories and fat, increased sedentary behavior, and more accepting body image could account for higher rates of obesity among black women. Future studies should further explore cultural attitudes and beliefs related to weight that could provide information for the development of culturally competent obesity interventions.     

Potent infection reservoir of crayfish plague now permanently established in Norway

Noble crayfish Astacus astacus is threatened in Europe due to invasive crayfish carrying the crayfish plague agent Aphanomyces astaci. Norway is among the last countries in which the introduction of non-indigenous crayfish has been limited through strict legislation practices. However, North American signal crayfish Pacifastacus leniusculus were recently discovered in a water-course that has been repeatedly hit by the plague. We mapped the distribution and relative density (catch per unit effort) of signal crayfish within this lake, and performed agent-specific real-time PCR to estimate the prevalence of A. astaci in the population. The resulting length frequencies and relative density estimates clearly demonstrate a well-established signal crayfish population, in which 86.4% of the analysed individuals were confirmed carriers. The success of detection was significantly higher (84.1%) in the crayfish tailfan (i.e. uropods) than in the soft abdominal cuticle (38.4%), which is commonly used in prevalence studies. We therefore propose tailfan (uropods and telson) as the preferred tissue for studying A. astaci prevalence in signal crayfish populations. The likelihood of detecting an A. astaci-positive signal crayfish increased significantly with increasing crayfish length. Further, large female crayfish expressed significantly higher PCR-forming units values than large males. In surveys primarily exploring the presence of A. astaci-positive individuals in a population, large females should be selected for molecular analyses. Our study demonstrates that a potent crayfish plague infection reservoir, evidently originating from the illegal human introduction of signal crayfish, has permanently been established in Norway.     

Exercise training does not correct abnormal cardiac glycogen accumulation in the db/db mouse model of type 2 diabetes

Substrate imbalance is a well-recognized feature of diabetic cardiomyopathy. Insulin resistance effectively limits carbohydrate oxidation, resulting in abnormal cardiac glycogen accumulation. Aims of the present study were to 1) characterize the role of glycogen-associated proteins involved in excessive glycogen accumulation in type 2 diabetic hearts and 2) determine if exercise training can attenuate abnormal cardiac glycogen accumulation. Control (db(+)) and genetically diabetic (db/db) C57BL/KsJ-lepr(db)/lepr(db) mice were subjected to sedentary or treadmill exercise regimens. Exercise training consisted of high-intensity/short-duration (10 days) and low-intensity/long-duration (6 wk) protocols. Glycogen levels were elevated by 35-50% in db/db hearts. Exercise training further increased (2- to 3-fold) glycogen levels in db/db hearts. Analysis of soluble and insoluble glycogen pools revealed no differential accumulation of one glycogen subspecies. Phosphorylation (Ser(640)) of glycogen synthase, an indicator of enzymatic fractional activity, was greater in db/db mice subjected to sedentary and exercise regimens. Elevated glycogen levels were accompanied by decreased phosphorylation (Thr(172)) of 5'-AMP-activated kinase and phosphorylation (Ser(79)) of its downstream substrate acetyl-CoA carboxylase. Glycogen concentration was not associated with increases in other glycogen-associated proteins, including malin and laforin. Novel observations show that exercise training does not correct diabetes-induced elevations in cardiac glycogen but, rather, precipitates further accumulation.     

Proof of the Concept to Use a Malignant B Cell Line Drug Screen Strategy for Identification and Weight of Melphalan Resistance Genes in Multiple Myeloma

In a conceptual study of drug resistance we have used a preclinical model of malignant B-cell lines by combining drug induced growth inhibition and gene expression profiling. In the current report a melphalan resistance profile of 19 genes were weighted by microarray data from the MRC Myeloma IX trial and time to progression following high dose melphalan, to generate an individual melphalan resistance index. The resistance index was subsequently validated in the HOVON65/GMMG-HD4 trial data set to prove the concept. Biologically, the assigned resistance indices were differentially distributed among translocations and cyclin D expression classes. Clinically, the 25% most melphalan resistant, the intermediate 50% and the 25% most sensitive patients had a median progression free survival of 18, 32 and 28 months, respectively (log-rank P-value  = 0.05). Furthermore, the median overall survival was 45 months for the resistant group and not reached for the intermediate and sensitive groups (log-rank P-value  = 0.003) following 38 months median observation. In a multivariate analysis, correcting for age, sex and ISS-staging, we found a high resistance index to be an independent variable associated with inferior progression free survival and overall survival. This study provides clinical proof of concept to use in vitro drug screen for identification of melphalan resistance gene signatures for future functional analysis.     

Antiparasite treatments reduce humoral immunity and impact oxidative status in raptor nestlings

Parasites are natural stressors that may have multiple negative effects on their host as they usurp energy and nutrients and may lead to costly immune responses that may cause oxidative stress. At early stages, animals may be more sensitive to infectious organisms because of their rapid growth and partly immature immune system. The objective of this study was to explore effects of parasites by treating chicks of two raptor species (northern goshawk Accipiter gentilis and white‐tailed sea eagle Haliaeetus albicilla) against both endoparasites (internal parasites) and ectoparasites (external parasites). Nests were either treated against ectoparasites by spraying with pyrethrin or left unsprayed as control nests. Within each nest, chicks were randomly orally treated with either an antihelminthic medication (fenbendazole) or sterile water as control treatment. We investigated treatment effects on plasma (1) total antioxidant capacity TAC (an index of nonenzymatic circulating antioxidant defenses), (2) total oxidant status TOS (a measure of plasmatic oxidants), and (3) immunoglobulin levels (a measure of humoral immune function). Treatment against ectoparasites led to a reduction in circulating immunoglobulin plasma levels in male chicks. TOS was higher when not receiving any parasite reduction treatment and when receiving both endo‐ and ectoparasitic reduction treatment compared with receiving only one treatment. TAC was higher in all treatment groups, when compared to controls. Despite the relatively low sample size, this experimental study suggests complex but similar relationships between treatment groups and oxidative status and immunoglobulin levels in two raptor species.     

Carboxamide SIRT1 inhibitors block DBC1 binding via an acetylation-independent mechanism

SIRT1 is an NAD⁺-dependent deacetylase that counteracts multiple disease states associated with aging and may underlie some of the health benefits of calorie restriction. Understanding how SIRT1 is regulated in vivo could therefore lead to new strategies to treat age-related diseases. SIRT1 forms a stable complex with DBC1, an endogenous inhibitor. Little is known regarding the biochemical nature of SIRT1-DBC1 complex formation, how it is regulated and whether or not it is possible to block this interaction pharmacologically. In this study, we show that critical residues within the catalytic core of SIRT1 mediate binding to DBC1 via its N-terminal region, and that several carboxamide SIRT1 inhibitors, including EX-527, can completely block this interaction. We identify two acetylation sites on DBC1 that regulate its ability to bind SIRT1 and suppress its activity. Furthermore, we show that DBC1 itself is a substrate for SIRT1. Surprisingly, the effect of EX-527 on SIRT1-DBC1 binding is independent of DBC1 acetylation. Together, these data show that protein acetylation serves as an endogenous regulatory mechanism for SIRT1-DBC1 binding and illuminate a new path to developing small-molecule modulators of SIRT1.     

Pigmentation and spectral absorbance in the deep-sea arctic amphipods Eurythenes gryllus and Anonyx sp.

As for many deep-sea animals, the red colouration of the two amphipods Eurythenes gryllus and Anonyx sp. has an important function providing camouflage, as the attenuation of the red wavelengths in seawater is higher than other colours within the visible range. Variation in colouration between different stages of colour intensity (related to size) is evident in both species. The red colour is caused by carotenoids, and the carotenoid composition was identified and quantified using spectral optical density signatures, high-performance liquid chromatography (HPLC) and liquid chromatography–mass spectrometry (LC–MS). The carotenoid astaxanthin was identified as the major carotenoid in both amphipods, both in pure and in esterified forms. In addition, minor amounts of lutein-like, canthaxanthin-like and several unidentified carotenoids were found in E. gryllus, while diatoxanthin, β,β-carotene and canthaxanthin-like carotenoids were detected in Anonyx sp. Generally, both species displayed an increase in the amount of carotenoids as a function of colour intensity and size. Shifts in λ_max in the OD (Optical density; dimensionless, acronym absorbance) spectra were evident in both species between the different colour stages in both the in vivo and the in vitro material, probably caused by changes in pigment composition. Similar shifts in λ_max were observed between the in vivo and in vitro pigment raw extracts in general, most likely caused by pigment-binding proteins. The differences in pigment composition and wavelength shifts suggest large intra- and inter-specific differences between the two species. Probable reasons for changes in pigment composition could be related to diet, season, moulting patterns, metabolic pathways and reproduction.