Eggshell pigment composition covaries with phylogeny but not with life history or with nesting ecology traits of British passerines

No single hypothesis is likely to explain the diversity in eggshell coloration and patterning across birds, suggesting that eggshell appearance is most likely to have evolved to fulfill many nonexclusive functions. By controlling for nonindependent phylogenetic associations between related species, we describe this diversity using museum eggshells of 71 British breeding passerine species to examine how eggshell pigment composition and concentrations vary with phylogeny and with life-history and nesting ecology traits. Across species, concentrations of biliverdin and protoporphyrin, the two main pigments found in eggshells, were strongly and positively correlated, and both pigments strongly covaried with phylogenetic relatedness. Controlling for phylogeny, cavity-nesting species laid eggs with lower protoporphyrin concentrations in the shell, while higher biliverdin concentrations were associated with thicker eggshells for species of all nest types. Overall, these relationships between eggshell pigment concentrations and the biology of passerines are similar to those previously found in nonpasserine eggs, and imply that phylogenetic dependence must be considered across the class in further explanations of the functional significance of avian eggshell coloration

A modified gas-trapping method for high-throughput metabolic experiments in Drosophila melanogaster

Metabolism is often studied in animal models, with the Drosophila melanogaster fruit fly model offering ease of genetic manipulation and high-throughput studies. Fly metabolism is typically studied using end-point assays that are simple but destructive, and do not provide information on the utilization of specific nutrients. To address these limitations, we adapted existing gas-trapping protocols to measure the oxidation of radiolabeled substrates (such as glucose) in multi-well plates. This protocol is cost effective, simple, and offers precise control over experimental diet and measurement time, thus being amenable to high-throughput studies. Furthermore, it is nondestructive, enabling time-course experiments and multiplexing with other parameters. Overall, this protocol is useful for merging fly genetics with metabolic studies to understand whole organism responses to different macronutrients

Communicating serum chemical concentrations to study participants: follow up survey

<p>Abstract</p> <p>Background</p> <p>A considerable literature now supports the importance of effective communication with study participants, including how best to develop communication plans focusing on the uncertainty of health risks associated with particular environmental exposures. Strategies for communicating individual concentrations of environmental chemicals in human biological samples in the absence of clearly established safe or hazardous levels have been discussed from a conceptual basis and to a lesser extent from an empirical basis. We designed and evaluated an empirically based communication strategy for women of reproductive age who previously participated in a prospective study focusing on persistent environmental chemicals and reproductive outcomes.</p> <p>Methods</p> <p>A cohort of women followed from preconception through pregnancy or up to 12 menstrual cycles without pregnancy was given their individual serum concentrations for lead, dichloro-2,2-bis<it>p</it>-chlorophenyl ethylene, and select polychlorinated biphenyl congeners. Two versions of standardized letters were prepared depending upon women's exposure status, which was characterized as low or high. Letters included an introduction, individual concentrations, population reference values and guidance for minimizing future exposures. Participants were actively monitored for any questions or concerns following receipt of letters.</p> <p>Results</p> <p>Ninety-eight women were sent letters informing them of their individual concentrations to select study chemicals. None of the 89 (91%) participating women irrespective of exposure status contacted the research team with questions or concerns about communicated exposures despite an invitation to do so.</p> <p>Conclusions</p> <p>Our findings suggest that study participants can be informed about their individual serum concentrations without generating unnecessary concern.</p

Inhibition of Rho Kinase Regulates Specification of Early Differentiation Events in P19 Embryonal Carcinoma Stem Cells

The Rho kinase pathway plays a key role in many early cell/tissue determination events that take place in embryogenesis. Rho and its downstream effector Rho kinase (ROCK) play pivotal roles in cell migration, apoptosis (membrane blebbing), cell proliferation/cell cycle, cell-cell adhesion and gene regulation. We and others have previously demonstrated that inhibition of ROCK blocks endoderm differentiation in embryonal carcinoma stem cells, however, the effect of ROCK inhibition on mesoderm and ectoderm specification has not been fully examined. In this study, the role of ROCK within the specification and differentiation of all three germ layers was examined.P19 cells were treated with the specific ROCK inhibitor Y-27623, and increase in differentiation efficiency into neuro-ectodermal and mesodermal lineages was observed. However, as expected a dramatic decrease in early endodermal markers was observed when ROCK was inhibited. Interestingly, within these ROCK-inhibited RA treated cultures, increased levels of mesodermal or ectodermal markers were not observed, instead it was found that the pluripotent markers SSEA-1 and Oct-4 remained up-regulated similar to that seen in undifferentiated cultures. Using standard and widely accepted methods for reproducible P19 differentiation into all three germ layers, an enhancement of mesoderm and ectoderm differentiation with a concurrent loss of endoderm lineage specification was observed with Y-27632 treatment. Evidence would suggest that this effect is in part mediated through TGF-β and SMAD signaling as ROCK-inhibited cells displayed aberrant SMAD activation and did not return to a 'ground' state after the inhibition had been removed.Given this data and the fact that only a partial rescue of normal differentiation capacity occurred when ROCK inhibition was alleviated, the effect of ROCK inhibition on the differentiation capacity of pluripotent cell populations should be further examined to elucidate the role of the Rho-ROCK pathway in early cellular 'fate' decision making processes

Insulin Tolerance Test under Anaesthesia to Measure Tissue-specific Insulin-stimulated Glucose Disposal.

Insulin resistance is a pathophysiological state defined by impaired responses to insulin and is a risk factor for several metabolic diseases, most notably type 2 diabetes. Insulin resistance occurs in insulin target tissues including liver, adipose and skeletal muscle. Methods such as insulin tolerance tests and hyperinsulinaemic-euglycaemic clamps permit assessment of insulin responses in specific tissues and allow the study of the progression and causes of insulin resistance. Here we detail a protocol for assessing insulin action in adipose and muscle tissues in anesthetized mice administered with insulin intravenously

Activation of AMPK by bitter melon triterpenoids involves CaMKKβ

We recently showed that bitter melon-derived triterpenoids (BMTs) activate AMPK and increase GLUT4 translocation to the plasma membrane in vitro, and improve glucose disposal in insulin resistant models in vivo. Here we interrogated the mechanism by which these novel compounds activate AMPK, a leading anti-diabetic drug target. BMTs did not activate AMPK directly in an allosteric manner as AMP or the Abbott compound (A-769662) does, nor did they activate AMPK by inhibiting cellular respiration like many commonly used anti-diabetic medications. BMTs increased AMPK activity in both L6 myotubes and LKB1-deficient HeLa cells by 20-35%. Incubation with the CaMKKß inhibitor, STO-609, completely attenuated this effect suggesting a key role for CaMKKß in this activation. Incubation of L6 myotubes with the calcium chelator EGTA-AM did not alter this activation suggesting that the BMT-dependent activation was Ca2+-independent. We therefore propose that CaMKKß is a key upstream kinase for BMT-induced activation of AMPK

A simple algorithm for quantifying DNA methylation levels on multiple independent CpG sites in bisulfite genomic sequencing electropherograms

DNA methylation at cytosines is a widely studied epigenetic modification. Methylation is commonly detected using bisulfite modification of DNA followed by PCR and additional techniques such as restriction digestion or sequencing. These additional techniques are either laborious, require specialized equipment, or are not quantitative. Here we describe a simple algorithm that yields quantitative results from analysis of conventional four-dye-trace sequencing. We call this method Mquant and we compare it with the established laboratory method of combined bisulfite restriction assay (COBRA). This analysis of sequencing electropherograms provides a simple, easily applied method to quantify DNA methylation at specific CpG sites