Simulation of Nanoparticle Permeation through a Lipid Membrane

A metric of nanoparticle toxicity is the passive permeability rate through cellular membranes. To assess the influence of nanoparticle morphology on this process, the permeability of buckyball-sized molecules through a representative lipid bilayer was investigated by molecular-dynamics simulation. When C₆₀ was compared with a prototypical opened C₆₀ molecule and a representative combustion-generated particle, C₆₈H₂₉, the calculated free-energy profiles along the permeation coordinate revealed a sizable variation in form and depth. The orientation of the anisotropic molecules was determined by monitoring the principal axis corresponding to the largest moment of inertia, and free rotation was shown to be hindered in the bilayer interior. Diffusion constant values of the permeant molecules were calculated from a statistical average of seven to 10 trajectories at five locations along the permeation coordinate. A relatively minor variation of the values was observed in the bilayer interior; however, local resistance values spanned up to 24 orders of magnitude from the water layer to the bilayer center, due primarily to its exponential dependence on free energy. The permeability coefficient values calculated for the three similarly sized but structurally distinct nanoparticles showed a significant variance. The use of C₆₀ to represent similarly sized carbonaceous nanoparticles for assessments of toxicity is questioned.     

Transposon-induced mutations in two loci of Listeria monocytogenes serotype 1/2a result in phage resistance and lack of N-acetylglucosamine in the teichoic acid of the cell wall.

Teichoic acid-associated N-acetylglucosamine and rhamnose have been shown to serve as phage receptors in Listeria monocytogenes serotype 1/2a. We generated and characterized two single-copy Tn916DeltaE mutants which were resistant to phage A118 and several other serotype 1/2a-specific phages. In one mutant the insertion was immediately upstream of the recently identified ptsHI locus, which encodes two proteins of the phosphoenolpyruvate-dependent carbohydrate uptake system, whereas in the other the insertion was immediately upstream of an operon whose most distal gene was clpC, involved in stress responses and virulence. Transduction experiments confirmed the association of the phage-resistant phenotype of these mutants with the transposon insertion. Phage A118 resistance of the mutants could be attributed to inability of the phage to adsorb onto the mutant cells, and biochemical analysis of cell wall composition showed that the teichoic acids of both mutants were deficient in N-acetylglucosamine. Rhamnose and other teichoic acid and cell wall components were not affected.     

Breakdown of exogenous insulin by langerhans islets of the pancreas in vitro

Pancreatic islets were isolated from Wistar rats, albino mice, spiny mice and sand rats (Psammomys obesus). Evidence is presented that pancreatic islets contain an enzyme system degrading insulin in the presence of glutathione or other sulfhydryl-containing compounds. Apparent K_m values for insulin and glutathione (in the presence of EDTA) are 14.0 μM (mol. wt 5700) and 1.28 mM, respectively. Maximum breakdown of ¹²⁵I-labeled insulin was found at about pH 7.2. After ultracentrifugation of islet homogenates the microsomal fraction contained the greatest relative specific insulin-degrading activity. The specific insulin-degrading actvitity was found to be higher in Wistar rats and albino mice than in spiny mice and sand rats. Starvation of Wistar rats for 72 h caused a decrease inthe enzymatic activity.     

Fermentation studies of the secretion of Serratia marcescens nuclease by Escherichia coli

The secretion of a Serratia marcescens nuclease was followed by fermentation with Escherichia coli. A plasmid, p403-SD2, carrying a 1.3-kilobase-pair insert with a 0.4-kilobase-pair region upstream of the nuclease gene caused a growth-phase-regulated expression of nuclease in E. coli in the same way as that seen in S. marcescens. Deletion of the regulatory gene generating plasmid p403-Rsa1 resulted in a constitutive expression of the nuclease. Anaerobiosis stimulated the expression from p403-SD2 in stationary growth phase by a factor of 10 compared with expression stimulated by cultivation in aerobic conditions; no such effect was found for plasmid p403-Rsa1. Different nutritional factors caused the expression level and the amount of extracellular nuclease to vary more when nuclease was expressed from plasmid p403-SD2 than when it was expressed from plasmid p403-Rsa1. A correlation between the regulatory gene and the extracellular secretion of nuclease is proposed.     

Ants benefit from attending facultatively myrmecophilous Lycaenidae caterpillars: evidence from a survival study

Workers of three ant species (Lasius niger, Lasius flavus, Myrmica rubra) were caged in the laboratory together with caterpillars and pupae of five species of lycaenid butterflies. Mortality of ants was 3–5 times higher when the ants were confined with larvae lacking a dorsal nectar organ (Lycaena phlaeas, Lycaena tityrus) rather than with caterpillars which possess a nectar gland (Aricia agestis, Polyommatus bellargus, P. icarus). For all five species, ant survival was always lower at the pupal stage (where a nectar organ is always absent) than at the caterpillar stage and was largely equivalent for the butterfly species tested. The experimental data confirm earlier estimates that ants can derive nutritive benefits from tending facultatively myrmecophilous lycaenid caterpillars, even though these caterpillars produce nectarlike secretions at low rates.     

Targeted and global pharmacometabolomics in everolimus-based immunosuppression: association of co-medication and lysophosphatidylcholines with dose requirement

Introduction

The immunosuppressive therapy with everolimus (ERL) after heart transplantation is characterized by a narrow therapeutic window and a substantial variability in dose requirement. Factors explaining this variability are largely unknown.

Objectives

Our aim was to evaluate factors affecting ERL metabolism and to identify novel metabolites associated with the individual ERL dose requirement to elucidate mechanisms underlying ERL dose response variability.

Method

We used liquid chromatography coupled with mass spectrometry for quantification of ERL metabolites in 41 heart transplant patients and evaluated the effect of clinical and genetic factors on ERL pharmacokinetics. Non-targeted plasma metabolic profiling by ultra-performance liquid chromatography and high resolution quadrupole-time-of-flight mass spectrometry was used to identify novel metabolites associated with ERL dose requirement.

Results

The determination of ERL metabolites revealed differences in metabolite patterns that were independent from clinical or genetic factors. Whereas higher ERL dose requirement was associated with co-administration of sodium-mycophenolic acid and the CYP3A5 expressor genotype, lower dose was required for patients receiving vitamin K antagonists. Global metabolic profiling revealed several novel metabolites associated with ERL dose requirement. One of them was identified as lysophosphatidylcholine (lysoPC) (16:0/0:0). Subsequent targeted analysis revealed that high levels of several lysoPCs were significantly associated with higher ERL dose requirement.

Conclusion

For the first time, this study describes distinct ERL metabolite patterns in heart transplant patients and detected potentially new drug–drug interactions. The global metabolic profiling facilitated the discovery of novel metabolites associated with ERL dose requirement that might represent new clinically valuable biomarkers to guide ERL therapy.