Gas-phase fragmentation and conformation of the sugar-modified antisense oligonucleotide tcDNA

Tricyclo-DNA (tcDNA) is a sugar- and backbone-modified analogue of DNA that is currently tested as antisense oligonucleotide for the treatment of Duchenne muscular dystrophy. The name tricyclo-DNA is derived from the modified sugar-moiety: the deoxyribose is extended to a three-membered ring system. This modification is designed to limit the flexibility of the structure, thus giving rise to entropically stabilized hybrid duplexes formed between tcDNA and complementary DNA or RNA oligonucleotides. While the structural modifications increase the biostability of the therapeutic agent, they also render the oligonucleotide inaccessible to enzyme-based sequencing methods. Tandem mass spectrometry constitutes an alternative sequencing technique for partially and fully modified oligonucleotides. For reliable sequencing, the fragmentation mechanism of the structure in question must be understood. Therefore, the presented work evaluates the effect of the modified sugar-moiety on the gas-phase dissociation of single stranded tcDNA. Moreover, our experiments reflect the exceptional gas-phase stability of hybrid duplexes that is most noticeable in the formation of truncated duplex ions upon collision-induced dissociation. The stability of the duplex arises from the modified sugar-moiety, as the rigid structure of the tcDNA single strand minimizes the change of the entropy for the annealing. Moreover, the tc-modification gives rise to extended conformations of the nucleic acids in the gas-phase, which was studied by ion mobility spectrometry-mass spectrometry.

Impact of Acquired Pellicle Modification on Adhesion of Early Colonizers

New preventive approaches against dental erosion caused by acidic drinks and beverages include fortification of beverages with natural polymers. We have shown that the mixture of casein and mucin significantly improved the erosion-inhibiting properties of the human pellicle layer. This study aimed to investigate the effect of pellicle modification by casein, mucin and a casein-mucin mixture on the adhesion of early bacterial colonizers. Test specimens of human tooth enamel were prepared, covered with saliva and coated with 0.5% aqueous (aq.) casein, 0.27% aq. mucin or with 0.5% aq. casein-0.27% aq. mucin, after which the adhesion of Streptococcus gordonii, Streptococcus oralis, and Actinomyces odontolyticus was measured after incubation for 30 min and 2 h. log10 colony-forming units were compared by nonparametric tests. All three bacterial strains adhered in higher number to pellicle-coated enamel than to native enamel. The protein modifications of pellicle all decreased the counts of adhering bacteria up to 0.34 log10/mm2, the most efficient being the casein-mucin mixture. In addition to the recently shown erosion-reducing effect by casein-mucin, modification of the pellicle may inhibit bacterial adherence compared to untreated human pellicle.

Regulation of the cardiac Na+ channel NaV1.5 by post-translational modifications.

The cardiac voltage-gated Na(+) channel, Na(V)1.5, is responsible for the upstroke of the action potential in cardiomyocytes and for efficient propagation of the electrical impulse in the myocardium. Even subtle alterations of Na(V)1.5 function, as caused by mutations in its gene SCN5A, may lead to many different arrhythmic phenotypes in carrier patients. In addition, acquired malfunctions of Na(V)1.5 that are secondary to cardiac disorders such as heart failure and cardiomyopathies, may also play significant roles in arrhythmogenesis. While it is clear that the regulation of Na(V)1.5 protein expression and function tightly depends on genetic mechanisms, recent studies have demonstrated that Na(V)1.5 is the target of various post-translational modifications that are pivotal not only in physiological conditions, but also in disease. In this review, we examine the recent literature demonstrating glycosylation, phosphorylation by Protein Kinases A and C, Ca(2+)/Calmodulin-dependent protein Kinase II, Phosphatidylinositol 3-Kinase, Serum- and Glucocorticoid-inducible Kinases, Fyn and Adenosine Monophosphate-activated Protein Kinase, methylation, acetylation, redox modifications, and ubiquitylation of Na(V)1.5. Modern and sensitive mass spectrometry approaches, applied directly to channel proteins that were purified from native cardiac tissues, have enabled the determination of the precise location of post-translational modification sites, thus providing essential information for understanding the mechanistic details of these regulations. The current challenge is first, to understand the roles of these modifications on the expression and the function of Na(V)1.5, and second, to further identify other chemical modifications. It is postulated that the diversity of phenotypes observed with Na(V)1.5-dependent disorders may partially arise from the complex post-translational modifications of channel protein components.

Modification of aviary design reduces incidence of falls, collisions and keel bone damage in laying hens

Non-cage housing systems for laying hens such as aviaries provide greater freedom to perform species-specific behavior and thus are thought to improve welfare of the birds; however, aviaries are associated with a high prevalence of keel bone damage (fractures and deviations), which is a major welfare problem in commercial laying hens. Potential causes of keel bone damage are falls and collisions with internal housing structures that occur as birds move between tiers or perches in the aviary. The aim of this study was to investigate the scope for reducing keel bone damage by reducing falls and collisions through modifications of aviary design. Birds were kept in 20 pens in a laying hen house (225 hens per pen) that were assigned to four different treatments (n = 5 pens per treatment group) including (1) control pens and pens modified by the addition of (2) perches, (3) platforms and (4) ramps. Video recordings at 19, 22, 29, 36 and 43 weeks of age were used to analyze controlled movements and falls (including details on occurrence of collision, cause of fall, height of fall and behavior after fall) during the transitional dusk and subsequent dark phase. Palpation assessments (focusing on fractures and deviations) using 20 focal hens per pen were conducted at 18, 20, 23, 30, 37, 44, 52 and 60 weeks of age. In comparison to the control group, we found 44% more controlled movements in the ramp (P = 0.003) and 47% more controlled movements in the platform treatments (P = 0.014) as well as 45% fewer falls (P = 0.006) and 59% fewer collisions (P < 0.001) in the ramp treatment. There were no significant differences between the control and perch treatments. Also, at 60 weeks of age, 23% fewer fractured keel bones were found in the ramp compared with the control treatment (P = 0.0053). After slaughter at 66 weeks of age, no difference in keel bone damage was found between treatment groups and the prevalence of fractures increased to an average of 86%. As a potential mechanism to explain the differences in locomotion, we suggest that ramps facilitated movement in the vertical plane by providing a continuous path between the tiers and thus supported more natural behavior (i.e. walking and running) of the birds. As a consequence of reducing events that potentially damage keel bones, the installation of ramps may have reduced the prevalence of keel fractures for a major portion of the flock cycle. We conclude that aviary design and installation of specific internal housing structures (i.e. ramps and platforms) have considerable potential to reduce keel bone damage of laying hens in aviary systems.

3D Direct Simulation Monte Carlo Modelling of the Inner Gas Coma of Comet 67P/Churyumov-Gerasimenko: A Parameter Study

Direct Simulation Monte Carlo (DSMC) is a powerful numerical method to study rarefied gas flows such as cometary comae and has been used by several authors over the past decade to study cometary outflow. However, the investigation of the parameter space in simulations can be time consuming since 3D DSMC is computationally highly intensive. For the target of ESA's Rosetta mission, comet 67P/Churyumov-Gerasimenko, we have identified to what extent modification of several parameters influence the 3D flow and gas temperature fields and have attempted to establish the reliability of inferences about the initial conditions from in situ and remote sensing measurements. A large number of DSMC runs have been completed with varying input parameters. In this work, we present the simulation results and conclude on the sensitivity of solutions to certain inputs. It is found that among cases of water outgassing, the surface production rate distribution is the most influential variable to the flow field.