X­irradiation induces acute and early term inflammatory responses in atherosclerosis­prone ApoE­/­ mice and in endothelial cells.

Thoracic radiotherapy is an effective treatment for many types of cancer; however it is also associated with an increased risk of developing cardiovascular disease (CVD), appearing mainly ≥10 years after radiation exposure. The present study investigated acute and early term physiological and molecular changes in the cardiovascular system after ionizing radiation exposure. Female and male ApoE­/­ mice received a single exposure of low or high dose X­ray thoracic irradiation (0.1 and 10 Gy). The level of cholesterol and triglycerides, as well as a large panel of inflammatory markers, were analyzed in serum samples obtained at 24 h and 1 month after irradiation. The secretion of inflammatory markers was further verified in vitro in coronary artery and microvascular endothelial cell lines after exposure to low and high dose of ionizing radiation (0.1 and 5 Gy). Local thoracic irradiation of ApoE­/­ mice increased serum growth differentiation factor­15 (GDF­15) and C­X­C motif chemokine ligand 10 (CXCL10) levels in both female and male mice 24 h after high dose irradiation, which were also secreted from coronary artery and microvascular endothelial cells in vitro. Sex­specific responses were observed for triglyceride and cholesterol levels, and some of the assessed inflammatory markers as detailed below. Male ApoE­/­ mice demonstrated elevated intercellular adhesion molecule­1 and P­selectin at 24 h, and adiponectin and plasminogen activator inhibitor­1 at 1 month after irradiation, while female ApoE­/­ mice exhibited decreased monocyte chemoattractant protein­1 and urokinase­type plasminogen activator receptor at 24 h, and basic fibroblast growth factor 1 month after irradiation. The inflammatory responses were mainly significant following high dose irradiation, but certain markers showed significant changes after low dose exposure. The present study revealed that acute/early inflammatory responses occurred after low and high dose thoracic irradiation. However, further research is required to elucidate early asymptomatic changes in the cardiovascular system post thoracic X­irradiation and to investigate whether GDF­15 and CXCL10 could be considered as potential biomarkers for the early detection of CVD risk in thoracic radiotherapy­treated patients.

Complex phylogenetic distribution of a non-canonical genetic code in green algae.

BACKGROUND: A non-canonical nuclear genetic code, in which TAG and TAA have been reassigned from stop codons to glutamine, has evolved independently in several eukaryotic lineages, including the ulvophycean green algal orders Dasycladales and Cladophorales. To study the phylogenetic distribution of the standard and non-canonical genetic codes, we generated sequence data of a representative set of ulvophycean green algae and used a robust green algal phylogeny to evaluate different evolutionary scenarios that may account for the origin of the non-canonical code. RESULTS: This study demonstrates that the Dasycladales and Cladophorales share this alternative genetic code with the related order Trentepohliales and the genus Blastophysa, but not with the Bryopsidales, which is sister to the Dasycladales. This complex phylogenetic distribution whereby all but one representative of a single natural lineage possesses an identical deviant genetic code is unique. CONCLUSIONS: We compare different evolutionary scenarios for the complex phylogenetic distribution of this non-canonical genetic code. A single transition to the non-canonical code followed by a reversal to the canonical code in the Bryopsidales is highly improbable due to the profound genetic changes that coincide with codon reassignment. Multiple independent gains of the non-canonical code, as hypothesized for ciliates, are also unlikely because the same deviant code has evolved in all lineages. Instead we favor a stepwise acquisition model, congruent with the ambiguous intermediate model, whereby the non-canonical code observed in these green algal orders has a single origin. We suggest that the final steps from an ambiguous intermediate situation to a non-canonical code have been completed in the Trentepohliales, Dasycladales, Cladophorales and Blastophysa but not in the Bryopsidales. We hypothesize that in the latter lineage an initial stage characterized by translational ambiguity was not followed by final reassignment of both stop codons to glutamine. Instead the standard code was retained by the disappearance of the ambiguously decoding tRNAs from the genome. We correlate the emergence of a non-canonical genetic code in the Ulvophyceae to their multinucleate nature.

Evolution and cytological diversification of the green seaweeds (Ulvophyceae).

The Ulvophyceae, one of the four classes of the Chlorophyta, is of particular evolutionary interest because it features an unrivaled morphological and cytological diversity. Morphological types range from unicells and simple multicellular filaments to sheet-like and complex corticated thalli. Cytological layouts range from typical small cells containing a single nucleus and chloroplast to giant cells containing millions of nuclei and chloroplasts. In order to understand the evolution of these morphological and cytological types, the present paper aims to assess whether the Ulvophyceae are monophyletic and elucidate the ancient relationships among its orders. Our approach consists of phylogenetic analyses (maximum likelihood and Bayesian inference) of seven nuclear genes, small subunit nuclear ribosomal DNA and two plastid markers with carefully chosen partitioning strategies, and models of sequence evolution. We introduce a procedure for fast site removal (site stripping) targeted at improving phylogenetic signal in a particular epoch of interest and evaluate the specificity of fast site removal to retain signal about ancient relationships. From our phylogenetic analyses, we conclude that the ancestral ulvophyte likely was a unicellular uninucleate organism and that macroscopic growth was achieved independently in various lineages involving radically different mechanisms: either by evolving multicellularity with coupled mitosis and cytokinesis (Ulvales-Ulotrichales and Trentepohliales), by obtaining a multinucleate siphonocladous organization where every nucleus provides for its own cytoplasmic domain (Cladophorales and Blastophysa), or by developing a siphonous organization characterized by either one macronucleus or millions of small nuclei and cytoplasmic streaming (Bryopsidales and Dasycladales). We compare different evolutionary scenarios giving rise to siphonous and siphonocladous cytologies and argue that these did not necessarily evolve from a multicellular or even multinucleate state but instead could have evolved independently from a unicellular ancestor.

Quantification of hydrophilic ethoxylates in polysorbate surfactants using diffusion H1 NMR spectroscopy.

Polysorbate surfactants (commercially available as Tween) are widely used in pharmaceutical, cosmetic and food products. They are generally considered as esters of ethoxylated sorbitan with fatty acids. Diffusion H1 NMR spectroscopy on a solution of polysorbate 20 in D2O revealed that only one diffusion coefficient was found for the fatty acyl part. Using the Stokes-Einstein equation, it became obvious that this diffusion behavior was caused by micelles. On the other hand, two significantly different diffusion coefficients were found for the methylene groups of ethylene oxide (EO). This indicates the presence of two distinct EO containing species in solution. Since the slowest diffusing EO species has the same diffusion coefficient as the fatty acyl part, it corresponds to the micellar (i.e. fatty acyl bound) ethoxylates. The diffusion coefficient of the fastest diffusing EO species was a factor of four larger than that of the slowly diffusing species and was attributed to water-soluble non-esterified ethoxylates. A solution of polysorbate 20 in the presence of NaOD was prepared to investigate if hydrolysis of the sorbitan ester could be the reason for the occurence of these hydrophilic ethoxylates. It was found that alkaline hydrolysis does lead to an increasing fraction of non-esterified ethoxylates, but is not the cause of its presence in untreated polysorbate samples since these species were also found in solutions of polyethylene glycol oleyl ether (commercially available as Brij), which are not susceptible to hydrolysis. Fractionation of the EO species present in polysorbate 20 into an amphiphilic and a hydrophilic fraction was only partly obtained by activated carbon adsorption. On the other hand, sequential extraction of aqueous polysorbate solutions by ethyl acetate and chloroform enabled a nearly complete fractionation. H1 NMR spectroscopy proved to be very useful since it allows in situ determination of the global composition of a surfactant sample, as well as quantification of both the amphiphilic and hydrophilic ethoxylate fractions via diffusion measurements.

Solubilization of flurbiprofen within non-ionic Tween 20 surfactant micelles: a 19F and 1H NMR study.

The solubilization of the poorly water soluble anti-inflammatory drug flurbiprofen in non-ionic Tween 20 surfactant micellar solutions was studied by both (19)F and (1)H NMR spectroscopy in an acidic environment. These non-destructive techniques allowed us to investigate the effect of temperature cycling in situ. Using (19)F NMR, an increased solubilisation capacity was observed as the temperature increased. This effect became more pronounced above the cloud point, which was reduced by more than 30 degrees C in the presence of an excess of flurbiprofen. Upon clouding, peak splitting was observed in the (19)F spectrum, which indicates that two pools of solubilised flurbiprofen exist that are in slow exchange on the NMR frequency timescale. The clouding and solubilization processes were found to be reversible, albeit with slow kinetics. Based on chemical shift differences of both Tween 20 and flurbiprofen, as well as NOESY experiments, the flurbiprofen was found to be accumulated within the palisade layer of the Tween 20 micelles.

Gain and loss of elongation factor genes in green algae.

BACKGROUND: Two key genes of the translational apparatus, elongation factor-1 alpha (EF-1alpha) and elongation factor-like (EFL) have an almost mutually exclusive distribution in eukaryotes. In the green plant lineage, the Chlorophyta encode EFL except Acetabularia where EF-1alpha is found, and the Streptophyta possess EF-1alpha except Mesostigma, which has EFL. These results raise questions about evolutionary patterns of gain and loss of EF-1alpha and EFL. A previous study launched the hypothesis that EF-1alpha was the primitive state and that EFL was gained once in the ancestor of the green plants, followed by differential loss of EF-1alpha or EFL in the principal clades of the Viridiplantae. In order to gain more insight in the distribution of EF-1alpha and EFL in green plants and test this hypothesis we screened the presence of the genes in a large sample of green algae and analyzed their gain-loss dynamics in a maximum likelihood framework using continuous-time Markov models. RESULTS: Within the Chlorophyta, EF-1alpha is shown to be present in three ulvophycean orders (i.e., Dasycladales, Bryopsidales, Siphonocladales) and the genus Ignatius. Models describing gene gain-loss dynamics revealed that the presence of EF-1alpha, EFL or both genes along the backbone of the green plant phylogeny is highly uncertain due to sensitivity to branch lengths and lack of prior knowledge about ancestral states or rates of gene gain and loss. Model refinements based on insights gained from the EF-1alpha phylogeny reduce uncertainty but still imply several equally likely possibilities: a primitive EF-1alpha state with multiple independent EFL gains or coexistence of both genes in the ancestor of the Viridiplantae or Chlorophyta followed by differential loss of one or the other gene in the various lineages. CONCLUSION: EF-1alpha is much more common among green algae than previously thought. The mutually exclusive distribution of EF-1alpha and EFL is confirmed in a large sample of green plants. Hypotheses about the gain-loss dynamics of elongation factor genes are hard to test analytically due to a relatively flat likelihood surface, even if prior knowledge is incorporated. Phylogenetic analysis of EFL genes indicates misinterpretations in the recent literature due to uncertainty regarding the root position.

A multi-locus time-calibrated phylogeny of the siphonous green algae.

The siphonous green algae are an assemblage of seaweeds that consist of a single giant cell. They comprise two sister orders, the Bryopsidales and Dasycladales. We infer the phylogenetic relationships among the siphonous green algae based on a five-locus data matrix and analyze temporal aspects of their diversification using relaxed molecular clock methods calibrated with the fossil record. The multi-locus approach resolves much of the previous phylogenetic uncertainty, but the radiation of families belonging to the core Halimedineae remains unresolved. In the Bryopsidales, three main clades were inferred, two of which correspond to previously described suborders (Bryopsidineae and Halimedineae) and a third lineage that contains only the limestone-boring genus Ostreobium. Relaxed molecular clock models indicate a Neoproterozoic origin of the siphonous green algae and a Paleozoic diversification of the orders into their families. The inferred node ages are used to resolve conflicting hypotheses about species ages in the tropical marine alga Halimeda.

NMR study of the sorption behavior of benzyl alcohol derivatives into sonicated and extruded dioctadecyldimethylammonium chloride (DODAC) dispersions: the relevance of membrane fluidity.

The sorption behavior of three benzyl alcohol derivatives with different hydrophobicities into sonicated and extruded DODAC dispersions has been studied using NMR spectroscopy and NMR diffusometry. We show that there is an increased sorption into a sonicated dispersion below the phase-transition temperature (T(m)) as compared to an extruded dispersion. This may be explained by the incomplete lipid chain freezing of charged lipids as a result of the sonication process. Around T(m), a sorption maximum is found that is attributed to the high bilayer disorder under this condition. In addition, a sorption increase and a fluidizing effect at increasing benzyl alcohol derivative concentrations are observed that provide additional evidence for the relevance of the bilayer fluidity on the sorption of hydrophobic components.