Wound-healing defect of CD18(-/-) mice due to a decrease in TGF-beta1 and myofibroblast differentiation

We studied the mechanisms underlying the severely impaired wound healing associated with human leukocyte-adhesion deficiency syndrome-1 (LAD1) using a murine disease model. In CD18(-/-) mice, healing of full-thickness wounds was severely delayed during granulation-tissue contraction, a phase where myofibroblasts play a major role. Interestingly, expression levels of myofibroblast markers alpha-smooth muscle actin and ED-A fibronectin were substantially reduced in wounds of CD18(-/-) mice, suggesting an impaired myofibroblast differentiation. TGF-beta signalling was clearly involved since TGF-beta1 and TGF-beta receptor type-II protein levels were decreased, while TGF-beta(1) injections into wound margins fully re-established wound closure. Since, in CD18(-/-) mice, defective migration leads to a severe reduction of neutrophils in wounds, infiltrating macrophages might not phagocytose apoptotic CD18(-/-) neutrophils. Macrophages would thus be lacking their main stimulus to secrete TGF-beta1. Indeed, in neutrophil-macrophage cocultures, lack of CD18 on either cell type leads to dramatically reduced TGF-beta1 release by macrophages due to defective adhesion to, and subsequent impaired phagocytic clearance of, neutrophils. Our data demonstrates that the paracrine secretion of growth factors is essential for cellular differentiation in wound healing.     

Genome sequence of Phaeobacter caeruleus type strain (DSM 24564T), a surface-associated member of the marine Roseobacter clade

In 2009 Phaeobacter caeruleus was described as a novel species affiliated with the marine Roseobacter clade, which, in turn, belongs to the class Alphaproteobacteria. The genus Phaeobacter is well known for members that produce various secondary metabolites. Here we report of putative quorum sensing systems, based on the finding of six N-acyl-homoserine lactone synthetases, and show that the blue color of P. caeruleus is probably due to the production of the secondary metabolite indigoidine. Therefore, P. caeruleus might have inhibitory effects on other bacteria. In this study the genome of the type strain DSM 24564^T was sequenced, annotated and characterized. The 5,344,419 bp long genome with its seven plasmids contains 5,227 protein-coding genes (3,904 with a predicted function) and 108 RNA genes.     

Physiological diversity of Roseobacter clade bacteria co-occurring during a phytoplankton bloom in the North Sea

Organisms of the Roseobacter clade are an important component in marine ecosystems, partially due to their metabolic variety. Not much is known, however, about the physiological diversity of different roseobacters present within one habitat. By using serial dilution cultures with low-nutrient media seven roseobacter strains, co-occurring during a phytoplankton bloom in the southern North Sea, were obtained in this study. Physiological characterization exhibited distinct substrate spectra of the isolates. Although no isolate showed growth on algal osmolyte dimethylsulfoniopropionate (DMSP), feeding experiments revealed that all new strains converted [²H₆]DMSP into a variety of volatile compounds. Six strains mainly decomposed DMSP via the demethylation pathway, but four strains were also capable of cleaving DMSP to DMS and acrylate. It is hypothesized that the great physiological diversity of the roseobacters reflects their ability to inhabit different ecological niches and enables the organisms to cope differently with changing substrate supplies during phytoplankton blooms. Denaturing gradient gel electrophoresis and sequencing of excised bands resulted in detection of five additional roseobacters. Three of these sequences showed affiliation with three of the four major clusters of the Roseobacter clade, consisting predominantly of uncultured organisms (i.e. the Roseobacter clade-affiliated (RCA)), the NAC11-7 and the CHAB-I-5 clusters.     

Selection,drift and community interactions shape microbial biogeographic patterns in the Pacific Ocean

Despite accumulating data on microbial biogeographic patterns in terrestrial and aquatic environments, we still lack a comprehensive understanding of how these patterns establish, in particular in ocean basins. Here we show the relative significance of the ecological mechanisms selection, dispersal and drift for shaping the composition of microbial communities in the Pacific Ocean over a transect of 12,400 km between subantarctic and subarctic regions. In the epipelagic, homogeneous selection contributes 50–60% and drift least to the three mechanism for the assembly of prokaryotic communities whereas in the upper mesopelagic, drift is relatively most important for the particle-associated subcommunities. Temperature is important for the relative significance of homogeneous selection and dispersal limitation for community assembly. The relative significance of both mechanisms was inverted with increasing temperature difference along the transect. For eukaryotes >8 µm, homogeneous selection is also the most important mechanisms at two epipelagic depths whereas at all other depths drift is predominant. As species interactions are essential for structuring microbial communities we further analyzed co-occurrence-based community metrics to assess biogeographic patterns over the transect. These interaction-adjusted indices explained much better variations in microbial community composition as a function of abiotic and biotic variables than compositional or phylogenetic distance measures like Bray–Curtis or UniFrac. Our analyses are important to better understand assembly processes of microbial communities in the upper layers of the largest ocean and how they adapt to effectively perform in global biogeochemical processes. Similar principles presumably act upon microbial community assembly in other ocean basins.Subject terms: Microbial ecology, Microbiology     

Configural Frequency Analysis (CFA) Revisited — a New Look at an Old Approach

Configural frequency analysis (CFA) is a widely used method for the identification of types and syndromes in contingency tables. However, the type model of CFA shows some major deficiencies. In this paper, we propose an alternative modeling of types eliminating the shortcomings of CFA. Basically, a type is modeled as a combination of traits or symptoms that deviates from the pattern of association holding true for the complementary configurations of the contingency table. The new approach is formulated in terms of a log-linear model. It is shown that parameter estimation can be performed with methods known from the analysis of incomplete contingency tables. Test procedures for confirmatory analysis and methods for exploratory search for type configurations are developed. We illustrate the methodology with two practical examples.     

Hydroxynitrile lyase from Hevea brasiliensis: Molecular characterization and mechanism of enzyme catalysis

(S)-Hydroxynitrile lyase (Hnl) from the tropical rubber tree Hevea brasiliensis is a 29 kDa single chain protein that catalyses the breakdown or formation of a C(SINGLE BOND)C bond by reversible addition of hydrocyanic acid to aldehydes or ketones. The primary sequence of Hnl has no significant homology to known proteins. Detailed homology investigations employing PROFILESEARCH and secondary structure prediction algorithms suggest that Hnl is a member of the α/β hydrolase fold protein family and contains a catalytic triad as functional residues for catalysis. The significance of the predicted catalytic residues was tested and confirmed by site-directed mutagenesis and expression of mutant and wild-type proteins in the yeast, Saccharomyces cerevisiae. Based on these data we suggest a mechanistic model for the (S)-cyanohydrin synthesis catalyzed by hydroxynitrile lyase from Hevea brasiliensis. Proteins 27:438–449, 1997. © 1997 Wiley-Liss, Inc.     

Distribution of Roseobacter RCA and SAR11 lineages and distinct bacterial communities from the subtropics to the Southern Ocean

We assessed the composition of the bacterioplankton in the Atlantic sector of the Southern Ocean in austral fall and winter and in New Zealand coastal waters in summer. The various water masses between the subtropics/Agulhas–Benguela boundary region and the Antarctic coastal current exhibited distinct bacterioplankton communities with the highest richness in the polar frontal region, as shown by denaturing gradient gel electrophoresis of 16S rRNA gene fragments. The SAR11 clade and the Roseobacter clade‐affiliated (RCA) cluster were quantified by real‐time quantitative PCR. SAR11 was detected in all samples analysed from subtropical waters to the coastal current and to depths of > 1000 m. In fall and winter, this clade constituted < 3% to 48% and 4–28% of total bacterial 16S rRNA genes respectively, with highest fractions in subtropical to polar frontal regions. The RCA cluster was only present in New Zealand coastal surface waters not exceeding 17°C, in the Agulhas–Benguela boundary region (visited only during the winter cruise), in subantarctic waters and in the Southern Ocean. In fall, this cluster constituted up to 36% of total bacterial 16S rRNA genes with highest fractions in the Antarctic coastal current and outnumbered the SAR11 clade at most stations in the polar frontal region and further south. In winter, the RCA cluster constituted lower proportions than the SAR11 clade and did not exceed 8% of total bacterial 16S rRNA genes. In fall, the RCA cluster exhibited significant positive correlations with latitude and ammonium concentrations and negative correlations with concentrations of nitrate, phosphate, and for near‐surface samples also with chlorophyll a, biomass production of heterotrophic prokaryotes and glucose turnover rates. The findings show that the various water masses between the subtropics and the Antarctic coastal current harbour distinct bacterioplankton communities. They further indicate that the RCA cluster, despite the narrow sequence similarity of > 98% of its 16S rRNA gene, is an abundant component of the heterotrophic bacterioplankton in the Southern Ocean, in particular in its coldest regions.     

Small molecular antioxidants effectively protect from PUVA-induced oxidative stress responses underlying fibroblast senescence and photoaging

Exposure of human fibroblasts to 8-methoxypsoralen plus ultraviolet-A irradiation (PUVA) results in stress-induced cellular senescence in fibroblasts. We here studied the role of the antioxidant defense system in the accumulation of reactive oxygen species (ROS) and the effect of the antioxidants alpha-tocopherol, N-acetylcysteine, and alpha-lipoic acid on PUVA-induced cellular senescence. PUVA treatment induced an immediate and increasing generation of intracellular ROS. Supplementation of PUVA-treated fibroblasts with alpha-tocopherol (alpha-Toc), N-acetylcysteine (NAC), or alpha-lipoic acid (alpha-LA) abrogated the increased ROS generation and rescued fibroblasts from the ROS-dependent changes into the cellular senescence phenotype, such as cytoplasmic enlargement, enhanced expression of senescence-associated-beta-galactosidase and matrix-metalloproteinase-1, hallmarks of photoaging and intrinsic aging. PUVA treatment disrupted the integrity of cellular membranes and impaired homeostasis and function of the cellular antioxidant system with a significant decrease in glutathione and hydrogen peroxide-detoxifying enzymes activities. Supplementation with NAC, alpha-LA, and alpha-Toc counteracted these changes. Our data provide causal evidence that (i) oxidative stress due to an imbalance in the overall cellular antioxidant capacity contributes to the induction and maintenance of the PUVA-induced fibroblast senescence and that (ii) low molecular antioxidants protect effectively against these deleterious alterations.