A GFP-MAP4 reporter gene for visualizing cortical microtubule rearrangements in living epidermal cells

Microtubules influence morphogenesis by forming distinct geometrical arrays in the cell cortex, which in turn affect the deposition of cellulose microfibrils. Although many chemical and physical factors affect microtubule orientation, it is unclear how cortical microtubules in elongating cells maintain their ordered transverse arrays and how they reorganize into new geometries. To visualize these reorientations in living cells, we constructed a microtubule reporter gene by fusing the microtubule binding domain of the mammalian microtubule-associated protein 4 (MAP4) gene with the green fluorescent protein (GFP) gene, and transient expression of the recombinant protein in epidermal cells of fava bean was induced. The reporter protein decorates microtubules in vivo and binds to microtubules in vitro. Confocal microscopy and time-course analysis of labeled cortical arrays along the outer epidermal wall revealed the lengthening, shortening, and movement of microtubules; localized microtubule reorientations; and global microtubule reorganizations. The global microtubule orientation in some cells fluctuates about the transverse axis and may be a result of a cyclic self-correcting mechanism to maintain a net transverse orientation during cellular elongation.     

Influence of nanofibers on growth and gene expression of human tendon derived fibroblast

Background  

Rotator cuff tears are a common and frequent lesion especially in older patients. The mechanisms of tendon repair are not fully understood. Common therapy options for tendon repair include mini-open or arthroscopic surgery. The use of growth factors in experimental studies is mentioned in the literature. Nanofiber scaffolds, which provide several criteria for the healing process, might be a suitable therapy option for operative treatment. The aim of this study was to explore the effects of nanofiber scaffolds on human tendon derived fibroblasts (TDF's), as well as the gene expression and matrix deposition of these fibroblasts.     

Migration, isolation, and speciation of hydrothermal vent limpets (Gastropoda; Lepetodrilidae) across the Blanco Transform Fault

The Sovanco Fracture Zone and Blanco Transform Fault separate the Explorer, Juan de Fuca, and Gorda ridge systems of the northeastern Pacific Ocean. To test whether such offsets in the ridge axis create barriers to along-axis dispersal of the endemic hydrothermal vent animals, we examined the genetic structure of limpet populations previously identified as Lepetodrilus fucensis McLean, 1988 (Gastropoda, Lepetodrilidae). Mitochondrial DNA sequences and patterns of allozyme variation revealed no evidence that the 150-km-long Sovanco Fracture Zone impeded gene flow between the Explorer and Juan de Fuca populations. In contrast, the 450-km-long Blanco Transform Fault separates the limpets into highly divergent northern and southern lineages that we recognize as distinct species. We describe southern populations from the Gorda Ridge (Seacliff) and Escanaba Trough as Lepetodrilus gordensis new species and refer northern populations from the Explorer and Juan de Fuca ridge systems to L. fucensis sensu stricto. The species are similar morphologically, but L. gordensis lacks a sensory neck papilla and has a more tightly coiled teleconch. To assess the degree of isolation between these closely related species, we used the Isolation with Migration method to estimate the time of population splitting, effective sizes of the ancestral and derived populations, and rates of migration across the Blanco Transform Fault.     

The genome of Rhizobium leguminosarum has recognizable core and accessory components

Background

Rhizobium leguminosarum is an α-proteobacterial N₂-fixing symbiont of legumes that has been the subject of more than a thousand publications. Genes for the symbiotic interaction with plants are well studied, but the adaptations that allow survival and growth in the soil environment are poorly understood. We have sequenced the genome of R. leguminosarum biovar viciae strain 3841.

Results

The 7.75 Mb genome comprises a circular chromosome and six circular plasmids, with 61% G+C overall. All three rRNA operons and 52 tRNA genes are on the chromosome; essential protein-encoding genes are largely chromosomal, but most functional classes occur on plasmids as well. Of the 7,263 protein-encoding genes, 2,056 had orthologs in each of three related genomes (Agrobacterium tumefaciens, Sinorhizobium meliloti, and Mesorhizobium loti), and these genes were over-represented in the chromosome and had above average G+C. Most supported the rRNA-based phylogeny, confirming A. tumefaciens to be the closest among these relatives, but 347 genes were incompatible with this phylogeny; these were scattered throughout the genome but were over-represented on the plasmids. An unexpectedly large number of genes were shared by all three rhizobia but were missing from A. tumefaciens.

Conclusion

Overall, the genome can be considered to have two main components: a 'core', which is higher in G+C, is mostly chromosomal, is shared with related organisms, and has a consistent phylogeny; and an 'accessory' component, which is sporadic in distribution, lower in G+C, and located on the plasmids and chromosomal islands. The accessory genome has a different nucleotide composition from the core despite a long history of coexistence.     

Periodontal therapy increases neutrophil extracellular trap degradation

In periodontitis, polymorphonuclear leucocytes (PMNs) are activated. They entrap and eliminate pathogens by releasing neutrophil extracellular traps (NETs). Abnormal NET degradation is part of a pro-inflammatory status, affecting co-morbidities such as cardiovascular disease. We aimed to investigate the ex vivo NET degradation capacity of plasma from periodontitis patients compared to controls (part 1) and to quantify NET degradation before and after periodontal therapy (part 2). Fresh NETs were obtained by stimulating blood-derived PMNs with phorbol 12-myristate 13-acetate. Plasma samples from untreated periodontitis patients and controls were incubated for 3 h onto freshly generated NETs (part 1). Similarly, for part 2, NET degradation was studied for 91 patients before and 3, 6 and 12 mo after non-surgical periodontal therapy with and without adjunctive systemic antibiotics. Finally, NET degradation was fluorospectrometrically quantified. NET degradation levels did not differ between periodontitis patients and controls, irrespective of subject-related background characteristics. NET degradation significantly increased from 65.6 ± 1.7% before periodontal treatment to 75.7 ± 1.2% at 3 mo post periodontal therapy, and this improvement was maintained at 6 and 12 mo, irrespective of systemic usage of antibiotics. Improved NET degradation after periodontitis treatment is another systemic biomarker reflecting a decreased pro-inflammatory status, which also contributes to an improved cardiovascular condition.     

Increased Platelet Reactivity Is Associated with Circulating Platelet-Monocyte Complexes and Macrophages in Human Atherosclerotic Plaques

Objective

Platelet reactivity, platelet binding to monocytes and monocyte infiltration play a detrimental role in atherosclerotic plaque progression. We investigated whether platelet reactivity was associated with levels of circulating platelet-monocyte complexes (PMCs) and macrophages in human atherosclerotic carotid plaques.

Methods

Platelet reactivity was determined by measuring platelet P-selectin expression after platelet stimulation with increasing concentrations of adenosine diphosphate (ADP), in two independent cohorts: the Circulating Cells cohort (n = 244) and the Athero-Express cohort (n = 91). Levels of PMCs were assessed by flow cytometry in blood samples of patients who were scheduled for percutaneous coronary intervention (Circulating Cells cohort). Monocyte infiltration was semi-quantitatively determined by histological examination of atherosclerotic carotid plaques collected during carotid endarterectomy (Athero-Express cohort).

Results

We found increased platelet reactivity in patients with high PMCs as compared to patients with low PMCs (median (interquartile range): 4153 (1585–11267) area under the curve (AUC) vs. 9633 (3580–21565) AUC, P<0.001). Also, we observed increased platelet reactivity in patients with high macrophage levels in atherosclerotic plaques as compared to patients with low macrophage levels in atherosclerotic plaques (mean±SD; 8969±3485 AUC vs. 7020±3442 AUC, P = 0.02). All associations remained significant after adjustment for age, sex and use of drugs against platelet activation.

Conclusion

Platelet reactivity towards ADP is associated with levels of PMCs and macrophages in human atherosclerotic carotid plaques.     

Multidimensional prognostic indices for use in COPD patient care. A systematic review

Background

A growing number of prognostic indices for chronic obstructive pulmonary disease (COPD) is developed for clinical use. Our aim is to identify, summarize and compare all published prognostic COPD indices, and to discuss their performance, usefulness and implementation in daily practice.

Methods

We performed a systematic literature search in both Pubmed and Embase up to September 2010. Selection criteria included primary publications of indices developed for stable COPD patients, that predict future outcome by a multidimensional scoring system, developed for and validated with COPD patients only. Two reviewers independently assessed the index quality using a structured screening form for systematically scoring prognostic studies.

Results

Of 7,028 articles screened, 13 studies comprising 15 indices were included. Only 1 index had been explored for its application in daily practice. We observed 21 different predictors and 7 prognostic outcomes, the latter reflecting mortality, hospitalization and exacerbation. Consistent strong predictors were FEV₁ percentage predicted, age and dyspnoea. The quality of the studies underlying the indices varied between fairly poor and good. Statistical methods to assess the predictive abilities of the indices were heterogenic. They generally revealed moderate to good discrimination, when measured. Limitations: We focused on prognostic indices for stable disease only and, inevitably, quality judgment was prone to subjectivity.

Conclusions

We identified 15 prognostic COPD indices. Although the prognostic performance of some of the indices has been validated, they all lack sufficient evidence for implementation. Whether or not the use of prognostic indices improves COPD disease management or patients'' health is currently unknown; impact studies are required to establish this.     

Acquisition of dwarf male "harems" by recently settled females of Osedax roseus n. sp. (Siboglinidae; Annelida)

After the deployment of several whale carcasses in Monterey Bay, California, a time-series analysis revealed the presence of a new species of Osedax, a genus of bone-eating siboglinid annelids. That species is described here as Osedax roseus n. sp. It is the fifth species described since the erection of this genus and, like its congeners, uses a ramifying network of "roots" to house symbiotic bacteria. In less than 2 months, Osedax roseus n. sp. colonized the exposed bones of a whale carcass deposited at 1018-m depth, and many of the females were fecund in about 3 months post-deployment. As with other Osedax spp., the females have dwarf males in their tube lumens. The males accrue over time until the sex ratio is markedly male-biased. This pattern of initial female settlement followed by gradual male accumulation is consistent with the hypothesis that male sex may be environmentally determined in Osedax. Of the previously described species in this genus, Osedax roseus n. sp. is most similar to O. rubiplumus, but it has several anatomical differences, as well as much smaller females, dwarf males, and eggs. Osedax roseus n. sp. is markedly divergent (minimally 16.6%) for mitochondrial cytochrome oxidase subunit I (mtCOI) sequences from any other Osedax species.     

Tyrosine phosphorylation of myosin heavy chain during skeletal muscle differentiation: an integrated bioinformatics approach

Background  

Previously it has been shown that insulin-mediated tyrosine phosphorylation of myosin heavy chain is concomitant with enhanced association of C-terminal SRC kinase during skeletal muscle differentiation. We sought to identify putative site(s) for this phosphorylation event.     

Epibiotic bacteria associated with the recently discovered Yeti crab, Kiwa hirsuta

The Yeti crab, Kiwa hirsuta Macpherson et al., is the single known species in a recently discovered crab family Kiwaidae (Decapoda: Galatheoidea) from deep-sea hydrothermal vents. Its chelipeds, walking legs and the ventral surface of its cephalothorax are covered with dense setae that, in turn, are covered with clusters of filamentous bacteria, making the crab appear extraordinarily 'hairy'. Electron microscopy revealed dense bacterial clusters attached to the chitinous outer layer of the setae. Molecular phylogenetic analyses revealed the setae-associated bacteria to be dominated by epsilon-Proteobacteria ( approximately 56% of the recovered ribotypes), gamma-Proteobacteria ( approximately 25%) and Bacteroidetes ( approximately 10%). Fluorescence in situ microscopy confirmed the attachment of filamentous epsilon-Proteobacteria on setae, but no specialized morphological structures appeared to exist for bacterial attachment. Key enzymes involved in the reductive tricarboxylic acid cycle (ATP-dependent citrate lyase) and sulfite oxidation or dissimilatory sulfate reduction (bidirectional APS reductase) were detected. Consequently, the potential for carbon fixation and cycling of reduced and oxidized sulfur appear to exist in the dense microflora that grows on the crab's setae.