Application of beta-irradiation through the struts of a previously deployed stent

The application of beta-radiation in coronary arteries is a promising new technique for the treatment of in-stent restenosis. This is the first case in which the 5 F. delivery catheter of the Beta-Cath trade mark system was advanced through the struts of a stent, previously deployed in an adjacent branch, so as to deliver radiation to the target vessel.     

Analysis of mutants of tetanus toxin Hc fragment: ganglioside binding, cell binding and retrograde axonal transport properties

Tetanus toxin binds neuronal tissue prior to internalization and trafficking to the central nervous system. Binding of the carboxy-terminal 50 kDa HC fragment of tetanus toxin to polysialogangliosides is important for this initial cell binding step. Using the three-dimensional structure of HC, mutants were designed to investigate the role of individual residues in ganglioside binding. Mutant proteins were tested for binding to GT1b gangliosides, to primary motoneurons and for their ability to undergo retrograde transport in mice. Two classes of mutant were obtained: (i) those containing deletions in loop regions within the C-terminal beta-trefoil domain which showed greatly reduced ganglioside and cell binding and did not undergo retrograde transport and (ii) those that showed reduced ganglioside binding, but retained primary neuronal cell binding and retrograde transport. The second class included point mutants of Histidine-1293, previously implicated in GT1b binding. Our deletion analysis is entirely consistent with recent structural studies which have identified sugar-binding sites in the immediate vicinity of the residues identified by mutagenesis. These results demonstrate that ganglioside binding can be severely impaired without abolishing cell binding and intracellular trafficking of tetanus toxin.     

Reaction mechanisms of non-heme diiron hydroxylases characterized in whole cells

Whole cells expressing the non-heme diiron hydroxylases AlkB and toluene 4-monooxygenase (T4MO) were used to probe enzyme reaction mechanisms. AlkB catalyzes the hydroxylation of the radical clock substrates bicyclo[4.1.0]heptane (norcarane), spirooctane and 1,1-diethylcyclopropane, and does not catalyze the hydroxylation of the radical clocks 1,1-dimethylcyclopropane or 1,1,2,2-tetramethylcyclopropane. The hydroxylation of norcarane yields a distribution of products consistent with an "oxygen-rebound" mechanism for the enzyme in both the wild type Pseudomonas putida GPo1 and AlkB from P. putida GPo1 expressed in Escherichia coli. Evidence for the presence of a substrate-based radical during the reaction mechanism is clear. With norcarane, the lifetime of that radical varies with experimental conditions. Experiments with higher substrate concentrations yield a shorter radical lifetime (approximately 1 ns), while experiments with lower substrate concentrations yield a longer radical lifetime (approximately 19 ns). Consistent results were obtained using either wild type or AlkB-equipped host organisms using either "resting cell" or "growing cell" approaches. T4MO expressed in E. coli also catalyzes the hydroxylation of norcarane with a radical lifetime of approximately 0.07 ns. No radical lifetime dependence on substrate concentration was seen. Results from experiments with diethylcyclopropane, spirooctane, dimethylcyclopropane, and diethylcyclopropane are consistent with a restricted active site for AlkB.     

Fgf8 drives myogenic progression of a novel lateral fast muscle fibre population in zebrafish

Fibroblast growth factors (Fgfs) have long been implicated in regulating vertebrate skeletal muscle differentiation, but their precise role(s) in vivo remain unclear. Here, we show that Fgf8 signalling in the somite is required for myod expression and terminal differentiation of a subset of fast muscle cells in the zebrafish lateral somite. In the absence of Fgf8, lateral somite cells transiently express myf5 but fail to make muscle and remain in a dermomyotome-like state characterised by pax3 and meox expression. Slow muscle fibres form and commence normal migration in the absence of Fgf8, but fail to traverse the expanded undifferentiated lateral somite. The Fgf8-independent residual population of medial fast muscle fibres is not Hedgehog dependent. However, Fgf8-independent medial fast muscle precursors are lacking in floatinghead mutants, suggesting that they require another ventral midline-derived signal. We conclude that Fgf8 drives terminal differentiation of a specific population of lateral muscle precursor cells within the early somite.     

Identification to the species level of <Emphasis Type="Italic">Lactobacillus</Emphasis> isolated in probiotic prospecting studies of human,animal or food origin by 16S-23S rRNA restriction profiling

Background  

The accurate identification of Lactobacillus and other co-isolated bacteria during microbial ecological studies of ecosystems such as the human or animal intestinal tracts and food products is a hard task by phenotypic methods requiring additional tests such as protein and/or lipids profiling.     

Functional caspase-1 is required for Langerhans cell migration and optimal contact sensitization in mice

Langerhans cell (LC) migration from epidermis to draining lymph node is a critical first step in cutaneous immune responses. Both TNF-alpha and IL-1 beta are important signals governing this process, but the potential regulatory role of IL-1 alpha processing by caspase-1 is unknown. In wild-type (WT) mice, application of the contact allergens 2,4-dinitrofluorobenzine and oxazolone lead to a marked reduction in epidermal LC numbers, but in caspase-1-deficient mice this reduction was not observed. Moreover, although intradermal injection of TNF-alpha (50 ng) induced epidermal LC migration in WT mice, this cytokine failed to induce LC migration in caspase-1-deficient mice. Intradermal IL-1 beta (50 ng) caused a similar reduction in epidermal LC numbers in both WT and caspase-1-deficient mice, indicating that, given an appropriate signal, caspase-1-deficient epidermal LC are capable of migration. Contact hypersensitivity to both 2,4-dinitrofluorobenzine and oxazolone was inhibited in caspase-1-deficient mice, indicating a functional consequence of the LC migration defect. In organ culture the caspase-1 inhibitor Ac-YVAD-cmk, but not control peptide, potently inhibited the epidermal LC migration that occurs in this system, and reduced spontaneous migration of LC was observed in skin derived from caspase-1-deficient mice. Moreover, Ac-YVAD-cmk applied to BALB/c mouse skin before application of contact sensitizers inhibited LC migration and contact hypersensitivity in vivo. Taken together, these data indicate that caspase-1 may play a central role in the regulation of LC migration and suggest that the activity of this enzyme is amenable to control by specific inhibitors both in vivo and in vitro.     

The three-component signalling system HbpS–SenS–SenR as an example of a redox sensing pathway in bacteria

The two-component system SenS–SenR and the extracellular HbpS protein of the cellulose degrader Streptomyces reticuli have been shown to act in concert as a novel system which detects redox stress. In vivo and in vitro experiments have led to the hypothesis that HbpS binds and degrades heme, communicating the extracellular presence of heme and oxidative stress to the membrane-embedded sensor histidine kinase SenS via a bound iron. The response regulator SenR would then up-regulate downstream signalling cascades, leading to the appropriate gene expression levels for bacterial survival in an oxidative environment. Sequence analysis has shown that homologs of HbpS and SenS–SenR exist in a number of ecologically and medically relevant bacterial species, suggesting the existence of a previously undescribed bacterial oxidative stress-response pathway common to both Gram-negative and Gram-positive bacteria. The presented report reviews the current knowledge of the function of this novel protein family consisting of an accessory protein and its cognate two-component system, which could be more properly described as a three-component system.