Natural antioxidants attenuate mycolactone toxicity to RAW 264.7 macrophages

Mycobacterium ulcerans produces a macrolide exotoxin, mycolactone which suppresses immune cells activity, is toxic to most cells and the key virulence factor in the pathogenesis of Buruli ulcer disease. Mycolactone is reported to mediate the production of reactive oxygen species in keratinocytes; cells that play critical role in wound healing. Increased levels of reactive oxygen species have been shown to disrupt the well-ordered process of wound repair; hence, the function of wound-healing cells such as macrophages, keratinocytes, and fibroblast could be impaired in the presence of the reactive oxygen species mediator, mycolactone. To ensure regeneration of tissues in chronic ulcers, with proper and timely healing of the wounds, natural antioxidants that can combat the effects of induced reactive oxygen species in wound-healing cells ought to be investigated. Reactive oxygen species activity was determined in mycolactone-treated RAW 264.7 macrophages and the scavenging ability of the antioxidants (ascorbic acid, gallic acid, and green tea kombucha) against mycolactone-induced reactive oxygen species (superoxide anions) was assessed using fluorescein probe (DCF-DA) and nitroblue tetrazolium dye. Cytotoxicity of the antioxidants, mycolactone, and the protective effect of the antioxidants on the cells upon treatment with mycolactone were determined using the Alamar blue assay. The expression levels of endogenous antioxidant enzyme genes (superoxide dismutase, catalase, and glutathione peroxidase) in response to mycolactone-mediated reactive oxygen species were determined using RT-qPCR. Mycolactone induced the production of reactive oxygen species in RAW 264.7 macrophages, and the resulting superoxide anions were scavenged by some of the antioxidants. The selected endogenous antioxidant enzyme genes in the macrophages were upregulated in the presence of the antioxidants and mycolactone. The exogenously supplied ascorbic acid and green tea kombucha offered moderate protection to the macrophages against the toxicity of mycolactone. We conclude that the results provide insights into alternate and adjunct therapeutic approaches in Buruli ulcer treatment, which could significantly attenuate the toxicity of the pathogenic factor; mycolactone.     

Genetic modification of lignin concentration affects fitness of perennial herbaceous plants

Populations of four perennial herbaceous species that were genetically modified for altered lignin content (or associated forage digestibility) by conventional plant breeding were evaluated for two agricultural fitness traits, plant survival and plant biomass, in three Northcentral USA environments for more than 4 years. Reduced lignin concentration or increased digestibility resulted in increased winter mortality in two of four species and reduced biomass in one species. Results from other experiment indicate that these apparent genetic correlations may be ephemeral, suggesting that selection for fitness can be successful within high-digestibility or low-lignin germplasm. Results indicate that perennial plants genetically engineered with altered lignin concentration or composition for use in livestock, pulp and paper, or bioenergy production should be evaluated for fitness in field environments prior to use in agriculture. Received: 12 December 2000 / Accepted: 27 February 2001     

Dissociation of cGMP accumulation and relaxation in myometrial smooth muscle: effects of S-nitroso-N-acetylpenicillamine and 3-morpholinosyndonimine

In guinea pig, primate and man, nitric oxide (NO)-induced regulation of myometrial smooth muscle contraction is distinct from other smooth muscles because cyclic guanosine 3',5'-cyclic monophosphate (cGMP) accumulation is neither necessary nor sufficient to relax the tissue. To further our understanding of the mechanism of action of NO in myometrium, we employed the NO donors, S-nitroso-N-acetylpenicillamine (SNAP), and 3-morpholinosyndonimine (SIN-1) proposed to relax airway smooth muscle by disparate mechanisms involving elevation in intracellular calcium ([Ca(2+)](i)) or cGMP accumulation, respectively. Treatment of guinea pig myometrial smooth muscle with either NO donor at concentrations thought to produce maximal relaxation of smooth muscles resulted in significant elevations in cGMP that were accompanied by phosphorylation of the cGMP-dependent protein kinase substrate vasodilator-stimulated phosphoprotein (VASP), shown here for the first time to be present and phosphorylated in myometrium. Stimulation of myometrial strips with oxytocin (OT, 1 microM) produced an immediate increase in contractile force that persisted in the continued presence of the agonist. Addition of SNAP (100 microM) in the presence of OT relaxed the tissue completely as might be expected of an NO donor. SIN-1 failed to relax the myometrium at any concentration tested up to 300 microM. In Fura-2 loaded myometrial cells prepared from guinea pig, addition of SNAP (100 microM) in the absence of other agonists caused a significant, reproducible elevation of intracellular calcium while SIN-1 employed under the same conditions did not. Our data further support the notion that NO action in myometrium is distinct from that in other smooth muscles and underscores the possibility that discrete regional changes in [Ca(2+)](i), rather than cGMP, signal NO-induced relaxation of the muscle.     

Infection of Sphyrion laevigatum (Copepoda: Sphyriidae) on Genypterus blacodes (Pisces: Ophidiidae) from the Falkland Islands,South Atlantic

The degree of parasitism by the parasitic copepod Sphyrion laevigatum on kingclip Genypterus blacodes in the Falkland Islands was investigated. In a sample of 719 kingclip, ranging in size from 35 to 145 cm total length, the prevalence of infection was 49%. The number of parasites per fish ranged from 0 to 24, with only 2% of the sample having more than 5 parasites. There was a significant increase in mean abundance of parasites with increasing fish age and length (P < 0.01 and P < 0.05, respectively). In addition, there was a significant difference in the prevalence in 1 of 3 geographical areas for the largest size class.     

Peptidic nanoparticles: for cancer diagnosis and therapy

A challenging topic in cancer research is to create drug delivery system that can bring in a specific and noncytotoxic manner a therapeutic compound. Usually, tumor targeting requires very specific compounds. Currently, peptide analogues like somatostatin, neurotensin, or bombesin are used to target G-coupled receptors, which are overexpressed on tumor cells. However, many of those analogues are rapidly degraded in the plasma and are cytotoxic [1–2]. Due to the limited efficiency and high toxicity of conventional chemotherapy different strategies have been developed for non-cytotoxic cancer treatment and cancer localization [3–5]. The recent development in bio-nanotechnology offers new avenues for cancer therapy. A lot of studies have been devoted to nanoparticulate delivery systems (10–100nm) like lipid or polymer particles [6–8]. Due to the nanometer sized of such cargos, the transportation of therapeutic compounds in the blood stream is increased in terms of time circulation. But their surface functionalization to improve drug-targeting properties is usually complicated and rather uneffective. We have recently designed a novel type of functional nanoparticles with regular icosahedral symmetry, mimicking small, rigid viral capsids (Fig. 1 (A)) and a diameter of about 17 nm (Fig. 1 (C)) which self-assemble from single polypeptide chains (Fig. 1 (B)).     

BioRAT: extracting biological information from full-length papers

MOTIVATION: Converting the vast quantity of free-format text found in journals into a concise, structured format makes the researcher's quest for information easier. Recently, several information extraction systems have been developed that attempt to simplify the retrieval and analysis of biological and medical data. Most of this work has used the abstract alone, owing to the convenience of access and the quality of data. Abstracts are generally available through central collections with easy direct access (e.g. PubMed). The full-text papers contain more information, but are distributed across many locations (e.g. publishers' web sites, journal web sites and local repositories), making access more difficult. In this paper, we present BioRAT, a new information extraction (IE) tool, specifically designed to perform biomedical IE, and which is able to locate and analyse both abstracts and full-length papers. BioRAT is a Biological Research Assistant for Text mining, and incorporates a document search ability with domain-specific IE. RESULTS: We show first, that BioRAT performs as well as existing systems, when applied to abstracts; and second, that significantly more information is available to BioRAT through the full-length papers than via the abstracts alone. Typically, less than half of the available information is extracted from the abstract, with the majority coming from the body of each paper. Overall, BioRAT recalled 20.31% of the target facts from the abstracts with 55.07% precision, and achieved 43.6% recall with 51.25% precision on full-length papers.     

Balance between four types of synaptic input for the integrate-and-fire model

We consider the integrate-and-fire model with AMPA, NMDA, GABA(A)and GABA(B)synaptic inputs, with model parameters based upon experimental data. An analytical approach is presented to determine when a post-synaptic balance between excitation and inhibition can be achieved. Secondly, we compare the model behaviour subject to these four types of input, with its behaviour subject to conventional point process inputs. We conclude that point processes are not a good approximation, even away from exact presynaptic balance. Thirdly, numerical simulations are presented which demonstrate that we can treat NMDA and GABA(B)as DC currents. Finally, we conclude that a balanced input is plausible neither pre-synaptically nor post-synaptically for the model and parameters we employed.     

A role for phospholipase D in GLUT4 glucose transporter translocation

Based on recent studies showing that phospholipase D (PLD)1 is associated with intracellular membranes and promotes membrane budding from the trans-Golgi, we tested its possible role in the membrane trafficking of GLUT4 glucose transporters. Using immunofluorescence confocal microscopy, expressed Myc epitope-tagged PLD1 was found to associate with intracellular vesicular structures by a mechanism that requires its N-terminal pleckstrin homology domain. Partial co-localization with expressed GLUT4 fused to green fluorescent protein in both 3T3-L1 adipocytes and Chinese hamster ovary cells was evident. Furthermore, microinjection of purified PLD into cultured adipocytes markedly potentiated the effect of a submaximal concentration of insulin to stimulate GLUT4 translocation to cell surface membranes. Insulin stimulated PLD activity in cells expressing high levels of insulin receptors but no such insulin effect was detected in 3T3-L1 adipocytes. Taken together, these results are consistent with the hypothesis that PLD1 associated with GLUT4-containing membranes acts in a constitutive manner to promote the mechanism of GLUT4 translocation by insulin.