Birefringence effects in multi-core fiber: coupled local-mode theory.

In this paper, we evaluate experimentally and model theoretically the intra- and inter-core crosstalk between the polarized core modes in single-mode multi-core fiber media including temporal and longitudinal birefringent effects. Specifically, extensive experimental results on a four-core fiber indicate that the temporal fluctuation of fiber birefringence modifies the intra- and inter-core crosstalk behavior in both linear and nonlinear optical power regimes. To gain theoretical insight into the experimental results, we introduce an accurate multi-core fiber model based on local modes and perturbation theory, which is derived from the Maxwell equations including both longitudinal and temporal birefringent effects. Numerical calculations based on the developed theory are found to be in good agreement with the experimental data.

Modeling of racetrack-resonator add-drop filters with arbitrary nonlinear directional couplers.

In this Letter we employ the general coupled-mode equations of the nonlinear directional coupler and demonstrate that the switching characteristics of prototypical nonlinear racetrack-resonator structures may differ considerably from those obtained when the standard, generally incorrect, coupled-mode equations are used.

Discovery of a small molecule insulin mimetic with antidiabetic activity in mice.

Insulin elicits a spectrum of biological responses by binding to its cell surface receptor. In a screen for small molecules that activate the human insulin receptor tyrosine kinase, a nonpeptidyl fungal metabolite (L-783,281) was identified that acted as an insulin mimetic in several biochemical and cellular assays. The compound was selective for insulin receptor versus insulin-like growth factor I (IGFI) receptor and other receptor tyrosine kinases. Oral administration of L-783,281 to two mouse models of diabetes resulted in significant lowering in blood glucose levels. These results demonstrate the feasibility of discovering novel insulin receptor activators that may lead to new therapies for diabetes.

Enhancement of antibiotic and secondary metabolite detection from filamentous fungi by growth on nutritional arrays.

AIMS: We asked to what extent does the application of the OSMAC (one strain, many compounds) approach lead to enhanced detection of antibiotics and secondary metabolites in fungi? Protocols for bacterial microfermentations were adapted to grow fungi in nutritional arrays. METHODS AND RESULTS: Protocols for microfermentations of non-sporulating fungi were validated using known antifungal-producing fungi. Detection of antifungal activity was often medium dependent. The effects of medium arrays and numbers of strains on detection of antifungal signals were modelled by interpolation of rarefaction curves derived from matrices of positive and negative extracts. Increasing the number of fermentation media for any given strain increased the probability of detection of growth inhibition of Candida albicans. Increasing biodiversity increased detection of antifungal phenotypes, however, nutritional arrays could partly compensate for lost antibiotic phenotypes when biodiversity was limiting. CONCLUSIONS: Growth and extraction in microtiter plates can enable a discovery strategy emphasizing low-cost medium arrays that can better exploit the metabolic potential of strains. SIGNIFICANCE AND IMPACT OF THE STUDY: Increasing fermentation parameters raise the probability of detecting bioactive metabolites from strains. The protocols can be used to pre-select strains and their growth conditions for scale up that will most likely yield antibiotics and secondary metabolites.

Attenuation of age-dependent oxidative damage to DNA and protein in brainstem of Tg Cu/Zn SOD mice.

Age-dependent accumulation of oxidative DNA and protein damage in brainstem and striatum was assessed in normal and transgenic (tg) mice which overexpress human Cu/Zn superoxide dismutase (h-SOD1). A marker of oxidative DNA damage, 8-hydroxy-2'-deoxyguanosine (oxo8dG), was measured at 3, 12, and 18 months of age in control and tg mice. Cu/Zn SOD, but not MnSOD, activities in brainstems and striata from tg mice were increased compared to controls at all ages. At 18 months, oxo8dG levels were increased by 58% in brainstem and by 21% in striatum of control mice. In the tg mice, brainstem and striatal oxo8dG levels were increased to a lesser extent than in the corresponding controls. Protein oxidation (carbonyl content), was increased by 59% at 18 months in control brainstem, but not in striatum, and the increase was significantly attenuated in the tg mice. In summary, oxidative damage to DNA and protein increased with age in brainstem (and to a lesser extent in striatum), and augmented Cu/Zn SOD activity modified the extent of DNA and protein damage.

Translational inhibition by eIF-2-phospholipid complex in mammalian cell-free systems.

The polypeptide chain initiation factor 2 (eIF-2) binds phospholipid (PL) and becomes a potent inhibitor of translation in hemin-supplemented reticulocyte lysates [De Haro et al. (1986) Proc. Natl. Acad. Sci. USA 83, 6711-6715]. This binding is independent of calcium ions and seems to be specific for phosphatidylinositol or phosphatidylserine; phosphatidic and arachidonic acids are inactive. Like alpha-subunit-phosphorylated eIF-2, eIF-2.PL traps GEF in a non-dissociable eIF-2.PL.GEF complex whereby GEF is no longer able to recycle. Initiation is inhibited when no free GEF is available. Translational inhibition by eIF-2.PL is rescued by equimolar amounts of eIF-2.GEF. On the basis of this stoichiometry, we have estimated that reticulocyte lysates contain about 60 pmol of GEF/ml (60 nM). eIF-2.PL also inhibits translation in cell-free mouse liver extracts and this inhibition is prevented by reticulocyte eIF-2.GEF suggesting that GEF also functions in liver. However, the eIF-2.PL complex does not affect translation in such non-mammalian eukaryotic systems as wheat germ and Drosophila embryos.

Comparison of base-excision repair capacity in proliferating and differentiated PC 12 cells following acute challenge with dieldrin.

Dieldrin, an organochlorine pesticide and known neurotoxicant, is ubiquitously distributed in the environment. Dieldrin depletes brain monoamines in some animal species and is toxic for dopaminergic neurons in vitro. Dieldrin interferes with mitochondrial electron transport and increases generation of superoxide anion. Reactive oxygen species have been shown to produce oxidative lesions to DNA bases, i.e., 8-hydroxy-2'-deoxyguanosine (8-oxodGuo). Accumulation of 8-oxodGuo has been shown to be promutagenic in proliferating cells, and can lead to degeneration in fully differentiated cells. The objective of this study was to determine the effects of dieldrin exposure on the activity of the enzyme responsible for removing 8-oxodGuo, OGG1, from undifferentiated (untreated with NGF) and differentiated (NGF-treated) PC 12 cells. Proliferating PC 12 cells exhibited a mild upregulation of glycosylase activity, reaching a maximum by 1 h and returning to baseline by 6 h. Differentiated (+) NGF cells showed a time-dependent decline in activity reaching a nadir at 3 h with a return towards baseline by 6 h. Levels of the damaged base, 8-oxodGuo, in the differentiated PC12 cells appeared to be regulated by the activity of OGG1. In contrast, levels of the damaged base in actively proliferating cells were independent of the OGG1 activity. This difference between actively dividing and differentiated cells in the regulation of base-excision repair and DNA damage accumulation explains, in part, the vulnerability of postmitotic neurons to oxidative stresses and neurotoxins.

DNA damage, repair, and antioxidant systems in brain regions: a correlative study.

8-Hydroxy-2'-deoxyguanosine (oxo(8)dG) has been used as a marker of free radical damage to DNA and has been shown to accumulate during aging. Oxidative stress affects some brain regions more than others as demonstrated by regional differences in steady state oxo(8)dG levels in mouse brain. In our study, we have shown that regions such as the midbrain, caudate putamen, and hippocampus show high levels of oxo(8)dG in total DNA, although regions such as the cerebellum, cortex, and pons and medulla have lower levels. These regional differences in basal levels of DNA damage inversely correlate with the regional capacity to remove oxo(8)dG from DNA. Additionally, the activities of antioxidant enzymes (Cu/Zn superoxide dismutase, mitochondrial superoxide dismutase, and glutathione peroxidase) and the levels of the endogenous antioxidant glutathione are not predictors of the degree of free radical induced damage to DNA in different brain regions. Although each brain region has significant differences in antioxidant defenses, the capacity to excise the oxidized base from DNA seems to be the major determinant of the steady state levels of oxo(8)dG in each brain region.

Geographical and seasonal influences on the distribution of fungal endophytes in Quercus ilex.

A systematic survey of the endophytic assemblages of Quercus ilex in central Spain has been performed, with the goal of evaluating the importance of geographical and seasonal factors on these fungal communities. Four sampling sites were selected; one of them was sampled twice, in the spring and the autumn. The collected plant material consisted of bark, twigs and leaves from eight trees per site. Fungal strains were isolated with the use of a surface-sterilization method with sodium hypochlorite. A total of 2921 fungal strains grouped into 149 'species' or morphological types were recovered. The 10 dominant species, with isolation frequencies >1.5%, were Pyrenochaeta sp., Periconiella anamorph of Biscogniauxia mediterranea (De Not.) Kuntze, Pseudonectria sp., Cryptosporiopsis quercina Petrak, Alternaria alternata (Fr:) Keissl., two undetermined coelomycetes, Penicillium funiculosum Thom, Diplodia mutila Fr. apud Mont. and Ascochyta sp. Medians of fungal species per tree were significantly different among the sampled sites. The isolation frequencies of the dominant species, as well as other less frequent species, were significantly dependent on the sampling site. The degree of endophytic infection and the diversity of fungal species were significantly higher in the spring. The frequencies of all dominant species at one of the sites depended significantly on the season, except for C. quercina, Acremonium sclerotigenum (F & V Moreau ex Valenta) Gams. and D. mutila. Cluster analysis of the whole endophytic mycoflora of the sampled trees suggested that the geographical factor affects the endophytic distribution patterns more significantly than the seasonal factor.

The X-gal caution in neural transplantation studies.

Cell transplantation into host brain requires a reliable cell marker to trace lineage and location of grafted cells in host tissue. The lacZ gene encodes the bacterial (E. coli) enzyme beta-galactosidase (beta-gal) and is commonly visualized as a blue intracellular precipitate following its incubation with a substrate, "X gal," in an oxidation reaction. LacZ is the "reporter gene" most commonly employed to follow gene expression in neural tissue or to track the fate of transplanted exogenous cells. If the reaction is not performed carefully-with adequate optimization and individualization of various parameters (e.g.. pH, concentration of reagents, addition of chelators, composition of fixatives) and the establishment of various controls--then misleading nonspecific background X-gal positivity can result, leading to the misidentification of cells. Some of this background results from endogenous nonbacterial beta-gal activity in discrete populations of neurons in the mammalian brain; some results from an excessive oxidation reaction. Surprisingly, few articles have empha sized how to recognize and to eliminate these potential confounding artifacts in order to maximize the utility and credibility of this histochemical technique as a cell marker. We briefly review the phenomenon in general, discuss a specific case that illustrates how an insufficiently scrutinized X-gal positivity can be a pitfall in cell transplantation studies, and then provide recommendations for optimizing the specificity and reliability of this histochemical reaction for discerning E. coli beta-gal activity.

Microbial natural products as a source of antifungals.

The vast number and variety of chemotherapeutic agents isolated from microbial natural products and used to treat bacterial infections have greatly contributed to the improvement of human health during the past century. However, only a limited number of antifungal agents (polyenes and azoles, plus the recently introduced caspofungin acetate) are currently available for the treatment of life-threatening fungal infections. Furthermore, the prevalence of systemic fungal infections has increased significantly during the past decade. For this reason, the development of new antifungal agents, preferably with novel mechanisms of action, is an urgent medical need. A selection of antifungal agents in early stages of development, produced by micro-organisms, is summarized in this review. The compounds are classified according to their mechanisms of action, covering inhibitors of the synthesis of cell wall components (glucan, chitin and mannoproteins), of sphingolipid synthesis (serine palmitoyltransferase, ceramide synthase, inositol phosphoceramide synthase and fatty acid elongation) and of protein synthesis (sordarins). In addition, some considerations related to the chemotaxonomy of the producing organisms and some issues relevant to antifungal drug discovery are also discussed.

Rubratoxin B elicits antioxidative and DNA repair responses in mouse brain.

Rubratoxin B (RB) is a mycotoxin with potential neurotoxic effects that have not yet been characterized. Based on existing evidence that RB interferes with mitochondrial electron transport to produce oxidative stress in peripheral tissues, we hypothesized that RB would produce oxidative damage to macromolecules in specific brain regions. Parameters of oxidative DNA damage and repair, lipid peroxidation, and superoxide dismutase (SOD) activity were measured across six mouse brain regions 24 h after administration of a single dose of RB. Lipid peroxidation and oxidative DNA damage were either unchanged or decreased in all brain regions in RB-treated mice compared with vehicle-treated mice. Concomitant with these decreased indices of oxidative macromolecular damage, SOD activity was significantly increased in all brain regions. Oxyguanosine glycosylase activity (OGG1), a key enzyme in the repair of oxidized DNA, was significantly increased in three brain regions--cerebellum (CB), caudate/putamen (CP), and cortex (CX)--but not in the hippocampus (HP), midbrain (MB), and pons/medulla (PM). The RB-enhanced OGG1 catalytic activity in these brain regions was not due to increased OGG1 protein expression, but was a result of enhanced catalytic activity of the enzyme. In conclusion, specific brain regions responded to an acute dose of RB by significantly altering SOD and OGG1 activities to maintain the degree of oxidative DNA damage equal to, or less than, that of normal steady-state levels.

The discovery of enfumafungin, a novel antifungal compound produced by an endophytic Hormonema species biological activity and taxonomy of the producing organisms.

In a screening of natural products with antifungal activity derived from endophytic fungi, we detected a potent activity in a culture belonging to the form-genus Hormonema, isolated from leaves of Juniperus communis. The compound is a new triterpene glycoside, showing an antifungal activity highly potent in vitro against Candida and Aspergillus and with moderate efficacy in an in vivo mouse model of disseminated candidiasis. The agent is especially interesting since its antifungal spectrum and its effect on morphology of Aspergillus fumigatus is comparable to that of the glucan synthase inhibitor pneumocandin B,,, the natural precursor of the clinical candidate MK-0991 (caspofungin acetate). An additional search for other Hormonema isolates producing improved titers or derivatives resulted in the isolation of two more strains recovered from the same plant host showing identical activity. The producing isolates were compared with other non-producing Hormonema strains by DNA fingerprinting and sequencing of the rDNA internal transcribed spacers. Comparison of rDNA sequences with other fungal species suggests that the producing fungus could be an undetermined Kabatina species. Kabatina is a coelomycetous genus whose members are known to produce Hormonema-like states in culture.

Evaluation of different PCR-based DNA fingerprinting techniques for assessing the genetic variability of isolates of the fungus Epicoccum nigrum.

Thirty-six strains of the fungus Epicoccum nigrum, isolated from different substrata and ecosystems of Europe, America and Africa, were analysed using 14 molecular markers included in 5 different genetic fingerprinting techniques: AP-PCR, tDNA-PCR, microsatellite-primed PCR, ARDRA and AFLP. All of the techniques used were able to differentiate the isolates, showing a high genetic diversity within the species. However, the different techniques detected different levels of similarity among the strains; ARDRA shows the most homogeneous results whilst AP-PCR shows the most heterogeneous. The similarity indices achieved for each strain were compared for the different techniques. The distribution obtained by microsatellite-primed PCR was similar to those shown by AP-PCR techniques. tDNA-PCR and AFLP rendered similar distributions, and ARDRA showed remarkably different results from the other techniques. The results also reveal the lack of an overall correlation between geographical or ecological origin of the isolates and their genotypes.

Production of a family of kinase-inhibiting lactones from fungal fermentations.

During the course of our screening for natural products from fungi, extracts of several cultures were found to make a family of related resorcylic acid lactone compounds, which are potent inhibitors of MEK kinase. Comparative and empirical studies of fermentation conditions improved the titers of the compounds of interest. Striking changes in the ratios and amounts of the major and minor compounds in some cases were achieved by manipulations of media composition.

Antimicrobial activity of viridiofungins.

A family of aminoacyl alkyl citrate compounds called viridiofungins, are novel squalene synthase inhibitors. The compounds have broad spectrum fungicidal activity but lack antibacterial activity. Although the compounds inhibit squalene synthase, the first committed step in ergosterol biosynthesis, results presented in this paper show that inhibition of fungal growth is not related to inhibition of ergosterol synthesis.

Discovery of an angiotensin II binding inhibitor from a Cytospora sp. using semi-automated screening procedures.

Cytosporin A, B and C, three antagonists of [125I]-angiotensin II binding to rat adrenal glands were discovered in fermentations of an endophytic Cytospora sp. during routine screening using semi-automated procedures. The most potent of these displayed an IC50 of 1.5-3 microM and was specific for angiotensin II AT2.

Nutritional preferences of a group of Streptosporangium soil isolates.

The nutritional preferences of twelve strains of genus Streptosporangium were studied as a first step in the design of production media to be used in a natural products screening program. The media ingredients included 20 alpha-amino acids, 35 carbon sources (monosaccharides, oligosaccharides, polysaccharides, polyalcohols and others), 22 complex sources (meals, peptones, yeast derivatives, etc.). The growth of these strains in the presence of these culture media ingredients and the production of inhibitory substances against Bacillus subtilis were analyzed in solid-phase cultures. The results show that the isolates studied did not exhibit a common trend in their preference of nutrient sources, either for to support their growth or the production of anti-B. subtilis substances.

Inhibitors of farnesylation of Ras from a microbial natural products screening program.

Mutant ras oncogenes are associated with various human tumors such as pancreas, colon, lung, thyroid, bladder and several types of leukemia. Prenylation of Ras proteins plays a major role in cell proliferation of both normal and cancerous cells. Normal and oncogenic Ras proteins are posttranslationally modified by a farnesyl group that promotes membrane binding. Inhibitors of farnesyl protein transferase (FPTase), the enzyme that catalyzes the prenylation of Ras proteins, inhibit growth of tumor cells. In an effort to identify structurally diverse and unique inhibitors of FPTase, a program devoted to screening of natural products was initiated. This effort led to the identification of 10 different families of compounds, all of which selectively inhibit FPTase with a variety of mechanisms that are reviewed in this manuscript. These compounds originated from the fermentations of a number of microorganisms, either actinomycetes or fungi, isolated from different substrates collected in tropical and temperate areas. A chemotaxonomic discussion on the distribution of each compound among single or different types of microorganisms, either phylogenetically related or unrelated species, is included.