Blueberry flavonoids inhibit matrix metalloproteinase activity in DU145 human prostate cancer cells.

Regulation of the matrix metalloproteinases (MMPs), the major mediators of extracellular matrix (ECM) degradation, is crucial to regulate ECM proteolysis, which is important in metastasis. This study examined the effects of 3 flavonoid-enriched fractions (a crude fraction, an anthocyanin-enriched fraction, and a proanthocyanidin-enriched fraction), which were prepared from lowbush blueberries (Vaccinium angustifolium), on MMP activity in DU145 human prostate cancer cells in vitro. Using gelatin gel electrophoresis, MMP activity was evaluated from cells after 24-hr exposure to blueberry fractions. All fractions elicited an ability to decrease the activity of MMP-2 and MMP-9. Of the fractions tested, the proanthocyanidin-enriched fraction was found to be the most effective at inhibiting MMP activity in these cells. No induction of either necrotic or apoptotic cell death was noted in these cells in response to treatment with the blueberry fractions. These findings indicate that flavonoids from blueberry possess the ability to effectively decrease MMP activity, which may decrease overall ECM degradation. This ability may be important in controlling tumor metastasis formation.     

Subtype-specific increase in G-protein alpha-subunit mRNA by interleukin 1 beta

The guanine nucleotide regulatory proteins (G-proteins) which are substrates for ADP-ribosylation by pertussis toxin (alpha i-1, alpha i-2, alpha i-3 and alpha o) transduce a variety of hormonal signals. Endothelial cells express mRNA for three alpha i subtypes although the level of alpha i-1 mRNA is very low. Interleukin 1 beta (IL 1 beta), a pleiotropic inflammatory mediator which stimulates a complex series of responses in human endothelial cells leading to increased coagulation and platelet adhesion, increases expression of one subtype of alpha i (alpha i-2) mRNA in human endothelial cells as determined by Northern blot analysis without affecting the level of mRNA for other alpha-subunits. These studies show that mRNA levels for alpha i subtypes are independently regulated, suggesting that there may be subtype specificity in the cell's requirements for the Gi class of signal-transducing proteins.     

13C NMR spectroscopy of Methanobacterium thermoautotrophicum. Carbon fluxes and primary metabolic pathways

The flux of 13C-labeled carbons from the soluble metabolite 2,3-cyclopyrophosphoglycerate (CPP), a novel compound found in high concentrations exclusively in methanobacteria and methanobrevibacter, into carbohydrate-containing material has been deduced by solid-state 13C NMR spectroscopy which strongly argues for a role in gluconeogenesis for this unique metabolite. The turnover rates, but not the steady-state levels, of CPP labeled by 13CO2 or [13C]acetate depend dramatically on cell growth conditions. When the demand for carbohydrate synthesis is reduced (i.e. in stationary phase), the rates of CPP biosynthesis and degradation decrease 10-fold, and the disaccharide alpha, alpha-trehalose accumulates. Valinomycin, a metabolic inhibitor of Methanobacterium thermoautotrophicum growth, does not affect steady-state levels of CPP, but does decrease 13C uptake into the CPP pool. The effects of these different conditions on CPP labeling suggest stringent regulation of CPP linked to cellular metabolism. Labeling of CPP by [6-(13)C]glucose, which does not serve as an energy or carbon source for this organism, provides strong evidence that glucose is cleaved by the reverse of the gluconeogenesis pathway. This metabolic pathway linking glucose with triose phosphate type precursors and an analysis of the 13C NMR spectrum of CPP labeled by incubating cells with [U-13C]glucose have established that in vivo phosphoenolpyruvate synthetase must be reversible.     

2'-Nor-cGMP: a seco-cyclic nucleotide with powerful anti-DNA-viral activity

As part of our study of antiherpetic acyclonucleosides, we synthesized a cyclic GMP analog, 9-[(2-hydroxy-1,3,2-dioxaphosphorinan-5-yl)oxymethyl]guanine P-oxide, sodium salt (2'-nor-cGMP), and discovered its potent and broad spectrum anti-DNA-viral activities. 2'-Nor-cGMP inhibits the replication of many DNA viruses, including herpes simplex virus, human cytomegalovirus, vaccinia, SV40, and adenovirus, but does not inhibit RNA viruses. In plaque reduction studies this potent antiviral agent is also approximately 10-fold more potent than 9-(1,3-dihydroxy-2-propoxymethyl)guanine (2'NDG) against varicella-zoster virus and inhibits cell transformation by bovine papilloma virus. Unlike 2'NDG, the potent activity of 2'-nor-cGMP against herpes virus is not dependent upon the action of virus-specified thymidine kinase. Intercellular metabolism of 2'-nor-cGMP produced small amounts of 2'NDG triphosphate which were insufficient to account for the antiviral activity observed, implying that this potent anti-DNA-viral agent operates by a mechanism different from that of known acyclonucleosides.     

Growth hormone stimulation of amino acid transport and utilization by the perfused rat liver.

The effects of growth hormone, administered in vivo or added in vitro, on amino acid transport and utilization have been studied in perfused livers of normal and hypophysectomized rats. A perfusion system employing a nonrecirculating medium was used in all of the studies. Two nonmetalbolizable amino acid analogues, alpha-aminoisobutyric acid (AIB) and 1-aminocyclopentane carboxylic acid (cycloleucine) were used to study transport. Accumulation of AIB increased linearly over a 60-min perfusion period, reaching distribution ratios of between 1 and 2 for both groups of animals. Treatment of both normal and hypophysectomized rats with growth hormone 60 min prior to the start of perfusion increased AIB distribution ratios by up to 84 and 108%, respectively. Accumulation of cycloleucine was linear for only about 20 min of perfusion and then plateaued. Steady state distribution ratios of this analogue ranged between 1 and 2 for both groups of animals. Growth hormone treatment had no apparent effect on the time necessary to reach these steady state levels, but significantly increased them in livers of both normal and hypophysectomized rats by 16 and 42%, respectively. Studies designed to analyze the kinetic properties of these hormone effects revealed that growth hormone treatment caused 2-fold i-crease in the maximum velocities of both the AIB and cycloleucine transport systems. The substrate concentration for half-maximal transport velocity was increased slightly for both systems by growth hormone. Direct effects of growth hormone were demonstrated in studies where livers of hypophysectomized rats were perfused under conditions simulationg those of experiments in which the hormone was administered in vivo. Following an initial 45-min period of perfusion the medium during the 20 min. Growth hormone added to the medium during the entire 65-min perfusion at a concentration of 1 mug per ml caused a 30% increase in the cycloleucine distribution ratio. Under similar experimental conditions growth hormone directly stimulated three hepatic pathways of amino acid utilization: (a) incorporation of [14C]valine into protein, (b) urea formation and (c) conversion of 14-C-amino-acids to labeled glucose. Intracellular concentrations of seven amino acids, including threonine, serine, proline, glycine, alanine, lysine, and arginine, were increased significantly in livers perfused with medium containing growth hormone...     

Conjugation of metallothionein to a murine monoclonal antibody

A method of conjugation of the metal-binding protein, metallothionein, to an anticarcinoma murine monoclonal antibody, B72.3, and its F(ab')2 fragment has been developed utilizing the heterobifunctional crosslinking reagent, succinimidyl 4-(N-maleimidomethyl)-cyclohexane 1-carboxylate. This crosslinking reagent is first reacted with the free amines on the immunoglobulin. After removal of unreacted crosslinker, conjugation is affected through a sulfhydryl group on metallothionein. Under the conditions employed all immunoglobulin aggregates contained metallothionein. The degree of undesired aggregation is directly proportional to the number of metallothioneins attached to the immunoglobulin. This aggregation can be controlled by the amount of crosslinker and metallothionein presented to the immunoglobulin. The immunoglobulin conjugate retains full immunoreactivity and can be readily purified from the unreacted metallothionein and high molecular weight aggregates. The metallothionein-B72.3 conjugate functions as an efficient and stable chelator of radiometals. Thus metallothionein-monoclonal antibody conjugates have potential utility in cancer diagnosis and therapy.     

A Potent,Selective, Non-Substrate Inhibitor of HSV-I Thymidine Kinase: (±)-9-[[(Z)-2-(Hydroxymethyl)Cyclohexyl]Methyl]Guanine and Related Compounds

Abstract

The title compound was prepared and found to be a potent and selective inhibitor of HSV-I thymidine kinase. This compound delayed the reactivation of latent virus from explanted mouse ganglia but exacerbated the primary HSV-I infection in mice.     

Depth-dependent indicators of algal turf herbivory throughout the Main Hawaiian Islands

Herbivorous fish are key to maintaining a balance between coral and algae on reefs, where reefs with greater herbivore biomass often show lower algal cover. For reefs worldwide, algal turf cover is expanding and is increasingly used as an indicator of disturbance. Water depth affects reef fish composition; thus, it may be expected that herbivory could also differ by depth. We examined relationships between algal turf cover and biomass (g m⁻²), density (# m⁻²) and size (cm) of herbivore groups (grazers, browsers and scrapers) across shallow (< 6 m), mid (6–18 m) and deep (18–30 m) coral reefs in the Main Hawaiian Islands. We find that across all depth classes, algal turf cover decreased with increasing grazer and scraper density, with steeper relationships observed at mid and deep reefs than in shallow reefs. In contrast, algal turf cover slightly increased with increasing grazer and browser biomass at deep reefs. Considering fish size, algal turf cover increased with larger grazer and scrapers at mid and deep reefs. The results indicate that herbivorous fish density, rather than biomass, is a better indicator of reductions in algal turf cover and resulting coral-algal balance on Hawaiian reefs, where smaller fish exert greater top-down control on cover than larger fish. Despite significant differences in herbivorous fish compositions, length-frequency distributions and fishing intensities across depth, algal turf cover remains similar across depths. Increases in fishing would have a disproportionately negative impact in deep than shallow reefs due to a lower overall fish density, where grazing functions in deep reefs are maintained by significantly fewer and smaller grazers and browsers, and larger scrapers, than in shallow reefs. Developing an understanding of patterns of algal turf herbivory by depth is important to understanding the spatial scale at which herbivory and regime shifts operate.

     

Impacts of remotely sensed environmental drivers on coral outplant survival

Globally, coral reefs are degrading due to a variety of stressors including climate change and pollution. Active restoration is an important effort for sustaining coral reefs where, typically, coral fragments are outplanted onto degraded reefs. Coral outplants, however, can experience mortality in response to a range of stressors. We pair results of outplant monitoring observations with satellite‐based measurements of multiple oceanographic variables to estimate the relative importance of each driver to coral outplant survival. We find that when considering mean environmental conditions experienced by outplants during the monitoring period, particulate organic carbon (POC) levels are most important in determining outplant survival, with certain levels of POC beneficial for outplants. Sea surface temperature anomalies (SSTA) are also important determinants of outplant survival, where survival is greatest in regions with minimal or slightly negative anomalies. Survival also increases with increasing distance to land, likely due to a reduction in negative ridge‐to‐reef effects on coral outplants. When considering the range (min–max) of environmental conditions experienced during the monitoring period, large fluctuations in photosynthetically active radiation (PAR) and POC are most important in determining outplant survival. Increasing outplant depth can help to counter the negative impacts of large fluctuations in environmental variables. We find that a variety of remotely sensed oceanographic variables have significant impacts on survival and should be considered in coral restoration planning to help evaluate potential restoration sites and ultimately maximize coral outplant survival.