Reactive oxygen species induce expression of vascular endothelial growth factor in chondrocytes and human articular cartilage explants

Vascular endothelial growth factor (VEGF) promotes cartilage-degrading pathways, and there is evidence for the involvement of reactive oxygen species (ROS) in cartilage degeneration. However, a relationship between ROS and VEGF has not been reported. Here, we investigate whether the expression of VEGF is modulated by ROS.     

Short-term effects of boron, germanium and high light intensity on membrane permeability in boron deficient leaves of sunflower

With the ultimate purpose of clarifying the mechanism for aluminium (Al) toxicity and for Al tolerance, we tried to isolate cDNAs whose expression is induced by Al treatment and phosphate (P_i) starvation. We performed P_i starvation and Al treatment (two-step treatment) on suspension-cultured cells of Nicotiana tabacum L. cv. Samsun and then constructed a cDNA library using poly(A)⁺-RNA derived from the treated cells. Four independent cDNA clones (pAL 111, 139, 141 and 142) were isolated from the library by differential screening. Northern blot hybridization analysis indicated that the expression of these clones was induced by P_i starvation. Furthermore, we found that pAL 111 and pAL 142 are also induced by Al treatment. The complete cDNA sequencing of these 4 clones was determined. The results indicated that pAL111 is identical to the parA gene of N. tabacum, which is described as an auxin-regulated gene and that pAL142 is highly homologous to the parB gene of N. tabacum whose product has glutathione S-transferase (GST, EC 2.5.1.18) activity. Furthermore, we found a cysteine-rich domain in the amino acid sequence of pAL139. No DNA and deduced amino acid sequences homologous to the pAL141 were found.     

Rat thymic epithelial cells in vitro and in situ: characterization by immunocytochemistry and morphology

A new method for the long-term culture of pure rat thymic epithelial cells was established. The cultures were characterized by immunocytochemistry, electron microscopy and proliferation assays. Non-epithelial thymic cells were eliminated with a reliable and reproducible pre-plating method, by differential trypsin treatment of the cultures and by addition of horse serum to the culture medium instead of fetal calf serum. The final cultures contained more than 95% pure epithelial cells as evidenced by immunostaining for cytokeratin. Ultrastructural studies indicated that these cells are physiologically active epithelial cells with tonofilaments, desmosomes and filopods. The subsets of the thymic epithelial cells in vitro were investigated by comparing their staining pattern with that obtained in situ using several subtype-selective antibodies. Thymic epithelial cells in vitro showed a preferential expression of subcapsular/perivascular and medullary markers. Only few cultivated cells were of cortical origin. In the first to the fourth subcultures, some cells were immunopositive for the thymus hormone/factor thymulin. The proliferation of thymic epithelial cells was stimulated by horse serum and to a lesser extend by fetal calf serum. The adenylate cyclase activators isoproterenol and forskolin, and the glucocorticoid cortisol inhibited the proliferation. Received: 12 May 1995 / Accepted: 13 October 1995     

Negative impacts of high temperatures on growth of black spruce forests intensify with the anticipated climate warming

An increasing number of studies conclude that water limitations and heat stress may hinder the capacity of black spruce (Picea mariana (Mill.) B.S.P.) trees, a dominant species of Canada's boreal forests, to grow and assimilate atmospheric carbon. However, there is currently no scientific consensus on the future of these forests over the next century in the context of widespread climate warming. The large spatial extent of black spruce forests across the Canadian boreal forest and associated variability in climate, demography, and site conditions pose challenges for projecting future climate change responses. Here we provide an evaluation of the impacts of climate warming and drying, as well as increasing [CO₂], on the aboveground productivity of black spruce forests across Canada south of 60°N for the period 1971 to 2100. We use a new extensive network of tree‐ring data obtained from Canada's National Forest Inventory, spatially explicit simulations of net primary productivity (NPP) and its drivers, and multivariate statistical modeling. We found that soil water availability is a significant driver of black spruce interannual variability in productivity across broad areas of the western to eastern Canadian boreal forest. Interannual variability in productivity was also found to be driven by autotrophic respiration in the warmest regions. In most regions, the impacts of soil water availability and respiration on interannual variability in productivity occurred during the phase of carbohydrate accumulation the year preceding tree‐ring formation. Results from projections suggest an increase in the importance of soil water availability and respiration as limiting factors on NPP over the next century due to warming, but this response may vary to the extent that other factors such as carbon dioxide fertilization, and respiration acclimation to high temperature, contribute to dampening these limitations.     

Replanting reduces frog diversity in oil palm

A growing body of literature has demonstrated significant biodiversity losses for many taxa when forest is converted to oil palm. However, no studies have directly investigated changes to biodiversity throughout the oil palm life cycle, in which oil palm matures for 25–30 yr before replanting. This process leads to major changes in the oil palm landscape that likely influence species assemblages and ecosystem function. We compare frog assemblages between mature (21–27‐yr old) and recently replanted (1–2‐yr old) oil palm in Sumatra, Indonesia. Across eighteen 2.25‐ha oil palm plots, we found 719 frogs from 14 species. Frog richness was 31 percent lower in replanted oil palm (nine species) than mature oil palm (13 species). Total frog abundance was 47 percent lower in replanted oil palm, and frog assemblage composition differed significantly between the two ages of oil palm. The majority of frog species were disturbance‐tolerant, although we encountered four forest‐associated frog species within mature oil palm despite a distance of 28 km between our study sites and the nearest extensive tract of forest. Although it is clear that protection of forest is of paramount importance for the conservation of tropical fauna, our results indicate that management decisions within tropical agricultural landscapes also have a profound impact on biodiversity. Practices such as staggered replanting or maintenance of connectivity among mature oil palm patches could help maintain frog diversity in the oil palm landscape.     

Subunit structure of D-galactose dehydrogenase from Pseudomonas saccharophila.

1. D-Galactose dehydrogenase from Pseudomonas saccharophila (molecular weight 102 000) dissociates in 8 M urea into its subunits (molecular weight 25 000) which migrate in polyacrylamide gels, containing 8 M urea, as a single band. 2. The N-terminal residue determination by the dansyl method revealed only serine. 3. The C-terminal group determination with carboxypeptidase A and B indicated the sequence -Tyr-His-Leu. Leucine as the single C-terminal amino acid was confirmed by the tritiation method and by tritiation and subsequent degradation with carboxypeptidases. 4. The fragmentation of D-galactose dehydrogenase (24 mol methionine per mol enzyme) by CNBr resulted in six peptides, as detected in disc electrophoresis and substantiated by end group determination, indicating the identity of the subunits. 5. The treatment of D-galactose dehydrogenase (24 mol lysine and 52 mol arginine per mol enzyme) with trypsin and subsequent peptide mapping showed 21, perhaps 22 peptides, indicating a structure comprising four identical subunits.     

Photoautotrophic cell suspension cultures of periwinkle (Catharanthus roseus (L.) G. Don): Transition from heterotrophic to photoautotrophic growth

Chlorophyllous, heterotrophic periwinkle (Catharanthus roseus (L.) G. Don) cells were capable of sustained photoautotrophic growth in sugar-free B5 medium containing naphthaleneacetic acid and kinetin when provided with a CO₂-enriched atmosphere. An increase in cell fresh weight, first observed approximately 2 weeks after transfer from heterotrophic to photoautotrophic conditions, coincided with the development of maximum chlorophyll content and photosynthetic activity. Electron micrographs revealed that chloroplasts of cells cultured photoautotrophically in continuous light contained large starch granules and exhibited a less extensive thylakoid system than did periwinkle mesophyll chloroplasts. Photoautotrophic cells did not accumulate vindoline or dimeric alkaloids.Abbreviations Chl chlorophyll - dry wt dry weight - fr wt fresh weight - 2,4-D 2,4-dichlorophenoxyacetic acid - NAA naphthaleneacetic acid     

Determination of heteronuclear long-range couplings to heteronuclei in natural abundance by two- and three-dimensional NMR spectroscopy

Summary A method to determine heteronuclear long-range couplings to carbon and nitrogen at natural abundance is presented and applied to two cyclic hexapeptides and the peptidomacrolide FK506. The method is applicable for proton-bearing heteronuclei. By introduction of heteronuclear half-filters in two- or three-dimensional experiments the spectra exhibit an E.COSY pattern when executed without heteronuclear decoupling. The extraction of the heteronuclear coupling constants is therefore independent of linewidth.Incyclo(-Ala-Ala-Ala-Pro-Ala-Pro-) a¹³C-_I-half-filtered TOCSY spectrum yields the³J(H^N-C) coupling constant, which can be used to remove ambiguity in the angle determination from³J(H^N-H). Incyclo(-d-Pro-Phe-Phe-Lys(Z)-Trp-Phe-) a¹⁵N-_I-half-filtered TOCSY was applied to individually assign the diastereotopic -methylene protons via the³J(H-N). In FK506 a 3D-HMQC-TOCSY without heteronuclear decoupling is used to obtain a number of heteronuclear coupling constants to carbons. These values have been applied for the assignment of diastereotopic methylene protons and determination of dihedral angles in the cyclic portion of the molecule.Dedicated to the memory of Professor V.F. Bystrov     

Characterization of a novel N-methyltransferase (NMT) from Catharanthus roseus plants

Young leaves from Catharanthus roseus plants contain a novel N-methyltransferase which transfers the methyl group from S-adenosyl-L-methionine specifically to position 1 of (2R, 3R)-2,3-dihydro-3-hydroxytabersonine, producing the N-methylated product. The enzyme shows a high degree of specificity toward substrates containing a reduced double bond at position 2,3 of tabersonine derivatives but the more substituted N-desmethyldeacetylvindoline did not act as a substrate. The enzyme catalyses the third last step in vindorosine and vindoline biosynthesis, and is associated with chlorophyll-containing fractions in partially purified enzyme preparations. The lack of vindoline accumulation in cell suspension cultures is correlated with the lack of expression of this enzyme activity as well as that of an acetyltransferase which catalyses the last step in vindoline biosynthesis. Neither fungal elicitor treatment of cell line #615 nor transfer to alkaloid production medium resulted in expression of these two enzyme activities, nor was either enzyme activity detected in photoautotrophic or hormone autotrophic cultures. Cell lines #200, 615–767 and 916 could not be induced to produce DAT or NMT enzyme activities.     

Semi-continuous production of sanguinarine and dihydrosanguinarine by Papaver somniferum L. cell suspension cultures treated with fungal homogenate

Papaver somniferum L. (opium poppy) cells were elicited with a Botrytis sp. homogenate and cultured by a semi-continuous process. Elicitation induced synthesis of sanguinarine and dihydrosanguinarine. Significant release of both alkaloids into the culture medium occurred. Medium exchange at 2-day intervals enabled product recovery from spent medium and maintained culture viability. Culture growth was not inhibited by elicitor treatment necessitating sub-culture prior to re-elicitation. Re-elicited cultures displayed an increasing sensitivity (reduced growth rate, higher alkaloid yield) to the elicitor with each successive treatment and did not survive a fourth elicitation.Abbreviations DW dry weight - FW fresh weight - 2,4-D 2,4-dichlorophenoxyacetic acid - SGE sanguinarine - DSGE dihydrosanguinarine Publication 29143 from the National Research Council of Canada