Minor triterpenes from Orthopterygium huancuy

Two new triterpenes, 3-oxo-20-hydroxy-lupan-28-oic and 3β,6β-dihydroxy-olean-18-en-28-oic acids, along with oleanonic, morolic and sumaresinolic acids have been isolated from Orthopterygium huancuy. The structure of the first new terpene was determined by single crystal X-ray analysis.     

CHARACTERIZATION OF OSTREOPSIS AND COOLIA (DINOPHYCEAE) ISOLATES IN THE WESTERN MEDITERRANEAN SEA BASED ON MORPHOLOGY,TOXICITY AND INTERNAL TRANSCRIBED SPACER 5.8S rDNA SEQUENCES1

Several isolates of epiphytic dinoflagellates belonging to the genera Ostreopsis Schmidt and Coolia Meunier from the western Mediterranean Sea were examined by LM and EM, toxicity assays, and internal transcribed spacer (ITS) regions of nuclear rDNA, and 5.8S rDNA were sequenced. Morphological comparisons based on the analyses of cell shape, size, thecal plates, and surface ornamentation revealed two distinct species in the western Mediterranean: O. cf. siamensis Schmidt from the Catalan, Andalusian, and Sicilian coasts and O. ovata Fukuyo from the Ligurian coast, southern Tyrrhenian Sea, and Balearic Islands. Both Ostreopsis species were toxic; however, no differences in toxicity were detected between the two Ostreopsis species. Coolia monotis Meunier was nontoxic. The morphological studies were supported by phylogenetic analyses; all western Mediterranean isolates of O. cf. siamensis showed ITS and 5.8S rDNA sequences identical to each other and so did those of O. ovata, whereas high genetic diversity was detected between the western Mediterranean and Asian isolates of O. ovata. The nucleotide sequence analyses of the C. monotis strains showed that all C. monotis isolates from Europe formed a homogeneous clade. Further, the genetic diversity was high between the European and Asian C. monotis isolates. In this study, genetic markers combined with morphology and toxicity analyses was useful in the taxonomic and phylogenetic studies of the Ostreopsidaceae in a temperate area.     

Prothymosin alpha expression in lymph nodes and tonsils: an optical and ultrastructural study.

Prothymosin alpha (ProT alpha) distribution in human and rat lymph nodes and human tonsils was studied by means of immunohistochemical methods, using specific antibodies raised against thymosin alpha 1. We observed ProT alpha immunoreactivity in lymphoid cells of the germinal centers both in humans and rats. In human tonsils, positive cells were also seen in the basal layer of the surface epithelium. These results support the hypothesis that ProT alpha expression is restricted to actively proliferating cells. Immunoelectron microscopy revealed that ProT alpha was located in the nucleus, mainly in the border between euchromatin and heterochromatin.     

Self-organizing tree-growing network for the classification of protein sequences.

The self-organizing tree algorithm (SOTA) was recently introduced to construct phylogenetic trees from biological sequences, based on the principles of Kohonen''s self-organizing maps and on Fritzke''s growing cell structures. SOTA is designed in such a way that the generation of new nodes can be stopped when the sequences assigned to a node are already above a certain similarity threshold. In this way a phylogenetic tree resolved at a high taxonomic level can be obtained. This capability is especially useful to classify sets of diversified sequences. SOTA was originally designed to analyze pre-aligned sequences. It is now adapted to be able to analyze patterns associated to the frequency of residues along a sequence, such as protein dipeptide composition and other n-gram compositions. In this work we show that the algorithm applied to these data is able to not only successfully construct phylogenetic trees of protein families, such as cytochrome c, triosephophate isomerase, and hemoglobin alpha chains, but also classify very diversified sequence data sets, such as a mixture of interleukins and their receptors.     

The microbiological transformation of epicandicandiol,ent-7α,18-dihydroxykaur-16-ene,by Gibberella fujikuroi

Incubation of ent-7α,18-dihydroxykaur-16-ene with Gibberella fujikuroi affords ent-7α,18,19-trihydroxykaur-16-ene and ent-7α,18-dihydroxykaur-16-en-19-oic acid. There was no transformation into 7,18-dihydroxykaurenolide.     

Vitamin D receptor ontogenesis in rat liver

Vitamin D through its receptor (VDR) plays a major role in bone mineral metabolism. However, VDR is also present in a variety of cell lines as well as in numerous tissues, suggesting other functions of the hormone beyond bone metabolism and mineral homeostasis. At the liver level, it has been shown that vitamin D induces numerous changes (i.e. enzyme activity level, stimulation of some metabolic pathways and stimulation of the normal liver recovery after partial hepatectomy). However, some works did not find VDR in the liver, and also used liver tissue as a negative control of VDR gene expression. In this paper, we examined fetal, neonatal and adult rat tissues for the presence of VDR using a sensitive RT-PCR technique and immunohistochemistry. We found VDR mRNA and VDR protein in rat liver at all different periods of rat life. Thus, we suggest that some of the actions of vitamin D on liver could be mediated at the genomic level through the VDR, and that the use of this tissue as a negative control of VDR gene expression is clearly inappropriate. Accepted: 7 June 1999     

Application of a high-throughput HPLC-MS/MS assay to Arabidopsis mutant screening; evidence that threonine aldolase plays a role in seed nutritional quality

Beyond their essential function as the building blocks of proteins, amino acids contribute to many aspects of plant biochemistry and physiology. Despite this, there are relatively large gaps in our understanding of the biochemical pathways and regulation of amino acid synthesis in plants. A rapid (1.5 min versus 20-90 min for standard methods) HPLC-MS/MS assay for separating 19 amino acids was developed for quantifying levels of free amino acids in plant tissue. This assay was used to determine the free amino acid content in the seeds of 10,000 randomly mutagenized Arabidopsis lines, and 322 Arabidopsis lines with increased levels of one or more amino acids were identified. The heritability of the mutant phenotype was confirmed for 43 lines with increased seed levels of the aspartate-derived amino acids Ile, Lys, Thr, or Met. Genetic mapping and DNA sequencing identified a mutation in an Arabidopsis threonine aldolase (AT1G08630, EC 4.1.2.5) as the cause of increased seed Thr levels in one mutant. The assay that was developed for this project has broad applicability to Arabidopsis and other plant species.     

Enalapril and captopril enhance glutathione-dependent antioxidant defenses in mouse tissues

The effect of enalapril and captopril on total glutathione content (GSSG + GSH) and selenium-dependent glutathione peroxidase (Se-GPx) and glutathione reductase (GSSG-Rd) activities was investigated in mouse tissues. CF-1 mice (4-mo-old females) received water containing enalapril (20 mg/l) or captopril (50 mg/l) for 11 wk. Enalapril increased GSSG + GSH content (P < 0.05) in erythrocytes (147%), brain (112%), and lung (67%), and captopril increased GSSG + GSH content in erythrocytes (190%) and brain (132%). Enalapril enhanced Se-GPx activity in kidney cortex (42%) and kidney medulla (23%) and captopril in kidney cortex (30%). GSSG-Rd activity was enhanced by enalapril in erythrocytes (21%), brain (21%), liver (18%), and kidney cortex (53%) and by captopril in erythrocytes (25%), brain (19%), and liver (34%). In vitro erythrocyte oxidant stress was evaluated by thiobarbituric acid-reactive substances (TBARS) production (control 365 +/- 11, enalapril 221 +/- 26, captopril 206 +/- 17 nmol TBARS x g Hb(-1) x h(-1); both P < 0.05 vs. control) and phenylhydrazine-induced methemoglobin (MetHb) formation (control 66.5 +/- 3.5, enalapril 52.9 +/- 0.4, captopril: 56.4 +/- 2.9 micromol MetHb/g Hb; both P < 0.05 vs. control). Both angiotensin-converting enzyme inhibitor treatments were associated with increased nitric oxide production, as assessed by plasma NO-(3) + NO-(2) level determination (control 9.22 +/- 0.64, enalapril 13.7 +/- 1.9, captopril 17.3 +/- 3.0 micromol NO-(3) + NO-(2)/l plasma; both P < 0.05 vs. control). These findings support our previous reports on the enalapril- and captopril-induced enhancement of endogenous antioxidant defenses and include new data on glutathione-dependent defenses, thus furthering current knowledge on the association of ACE inhibition and antioxidants.     

Tendency for oxidation of annelid hemoglobin at alkaline pH and dissociated states probed by redox titration

The redox titration of extracellular hemoglobin of Glossoscolex paulistus (Annelidea) was investigated in different pH conditions and after dissociation induced by pressure. Oxidation increased with increasing pH, as shown by the reduced amount of ferricyanide necessary for the oxidation of hemoglobin. This behavior was the opposite of that of vertebrate hemoglobins. The potential of half oxidation (E_{1 / 2}) changed from − 65.3 to + 146.8 mV when the pH increased from 4.50 to 8.75. The functional properties indicated a reduction in the log P₅₀ from 1.28 to 0.28 in this pH range. The dissociation at alkaline pH or induced by high pressure, confirmed by HPLC gel filtration, suggested that disassembly of the hemoglobin could be involved in the increased potential for oxidation. These results suggest that the high stability and prolonged lifetime common to invertebrate hemoglobins is related to their low tendency to oxidize at acidic pH, in contrast to vertebrate hemoglobins.