Inhibitory Effect of Ascorbic Acid on the Proliferation and Invasion of Hepatoma Cells in Culture

Effect of ascorbic acid (AsA) on the proliferation and invasion of rat ascites hepatoma AH109A cells was investigated by measuring [³H]thymidine incorporation into acid-insoluble fraction of the cells and by co-culturing the hepatoma cells with rat mesentery-derived mesothelial cells, respectively. AsA suppressed the invasion of AH109A cells in a dose-dependent manner at concentrations of 62.5–500 μM, while it inhibited the proliferation of the cells at higher concentrations of 250 and 500 μM. Hepatoma cells previously cultured with hypoxanthine (HX) and xanthine oxidase (XO) or with hydrogen peroxide showed increased invasive activities. AsA suppressed the reactive oxygen species-potentiated invasive capacity by simultaneously treating AH109A cells with AsA, HX and XO or with AsA and hydrogen peroxide. Furthermore, AsA reduced the intracellular peroxide levels in AH109A cells. These results suggest that the antioxidative property of AsA may be involved in its anti-invasive action on hepatoma cells.     

Innate immune response to viral infection

In viral infections the host innate immune system is meant to act as a first line defense to prevent viral invasion or replication before more specific protection by the adaptive immune system is generated. In the innate immune response, pattern recognition receptors (PRRs) are engaged to detect specific viral components such as viral RNA or DNA or viral intermediate products and to induce type I interferons (IFNs) and other pro-inflammatory cytokines in the infected cells and other immune cells. Recently these innate immune receptors and their unique downstream pathways have been identified. Here, we summarize their roles in the innate immune response to virus infection, discrimination between self and viral nucleic acids and inhibition by virulent factors and provide some recent advances in the coordination between innate and adaptive immune activation.     

Histochemical staining of exogenous chromium and iron with the catalytic oxidation of phenylamines

Summary A highly sensitive and specific method for staining exogenous chromium and iron in tissues is described. This method is superior to conventional complex-forming methods with regard to its sensitivity and specificity for these metals. The staining reaction is based on the metalcatalysed oxidation of phenylamines. Tissue sections were incubated in a medium containing hydrogen peroxide and phenylamines, p-phenylenediamine or phenylhydrazine. Results obtained from test-tube experiments concerning the catalytic activities of metals indicated that the staining reactions depends on the activities of metals in tissues.     

Accumulation of Phenylpropanoids in Cotyledons of Morning Glory (Pharbitis nil) Seedlings during the Induction of Flowering by Poor Nutrition

Flowering of Pharbitis nil strain Violet is induced in continuouslight under poor nutritional conditions. High-performance liquidchromatography of extracts of the cotyledons revealed that twocompounds in addition to chlorogenic acid accumulate under suchconditions. The compounds were identified as pinoresinol glucosideand p-coumaroylquinic acid. The endogenous levels of these phenylpropanoidswere correlated with the flowering response when nutrition waspoor. However, activation of phenylpropanoid biosynthesis seemednot to be essential for the induction of flowering. (Received May 17, 1993; Accepted July 26, 1993)     

Patterns of Endopolyploidy During Seedling Development in Cabbage (Brassica oleracea L.)

Flow cytometric analysis of nuclear DNA contents of somatictissues of cabbage (Brassica oleracea L.) has revealed extensiveendopolyploidization, resulting in tissues that comprise mixturesof cells with different DNA contents, ranging from 2C to 16C.Patterns of endopolyploidy are specific to each developmentalstage. Multiple polyploidy was not present in the embryos ofdry seeds. Rapid endoreduplication occurred in the radicle andthe hypocotyl of the embryos during seed germination. Furtherendoreduplication cycles were detected in all tissues exceptthose of the shoot tips. In five cabbage cultivars tested, seedlingscontained cells of four ploidy levels, corresponding to 2C,4C, 8C and 16C. Multiploidy may be an integral part of differentiationprograms in cabbage plants. The biological significance of endoreduplicationin cabbage plants is discussed. Copyright 2001 Annals of BotanyCompany Cabbage (Brassica oleracea L.), endopolyploidy, endoreduplication, flow cytometry     

Influence of maceration temperature in red wine vinification on extraction of phenolics from berry skins and seeds of grape (Vitis vinifera)

The extraction of phenolics from berry skins and seeds of the grape, Vitis vinifera cv. Cabernet Sauvignon, during red wine maceration and the influence of different temperature conditions (cold soak and/or heating at the end of maceration) were examined. Phenolics contained mainly in berry skins, viz., anthocyanin, flavonol, and epigallocatechin units within proanthocyanidins, were extracted during the early stage of maceration, whereas those in seeds, viz., gallic acid, flavan-3-ol monomers, and epicatechin-gallate units within proanthocyanidins, were gradually extracted. In addition to their localization, the molecular size and composition of the proanthocyanidins possibly influenced the kinetics of their extraction. Cold soak reduced the extraction of phenolics from the seeds. Heating at the end of maceration decreased the concentration of proanthocyanidins. Thus, modification of the temperature condition during maceration affected the progress of the concentration of phenolics, resulting in an alteration of their make-up in the finished wine.     

Population genetic structure and demography of Magnolia kobus: variety borealis is not supported genetically

Species delimitations by morphological and by genetic markers are not always congruent. Magnolia kobus consists of two morphologically different varieties, kobus and borealis. The latter variety is characterized by larger leaves than the former. For the conservation of M. kobus genetic resources in natural forests, the relationships between morphological and genetic variation should be clarified. We investigated variations in nuclear microsatellites, chloroplast DNA (cpDNA) sequences and leaf morphological traits in 23 populations of M. kobus over the range of species. Two genetically divergent lineages, northern and southern were detected and their geographical boundary was estimated to be at 39°N. The northern lineage consisted of two genetic clusters and a single cpDNA haplotype, while the southern one had multiple genetic clusters and cpDNA haplotypes. The northern lineage showed significantly lower genetic diversity than the southern. Approximate Bayesian computation indicated that the northern and southern lineages had experienced, respectively, population expansion and long-term stable population size. The divergence time between the two lineages was estimated to be 565,000 years ago and no signature of migration between the two lineages after divergence was detected. Ecological niche modeling showed that the potential distribution area in northern Japan at the last glacial maximum was very small. It is thus considered that the two lineages have experienced different population histories over several glacial-inter-glacial cycles. Individuals of populations in the central to northern part of Honshu on the Sea of Japan side and in Hokkaido had large leaf width and area. These leaf characteristics corresponded with those of variety borealis. However, the delimitation of the northern and southern lineages detected by genetic markers (39°N) was not congruent with that detected by leaf morphologies (36°N). It is therefore suggested that variety borealis is not supported genetically and the northern and southern lineages should be considered separately when identifying conservation units based not on morphology but on genetic markers.

     

Phosphorylation and Inactivation of Brain Glycogen Synthase by a Multifunctional Calmodulin-Dependent Protein Kinase

Glycogen synthase was partially purified from canine brain to about 70% purity. The purified enzyme showed differences from the properties of the skeletal muscle enzyme with respect to molecular weights of the holoenzyme and subunit and phosphopeptide mapping. The multifunctional calmodulin-dependent protein kinase from the brain phosphorylated brain glycogen synthase with concomitant inactivation of the enzyme. Although about 1.3 mol of phosphate/mol subunit was maximally incorporated into glycogen synthase, 0.4 mol of phosphate/mol subunit was sufficient for the maximal inactivation of the enzyme. The results indicate that brain glycogen synthase is regulated in a calmodulin-dependent manner similarly to the skeletal muscle enzyme, but that the brain enzyme is different from the skeletal muscle enzyme.     

Importance of renal mitochondria in the reduction of TEMPOL,a nitroxide radical

Spin probing methods using an electron spin resonance (ESR) spectrometer are used extensively and bring us a lot of information about in vivo redox mechanisms. However, the in vivo reducing mechanisms of exogenous nitroxide radicals, which serve as typical spin probing reagents are not clear. To clarify this, we examined the sequential kinetics of a spin probe, 4-hydroxy 2,2,6,6-tetramethylpiperidine-N-oxyl (TEMPOL) in the in vivo organs, tissue homogenates and subcellular fractions of kidney and liver using an in vivo and X-band ESR spectrometers. As a parameter of reducing activity, we calculated the half-life of TEMPOL from the decay curve of ESR signal intensity. The half-life of TEMPOL in the whole organs and homogenates of the kidney was significantly shorter than that of the liver, this indicates that the kidney has more reducing activity against TEMPOL as compared to the liver. Subcellular fractional studies revealed that this reducing activity of the kidney mainly exists in the mitochondria. Contrarily, in addition to reduction in the mitochondria, TEMPOL in the liver was reduced by the microsome and cytosol.     

Molecular cloning and expression ofThiobacillus ferrooxidans chromosomal ribulose bisphosphate carboxylase genes inEscherichia coli

The genes coding ford-ribulose-1,5-bisphosphate carboxylase (RuBPCase) from an iron-oxidizing bacterium,Thiobacillus ferrooxidans, were cloned into anEscherichia coli plasmid, pUC18. The recombinant plasmid, termed pTR11, contained a 4.0-kb PstI fragment including the entire coding regions for both large and small subunits of RuBPCase.Escherichia coli carrying pTR11 did not show any CO₂-fixing activity. However, a derivative plasmid with an appropriate deletion, which was placed under the control of atac promoter, conferred ribulose bisphosphate-dependent CO₂-fixing activity on the host cell. Analysis of gel-filtration chromatography of the RuBPCase synthesized inE. coli revealed that it had a hexadecameric form like the native enzyme ofT. ferrooxidans.