Metabolic pathways involved in the oxidation of isopropanol into acetone by the intact rat

Isopropanol administered in a large (6 g/kg, orally) as well as in a lower dose (1 g/kg, I.P.) is slowly oxidized into acetone by the intact rat. Using two inhibitors, 3 amino-1,2,4-triazole and pyrazole, investigations on the hepatic enzymatic system involved in the oxidation of isopropanol show that catalase does not play an important part in this pathway, contrary to alcohol dehydrogenase which is the major enzyme responsible for this oxidation. Although isopropanol oxidation is mainly catalysed in the liver through alcohol dehydrogenase, no alteration of the hepatic extramitochondrial redox state occurs after the administration of a large as well as of a lower dose of isopropanol. From these experiments it may be concluded that alterations of the liver NAD⁺/NADH ratio, which seem to play an important part in the ethanol induced fatty liver, are not involved in the isopropanol induced one.     

Untersuchungen zur Pollenentwicklung und Pollenschlauchbildung bei höheren Pflanzen

Zusammenfassung In reifen Pollenkörnern der beiden Sommergerstensorten Amsel und Wisa sowie der F₁-Pflanzen, die aus den Sorten-Kreuzungen Impala X Wisa und Union X Wisa hervorgegangen sind, wurde die DNS-Menge der Kerne cytophotometrisch bestimmt. Die Messungen wurden zugleich bei Spermakernen und vegetativen Kernen eines Pollenkorns vorgenommen. Außerdem wurde der DNS-Gehalt von Kernen von Wurzelspitzen-Zellen der Sorten Amsel und Wisa ermittelt.Amsel und Wisa unterscheiden sich signifikant im DNS-Gehalt der Kerne von Wurzelspitzen-Zellen.Die Befunde der Messungen des DNS-Gehalts von vegetativen und Sperma-Kernen bei vier Gerstenformen zeigen, daß zum Zeitpunkt der Anthese die DNS-Replikationsphase bei vegetativen und Sperma-Kernen noch nicht abgeschlossen ist. Der DNS-Gehalt vegetativer Kerne von Wisa ist signifikant niedriger als die entsprechenden Werte der übrigen drei Gerstenformen. Der Verlauf der DNS-Replikation erfolgt bei beiden Spermakernen synchron. Hingegen verläuft die DNS-Replikation bei vegetativen und Sperma-Kernen mit großer Wahrscheinlichkeit nicht gleichsinnig.Im Diskussionsteil wird erstens erläutert, daß bei allen bisher analysierten Pflanzenarten des zwei- oder dreikernigen Pollenkorn-Typs zum Zeitpunkt der Pollenreife die DNS-Replikation der generativen bzw. Sperma-Kerne eingesetzt hat, aber je nach Pflanzenart noch nicht beendet sein muß. Zum gleichen Zeitpunkt der Pollenkornentwicklung kann der vegetative Kern in Abhängigkeit von der Pflanzenart auf dem C-Niveau verharren, eine teilweise oder bereits abgeschlossene DNS-Replikation erfahren haben oder schon teilweise oder ganz degeneriert sein, ohne zuvor eine DNS-Replikation vollzogen zu haben. Zweitens wird in diesem Abschnitt diskutiert, daß mit großer Wahrscheinlichkeit im Ablauf der DNS-Replikation zwischen zwei- und dreikernigen Pollenkorn-Typen keine Unterschiede bestehen. Drittens wird die Hypothese vertreten, daß nur auf einem sehr frühen Stadium die normale Pollenkornentwicklung einschließlich des Ablaufs der DNS-Replikation insbesondere des vegetativen Kerns so abgewandelt werden kann, daß aus Pollenkörnern haploide Pflanzen erzeugt werden können.

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The development of pollen grains and formation of pollen tubes in higher plantsIII. DNA-replication of vegetative and sperm nuclei in mature pollen grains of barley

Summary The DNA-content of vegetative and sperm nuclei in mature pollen grains of the barley varieties Amsel and Wisa and the F₁-plants of crossings of the barley varieties Impala X Wisa and Union X Wisa was determined by cytophotometry. In addition, the DNA-content of nuclei of root tips of Amsel and Wisa was cytophotometrically measured.The DNA contents of the nuclei in root tips of Amsel and Wisa differed significantly.The data obtained from the measurements of the vegetative and sperm nuclei of the four types of barley show that DNA-replication continues in the nuclei of mature pollen grains. The DNA values of vegetative nuclei of Wisa are significantly lower than the values of Amsel and of the F₁ plants. The DNA values of the different nuclei indicate that DNA replication of both types of sperm nuclei is synchronous, whereas it probably is not synchronous in vegetative and sperm nuclei respectively.In the discussion it is pointed out that a survey of the literature shows that in all of the plant species having binucleate or trinucleate pollen DNA replication of generative and of sperm nuclei has started at the time of pollen grain maturation. Depending on the plant species, replication may or may not be completed in the mature pollen grain. At a given stage of development of the pollen grain the vegetative nucleus may be arrested at the C-stage, may have partially or completely finished its DNA replication or may be partially or completely degenerated without prior replication of DNA.In the second part of the discussion it is stated that the course of DNA replication is likely to be similar in binucleate and trinucleate pollen grains. Thirdly, the hypothesis is discussed that in order to get haploid plants from pollen grains, changes in the normal development of the pollen grain and in the pattern of DNA replication must occur at a very early stage of pollen grain development.

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Mit Unterstützung durch die Deutsche Forschungsgemeinschaft.

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Angenommen durch F. Mechelke

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Mein Dank grit Herrn Prof. Dr. F. Mechelke fiir die Anregung zu diesen Untersuchungen sowie fiir die Unterstfitzung und die kritischen Diskussionen w/ihrend ihres Verlaufs und Fr/iulein H. Nagel fiir zuverl/issige technische Hilfe.     

Sequence analysis of a 101-kilobase plasmid required for agar degradation by a Microscilla isolate.

An agar-degrading marine bacterium identified as a Microscilla species was isolated from coastal California marine sediment. This organism harbored a single 101-kb circular DNA plasmid designated pSD15. The complete nucleotide sequence of pSD15 was obtained, and sequence analysis indicated a number of genes putatively encoding a variety of enzymes involved in polysaccharide utilization. The most striking feature was the occurrence of five putative agarase genes. Loss of the plasmid, which occurred at a surprisingly high frequency, was associated with loss of agarase activity, supporting the sequence analysis results.     

Identification of clusterin sequences mediating renal tubular cell interactions.

Expression of the glycoprotein clusterin is markedly increased following tissue injury. One function of clusterin is to promote cell interactions which are perturbed in these pathologic settings. Clusterin causes cell aggregation and adhesion in vitro yet the molecular mechanism for this effect is not known. In order to identify the active site(s) of clusterin, 34 peptides, each 15 amino acid residues in length, were synthesized from hydrophilic regions of human clusterin. When studied individually, none of the peptides caused aggregation of LLC-PK1 cells, a porcine renal epithelial cell line. However, two out of the 34 peptides inhibited clusterin-induced cell aggregation in a dose-dependent manner. Scrambled versions of these two 'active' peptides did not inhibit cell aggregation. Seven peptides promoted cell adhesion. In conclusion, these findings provide evidence for novel amino acid sequences mediating clusterin-induced renal cell interactions.     

Dapper Antagonist of Catenin-1 Cooperates with Dishevelled-1 during Postsynaptic Development in Mouse Forebrain GABAergic Interneurons

Synaptogenesis has been extensively studied along with dendritic spine development in glutamatergic pyramidal neurons, however synapse development in cortical interneurons, which are largely aspiny, is comparatively less well understood. Dact1, one of 3 paralogous Dact (Dapper/Frodo) family members in mammals, is a scaffold protein implicated in both the Wnt/β-catenin and the Wnt/Planar Cell Polarity pathways. We show here that Dact1 is expressed in immature cortical interneurons. Although Dact1 is first expressed in interneuron precursors during proliferative and migratory stages, constitutive Dact1 mutant mice have no major defects in numbers or migration of these neurons. However, cultured cortical interneurons derived from these mice have reduced numbers of excitatory synapses on their dendrites. We selectively eliminated Dact1 from mouse cortical interneurons using a conditional knock-out strategy with a Dlx-I12b enhancer-Cre allele, and thereby demonstrate a cell-autonomous role for Dact1 during postsynaptic development. Confirming this cell-autonomous role, we show that synapse numbers in Dact1 deficient cortical interneurons are rescued by virally-mediated re-expression of Dact1 specifically targeted to these cells. Synapse numbers in these neurons are also rescued by similarly targeted expression of the Dact1 binding partner Dishevelled-1, and partially rescued by expression of Disrupted in Schizophrenia-1, a synaptic protein genetically implicated in susceptibility to several major mental illnesses. In sum, our results support a novel cell-autonomous postsynaptic role for Dact1, in cooperation with Dishevelled-1 and possibly Disrupted in Schizophrenia-1, in the formation of synapses on cortical interneuron dendrites.     

Structure of Haze Forming Proteins in White Wines: Vitis vinifera Thaumatin-Like Proteins

Grape thaumatin-like proteins (TLPs) play roles in plant-pathogen interactions and can cause protein haze in white wine unless removed prior to bottling. Different isoforms of TLPs have different hazing potential and aggregation behavior. Here we present the elucidation of the molecular structures of three grape TLPs that display different hazing potential. The three TLPs have very similar structures despite belonging to two different classes (F2/4JRU is a thaumatin-like protein while I/4L5H and H2/4MBT are VVTL1), and having different unfolding temperatures (56 vs. 62°C), with protein F2/4JRU being heat unstable and forming haze, while I/4L5H does not. These differences in properties are attributable to the conformation of a single loop and the amino acid composition of its flanking regions.     

Synthesis of 18O-labeled RNA for application to kinetic studies and imaging

Radioisotopes and fluorescent compounds are frequently used for RNA labeling but are unsuitable for clinical studies of RNA drugs because of the risk from radiation exposure or the nonequivalence arising from covalently attached fluorophores. Here, we report a practical phosphoramidite solid-phase synthesis of ¹⁸O-labeled RNA that avoids these disadvantages, and we demonstrate its application to quantification and imaging. The synthesis involves the introduction of a nonbridging ¹⁸O atom into the phosphate group during the oxidation step of the synthetic cycle by using ¹⁸O water as the oxygen donor. The ¹⁸O label in the RNA was stable at pH 3–8.5, while the physicochemical and biological properties of labeled and unlabeled short interfering RNA were indistinguishable by circular dichroism, melting temperature and RNA-interference activity. The ¹⁸O/¹⁶O ratio as measured by isotope ratio mass spectrometry increased linearly with the concentration of ¹⁸O-labeled RNA, and this technique was used to determine the blood concentration of ¹⁸O-labeled RNA after administration to mice. ¹⁸O-labeled RNA transfected into human A549 cells was visualized by isotope microscopy. The RNA was observed in foci in the cytoplasm around the nucleus, presumably corresponding to endosomes. These methodologies may be useful for kinetic and cellular-localization studies of RNA in basic and pharmaceutical studies.     

Savignin, a potent thrombin inhibitor isolated from the salivary glands of the tick Ornithodoros savignyi (Acari: Argasidae).

A thrombin (E.C. 3.4.21.5) inhibitor, savignin, was isolated from the salivary glands of Ornithodoros savignyi by a combination of size exclusion, anion-exchange, and reversed-phase chromatography. The inhibitor has a molecular mass of 12,430.4 Da as determined by electrospray mass spectrometry. The behavior of savignin during anion-exchange chromatography indicated that it has an acidic pI. The available N-terminal sequence (residues 1-11) differed from that of ornithodorin with only one residue. Savignin inhibits thrombin-induced platelet aggregation, but has no effect on ADP- or collagen-induced aggregation. Kinetic studies indicated that savignin is a competitive, slow-, tight-binding inhibitor of alpha-thrombin (K(i) = 4.89 +/- 1.39 pM). Tight-binding kinetics showed that the inhibitor has a lower affinity for gamma-thrombin (K(i) = 22.3 +/- 5.9 nM). Plasmin, factor Xa, and trypsin are not inhibited by savignin.     

Lung tissue mechanics and extracellular matrix composition in a murine model of silicosis.

The dynamic mechanical properties of lung tissue and its contents of collagen and elastic fibers were studied in strips prepared from mice instilled intratracheally with saline (C) or silica [15 (S15) and 30 days (S30) after instillation]. Resistance, elastance, and hysteresivity were studied during oscillations at different frequencies on S15 and S30. Elastance increased from C to silica groups but was similar between S15 and S30. Resistance was augmented from C to S15 and S30 and was greater in S30 than in S15 at higher frequencies. Hysteresivity was higher in S30 than in C and S15. Silica groups presented a greater amount of collagen than did C. Elastic fiber content increased progressively along time. This increment was related to the higher amount of oxytalan fibers at 15 and 30 days, whereas elaunin and fully developed elastic fibers were augmented only at 30 days. Silicosis led not only to pulmonary fibrosis but also to fibroelastosis, thus assigning a major role to the elastic system in the silicotic lung.     

Distribution of matter in the current-carrying plasma and dense core of the discharge channel formed upon electrical wire explosion

Distribution of matter in the discharge channel formed upon a nanosecond electrical explosion of a single wire in air and vacuum was studied experimentally. Simultaneous use of optical, UV, and X-ray diagnostics made it possible to distinguish qualitatively different regions of the discharge channel, such as the current-carrying layers and the region occupied by a weakly conducting cold plasma. Several series of experiments with 25-μm-diameter 12-mm-long wires made of different materials were performed. The charging voltage and the current amplitude were varied in the ranges of U ₀ = 10–20 kV and I _max ∼ 5–10 kA, respectively. Explosion regimes with a current pause and with and without current interruption, as well as with wire preheating in air and vacuum, were studied. Shadow and schlieren images of the discharge channel were obtained using optical probing at the second harmonic of a YAG: Nd⁺³ laser (λ = 0.532 μm, τ ∼ 10 ns). In the experiments carried out in vacuum, X-ray images of the discharge channel were also obtained using an X-pinch as a point source of probing radiation and UV images were recorded using a four-frame MCP camera.