Nucleosides,XLIII1) Synthesis and Properties of 04-Alkylthymidines

Abstract

Various 0⁴-alkylthymidines 14–20 have been synthesized by two different methods. 0⁴-Alkylation takes place with 3′,5′-di-0-acetyl- (2) and 3′,5′-di-0-benzoylthymidine (3) respectively in a silver ion catalysed reaction with alkyl halides, whereas the azolide approach makes use of a nucleophilic displacement of the appropriate intermediate by alkoxides and subsequent deacylation to the free nucleosides. Structural proofs are based on elemental analyses, UV- and ¹H-NMR-spectra.     

Effect of Oxidative Stress on the Junctional Proteins of Cultured Cerebral Endothelial Cells

Summary 1. There is increasing evidence that the cerebral endothelium and the blood–brain barrier (BBB) plays an important role in the oxidative stress-induced brain damage. The aim of the present study was to investigate the role of interendothelial junctional proteins in the BBB permeability increase induced by oxidative stress.2. For the experiments, we have used cultured cerebral endothelial cells exposed to hypoxia/reoxygenation or treated with the redox cycling quinone 2,3-Dimethoxy-1,4-naphthoquinone (DMNQ) in the presence or absence of glucose. The expression of junctional proteins and activation of mitogen activated protein kinases (MAPK) was followed by Western-blotting, the interaction of junctional proteins was investigated using coimmunoprecipitation.3. Oxidative stress induces a downregulation of the tight junction protein occludin expression which is more pronounced in the absence of glucose. Furthermore, oxidative stress leads to disruption of the cadherin--catenin complex and an activation of extracellular signal-regulated kinase (ERK1/2), which is more intense in the absence of glucose.4. We have shown that one of the causes of the BBB breakdown is probably the structural alteration of the junctional complex caused by oxidative stress, a process in which ERK1/2 may play an important role.This revised article was published online in May 2005 with a February 2005 cover date.     

An indirect method for quantitation of cellular zinc content of timm-stained cerebellar samples by energy dispersive X-ray microanalysis

Summary The absolute concentration of zinc in the Purkinje cells of the rat cerebellum was determined by means of energy dispersive X-ray microanalysis (EDAX). Gelatine blocks with known zinc concentrations were stained by Timm's sulphide-silver method, and their silver concentrations were measured by EDAX. A linear correlation was found between the zinc and silver concentrations and this linear function was used as a quantitative calibration for evaluation of sulphide-silver staining, after perfusion with sodium-sulphide solution, fixation with glutaraldehyde, cryostat sectioning and staining of cerebellar samples in Timm's reagent.     

Effect of Basic and Acidic Additives on the Separation of Some Basic Drug Enantiomers on Polysaccharide‐Based Chiral Columns With Acetonitrile as Mobile Phase

The separation of enantiomers of 16 basic drugs was studied using polysaccharide‐based chiral selectors and acetonitrile as mobile phase with emphasis on the role of basic and acidic additives on the separation and elution order of enantiomers. Out of the studied chiral selectors, amylose phenylcarbamate‐based ones more often showed a chiral recognition ability compared to cellulose phenylcarbamate derivatives. An interesting effect was observed with formic acid as additive on enantiomer resolution and enantiomer elution order for some basic drugs. Thus, for instance, the enantioseparation of several β‐blockers (atenolol, sotalol, toliprolol) improved not only by the addition of a more conventional basic additive to the mobile phase, but also by the addition of an acidic additive. Moreover, an opposite elution order of enantiomers was observed depending on the nature of the additive (basic or acidic) in the mobile phase. Chirality 27:228–234, 2015. © 2015 Wiley Periodicals, Inc.     

Soil carbon dioxide efflux determined from large undisturbed soil cores collected in different soil management systems

The aim of this study was to evaluate a measuring technique for determining soil CO₂ efflux from large soil samples having undisturbed structure under controlled laboratory conditions. Further objectives were to use the developed measuring method for comparing soil CO₂ efflux from samples, collected in three different soil management systems at various soil water content values. The experimental technique was tested and optimised for timing of sampling by taking air samples after 1, 3 and 6 hours of incubation. Based on the results, the incubation time was set to three hours. The CO₂ efflux measured for different soil management systems was the highest in the no-till and the lowest in the ploughing treatment, which was in accordance with measurements on accessible organic carbon for microbes. An increase in CO₂ efflux with increasing soil water content was found in the studied soil water content range. Our results indicate that soil respiration rates, measured directly after tillage operations, can highly differ from those measured long after.     

Copper deficiency induced emphysema is associated with focal adhesion kinase inactivation

Background

Copper is an important regulator of hypoxia inducible factor 1 alpha (HIF-1α) dependent vascular endothelial growth factor (VEGF) expression, and is also required for the activity of lysyl oxidase (LOX) to effect matrix protein cross-linking. Cell detachment from the extracellular matrix can induce apoptosis (anoikis) via inactivation of focal adhesion kinase (FAK).

Methodology

To examine the molecular mechanisms whereby copper depletion causes the destruction of the normal alveolar architecture via anoikis, Male Sprague-Dawley rats were fed a copper deficient diet for 6 weeks while being treated with the copper chelator, tetrathiomolybdate. Other groups of rats were treated with the inhibitor of auto-phosphorylation of FAK, 1,2,4,5-benzenetetraamine tetrahydrochloride (1,2,4,5-BT) or FAK small interfering RNA (siRNA).

Principal Findings

Copper depletion caused emphysematous changes, decreased HIF-1α activity, and downregulated VEGF expression in the rat lungs. Cleaved caspase-3, caspase-8 and Bcl-2 interacting mediator of cell death (Bim) expression was increased, and the phosphorylation of FAK was decreased in copper depleted rat lungs. Administration of 1,2,4,5-BT and FAK siRNA caused emphysematous lung destruction associated with increased expression of cleaved capase-3, caspase-8 and Bim.

Conclusions

These data indicate that copper-dependent mechanisms contribute to the pathogenesis of emphysema, which may be associated with decreased HIF-1α and FAK activity in the lung.     

Interference by unstable nucleotides in the study of enzymes that cleave N-glycoside bonds in nucleic acids

The guanylation of tRNA involves the excision of a base from within the polynucleotide chain by cleavage of the N-glycoside bond. The excised base is replaced by guanine. During studies using chemically tritiated tRNA to study the guanylation reaction, spurious radioactivity was released from tRNA. We have identified the substance released as the hypermodified compound known as Y base. Our report (i) warns workers, who are studying enzymes that excise bases at the polynucleotide level, against this pitfall; (ii) indicates how to determine if counts released from RNA or DNA are spurious; and (iii) describes methods that avoid release of spurious counts while studying enzymes that cleave N-glycoside bonds in polynucleotides.