Melatonin attenuates the effects of sub-acute administration of lead on kidneys in rats without altering the lead-induced reduction in nitric oxide

Exposure to lead induces oxidative stress and renal damage. Although most forms of oxidative stress are characterized by simultaneous elevation of nitrogen and oxidative species, lead-induced oxidative stress is unusual in that it is associated with a reduction in nitric oxide (NO) levels in the kidney. The role of NO in kidney injury is controversial; some studies suggest that it is associated with renal injury, whereas others show that it exerts protective effects. Concentration-dependent effects have also been proposed, linking low levels with vasodilatation and high levels with toxicity. The aim of this study was to evaluate the effects of melatonin co-exposure on the lead-induced reduction in renal NO levels. We found that sub-acute intraperitoneal administration of 10 mg/kg/day of lead for 15 days induced toxic levels of lead in the blood and caused renal toxicity (pathological and functional). Under our experimental conditions, lead induced an increase in lipid peroxidation and a decrease in NO. Melatonin co-treatment decreased lead-induced oxidative stress (peroxidation level) and toxic effects on kidneys without altering the lead-induced reduction in renal NO. These results suggest that, in our experimental model, the reduction in renal NO levels by lead exposure is not the only responsible factor for lead-induced kidney damage.     

基于杂交链式(HCR)反应的甲型流感病毒检测技术研究

甲型流感病毒的现场快速检测对于流感的及时有效防控具有重要意义．本研究基于杂交链式(HCR)反应,利用GO对荧光基团的猝灭作用及共同实现了对甲型流感病毒的快速检测．当目标序列存在时,可引发HCR反应,使短链DNA形成长链,保护FAM荧光基团不被猝灭,从而实现目标物的检测．实验结果表明,该方法在10～40 nmol/L范围内荧光强度与目标检测物浓度表现出了良好的线性关系,检测范围为5～100 nmol/L．这种HCR等温扩增检测技术具有较好的样本检测能力,具有等温、无酶、反应体系简单、操作步骤简便等优点,表现出良好的现场检测应用前景．     

Denitrosylation of S-nitrosylated OGT is triggered in LPS-stimulated innate immune response

O-linked N-acetylglucosaminyltransferase (OGT)-mediated protein O-GlcNAcylation has been revealing various aspects of functional significance in biological processes, such as cellular signaling and activation of immune system. We found that OGT is maintained as S-nitrosylated form in resting cells, and its denitrosylation is triggered in innate immune response of lipopolysaccharide (LPS)-treated macrophage cells. S-nitrosylation of OGT strongly inhibits its catalytic activity up to more than 80% of native OGT, and denitrosylation of OGT leads to protein hyper-O-GlcNAcylation. Furthermore, blockage of increased protein O-GlcNAcylation results in significant loss of nitric oxide and cytokine production. We propose that denitrosylation of S-nitrosylated OGT is a direct mechanism for upregulation of OGT activity by which immune defense is critically controlled in LPS-stimulated innate immune response.     

Spectrofluorimetric analysis of the interaction of amyloid peptides with neuronal nitric oxide synthase: Implications in Alzheimer's disease

Background

The deposition of aggregated β-amyloid peptide senile plaques and the accumulation of arginine within the astrocytes in the brain of an Alzheimer's patient are classic observations in the neuropathology of the disease. It would be logical, in the aetiology and pathogenesis, to investigate arginine-metabolising enzymes and their intimate association with amyloid peptides.

Methods

Neuronal nitric oxide synthase (nNOS) was isolated, purified and shown, through fluorescence quenching spectroscopy and fluorescence resonance energy transfer (FRET), to interact with structural fragments of Aβ_1–40 and be catalytic towards amyloid fibril formation.

Results

Only one binding site on the enzyme was available for binding. Two amyloid peptide fragments of Aβ_1–40 (Aβ_17–28 and Aβ_25–35) had Stern–Volmer values (K_SV) of 0.111 μM⁻¹ and 0.135 μM⁻¹ indicating tight binding affinity to nNOS and easier accessibility to fluor molecules during binding. The polarity of this active site precludes binding of the predominantly hydrophobic amyloid peptide fragments contained within Aβ_17–28 and within two glycine zipper motifs [G-X-X-X-G-X-X-X-G] [Aβ_29–37] and bind to the enzyme at a site remote to the active region.

Conclusions

The interaction and binding of Aβ_17–28 and Aβ_25–35 to nNOS causes the movement of two critical tryptophan residues of 0.77 nm and 0.57 nm respectively towards the surface of the enzyme.

General significance

The binding of Aβ-peptide fragments with nNOS has been studied by spectrofluorimetry. The information and data presented should contribute towards understanding the mechanism for deposition of aggregated Aβ-peptides and fibrillogenesis in senile plaques in an AD brain.     

Changes in nitrogen cycling and retention processes in soils under spruce forests along a nitrogen enrichment gradient in Germany

A network of long-term monitoring sites on nitrogen (N) input and output of forests across Germany showed that a number of Germany's forests are subject to or are experiencing N saturation and that spruce (Picea abies) stands have high risk. Our study was aimed at (1) quantifying the changes in gross rates of microbial N cycling and retention processes in forest soils along an N enrichment gradient and (2) relating the changes in soil N dynamics to N losses. We selected spruce sites representing an N enrichment gradient (indicated by leaching : throughfall N ratios) ranging from 0.04–0.13 (low N),≤0.26 (intermediate N enrichment) to≥0.42 (highly N enriched). To our knowledge, our study is the first to report on mechanistic changes in gross rates of soil N cycling and abiotic NO₃⁻ retention under ambient N enrichment gradient. Gross N mineralization, NH₄⁺ immobilization, gross nitrification, and NO₃⁻ immobilization rates increased up to intermediate N enrichment level and somewhat decreased at highly N-enriched condition. The turnover rates of NH₄⁺ and microbial N pools increased while the turnover rates of the NO₃⁻ pool decreased across the N enrichment gradient. Abiotic immobilization of NH₄⁺ did not differ across sites and was lower than that of NO₃⁻. Abiotic NO₃⁻ immobilization decreased across the N enrichment gradient. Microbial assimilation and turnover appeared to contribute largely to the retention of NH₄⁺. The increasing NO₃⁻ deposition and decreasing turnover rates of the NO₃⁻ pool, combined with decreasing abiotic NO₃⁻ retention, possibly contributed to increasing NO₃⁻ leaching and gaseous emissions across the N enrichment gradient. The empirical relationships of changes in microbial N cycling across the N enrichment gradient may be integrated in models used to predict responses of forest ecosystems (e.g. spruce) to increasing N deposition.     

The effect of copper and nitrogen on the amino acid composition of oat straw

Summary The effect of fertilization with nitrogen and copper on the amino acid composition of oat straw has been studied.The plants (Avena sativa cv Yielder) were grown in peat with a very low copper content and supplied with two levels of nitrogen (NH₄ or NO₃) and three levels of copper sulphate.The higher level of nitrogen stimulated growth only when copper was added, whereas, without copper, it had an adverse effect on growth and prevented grain formation altogether. The higher level of nitrogen increased the nitrogen content of the straw at all levels of copper, but particularly in plants receiving no copper.Total amino acids in the straw hydrolysate of copper sufficient oats accounted for about 50% of the total N and was about 20% higher in copper-deficient tissues. The addition of copper caused a decrease in the amounts of all amino acids. The relative proportions of most of the amino acids to glycine remained fairly constant. Threonine, serine, alanine, iso-leucine, histidine and arginine showed small significant differences with copper treatment, whereas valine, tyrosine, phenylalanine, proline, lysine and cysteic acid (derived from cysteine and cystine) showed no differences. The proportion of aspartic acid relative to glycine in the straw hydrolysate was greatly increased in copper deficient plants supplied with the higher level of nitrogen, particularly as ammonium. The proportion of glutamic acid was also increased by the higher level of nitrogen, but showed no effect of added copper. Most of the difference in aspartic acid could be accounted for as free asparagine. The possible reasons for higher proportions of asparagine are discussed in relation to the metabolism of the oat plant.     

Identification and quantification of free radicals during myocardial ischemia and reperfusion using electron paramagnetic resonance spectroscopy

There is general agreement that free radicals are involved in reperfusion injury. Electron paramagnetic resonance (EPR) spectroscopy can be considered as the more suitable technique to directly measure and characterize free radical generation during myocardial ischemia and reperfusion. There are essentially two approaches used in the detection of unstable reactive species: freezing technique and spin traps. The detection of secondary free radicals or ascorbyl free radicals during reperfusion might provide an index of oxidative stress. Spin trapping can also characterize nitric oxide. EPR spectroscopy can provide important data regarding redox state and free radical metabolism but ideally, the spin traps must not interfere with cell or organism function.     

Seasonal periodicity of enteric nitric oxide synthesis and its regulation in the snail, Helix lucorum

Abstract. The snail Helix lucorum has been used as a model to study the adaptation of a nitric oxide (NO)‐forming enteric neural network to the long‐term resting period of summer estivation or winter hibernation. Quantification of the NO‐derived nitrite established that NO formation is confined to the nitric oxide synthase (NOS)‐containing myenteric network of the mid‐intestine. In active snails but not in resting snails, NO production could be enhanced by the NOS substrate l ‐arginine (l ‐ARG, 1 mM). We followed the enteric NO synthesis in a snail population kept at natural conditions for 1 year. Our findings indicate that NO synthesis was depressed in July during entry to the estivation, had a peak in autumn before hibernation, and finally was reduced during hibernation. Monoamines (histamine, serotonin, and adrenalin) could inhibit the NO liberation in active snails. Cofactors of NOS (β‐NADPH, β‐NAD, FAD, FMN, Ca²⁺, TH4) did not alter the low nitrite production in hibernating snails. We conclude that enteric NO synthesis in H. lucorum has a regular seasonal periodicity following the annual physiological cycles of terrestrial snails. During estivation or hibernation, NOS activity is blocked. Monoamines, the levels of which are elevated during hibernation, can trigger decreased NOS activity. The reduced activity of NOS cannot be restored by the administration of NOS cofactors; therefore, their absence cannot be the cause of the temporarily blocked L‐ARG/NO conversion ability of NOS.     

Formation of unidentified nitrogen in plants: an implication for a novel nitrogen metabolism

Plants take up inorganic nitrogen and store it unchanged or convert it to organic forms. The nitrogen in such organic compounds is stoichiometrically recoverable by the Kjeldahl method. The sum of inorganic nitrogen and Kjeldahl nitrogen has long been known to equal the total nitrogen in plants. However, in our attempt to study the mechanism of nitrogen dioxide (NO₂) metabolism, we unexpectedly discovered that about one-third of the total nitrogen derived from ¹⁵N-labeled NO₂ taken up by Arabidopsis thaliana (L.) Heynh. plants was converted to neither inorganic nor Kjeldahl nitrogen, but instead to an as yet unknown nitrogen compound(s). We here refer to this nitrogen as unidentified nitrogen (UN). The generality of the formation of UN across species, nitrogen sources and cultivation environments for plants has been shown as follows. Firstly, all of the other 11 plant species studied were found to form the UN in response to fumigation with ¹⁵NO₂. Secondly, tobacco (Nicotiana tabacum L.) plants fed with ¹⁵N-nitrate appeared to form the UN. And lastly, the leaves of naturally fed vegetables, grass and roadside trees were found to possess the UN. In addition, the UN appeared to comprise a substantial proportion of total nitrogen in these plant species. Collectively, all of our present findings imply that there is a novel nitrogen mechanism for the formation of UN in plants. Based on the analyses of the exhaust gas and residue fractions of the Kjeldahl digestion of a plant sample containing the UN, probable candidates for compounds that bear the UN were deduced to be those containing the heat-labile nitrogen–oxygen functions and those recalcitrant to Kjeldahl digestion, including organic nitro and nitroso compounds. We propose UN-bearing compounds may provide a chemical basis for the mechanism of the reactive nitrogen species (RNS), and thus that cross-talk may occur between UN and RNS metabolisms in plants. A mechanism for the formation of UN-bearing compounds, in which RNS are involved as intermediates, is proposed. The important broad impact of this novel nitrogen metabolism, not only on the general physiology of plants, but also on plant substances as human and animal food, and on plants as an integral part of the global environment, is discussed.Abbreviations NO Nitric oxide - NO₂ Nitrogen dioxide - RNS Reactive nitrogen species - UN Unidentified nitrogen - TNNAT, RNNAT, INNAT and UNNAT Total, Kjeldahl, inorganic and unidentified nitrogen in naturally fed plants, respectively - TNNIT, RNNIT, INNIT and UNNIT Total, Kjeldahl, inorganic and unidentified nitrogen derived from nitrate, respectively - TNNO2, RNNO2, INNO2 and UNNO2 Total, Kjeldahl, inorganic and unidentified nitrogen derived from NO₂, respectively