Oxidative stress increases levels of endogenous amyloid-beta peptides secreted from primary chick brain neurons

Oxidative damage is associated with Alzheimer's disease and mild cognitive impairment, but its relationship to the development of neuropathological lesions involving accumulation of amyloid-beta (Abeta) peptides and hyperphosphorylated tau protein remains poorly understood. We show that inducing oxidative stress in primary chick brain neurons by exposure to sublethal doses of H(2)O(2 )increases levels of total secreted endogenous Abeta by 2.4-fold after 20 h. This occurs in the absence of changes to intracellular amyloid precursor protein or tau protein levels, while heat-shock protein 90 is elevated 2.5-fold. These results are consistent with the hypothesis that aging-associated oxidative stress contributes to increasing Abeta generation and up-regulation of molecular chaperones in Alzheimer's disease.     

Aluminum-mediated metabolic changes in rat serum and urine: a proton nuclear magnetic resonance study

The toxic effects of Al(3+) have been studied in 90-days AlCl(3) orally treated male albino rats (n = 7) using (1)H NMR spectroscopy-based metabolic profile of rat serum and urine, serum enzyme tests, behavioral impairment, and histopathology of kidney and liver. Metabolic profile of 90-days Al(3+)-treated rat sera showed significantly elevated levels of alanine, glutamine, beta-hydroxy-butyrate, and acetoacetate and significantly decreased level of acetone when compared with that of control rats. However, metabolic profile of 90-days Al(3+)-treated rat urine showed significantly decreased levels of citrate, creatinine, allantoin, trans-aconitate, and succinate and significantly increased level of acetate when compared to control rats. The overall perturbations observed in the metabolic profile of serum and urine demonstrate the impairment in the tricarboxylic acid cycle, liver and kidney metabolism, which was further reinstated by clinical chemistry and histopathological observations. Moreover, "in vivo" behavioral impairment has also been observed as the indication of aluminum neurotoxicity.     

Role of the JAK-STAT pathway in protection of hydrogen peroxide preconditioning against apoptosis induced by oxidative stress in PC12 cells

The aim of this study was to investigate the role of JAK-STAT pathway in the cytoprotection afforded by preconditioning with H₂O₂. It was shown that (1) Preconditioning with 100 μmol/L H₂O₂ can markedly protect PC12 cells against apoptosis and cytotoxicity induced by 300 μmol/L H₂O₂; (2) The expression and tyrosine phosphorylation of JAK2, not JAK1 were rapidly increased at 5 min after H₂O₂ preconditioning; (3) The expression of STAT1 and STAT3 were significantly increased at 15 min after H₂O₂ preconditioning, and the pTyr-STAT1 and pTyr-STAT3 were markedly increased at 60 min after H₂O₂ preconditioning; (4) Pretreatment with the JAK inhibitor AG-490 (10 μmol/L) 20 min before H₂O₂ preconditioning blocked not only the activation of JAK2, STAT1 and STAT3, but also the cytoprotection of H₂O₂ preconditioning against apoptosis and cytotoxicity induced by oxidative stress. These findings suggested that preconditioning with H₂O₂ activated the JAK-STAT pathway that played an important role in the cytoprotection induced by H₂O₂ preconditioning.     

鸭瘟病毒gB蛋白N端主要抗原域的表达及间接ELISA检测

在分析鸭瘟病毒gB蛋白抗原性的基础上,设计一对引物克隆gB蛋白N端抗原性较好的抗原域编码基因.将克隆的基因定向插入pET-32a的EcoR Ⅰ和HindⅢ之间,构建了gB蛋白主要抗原域原核表达载体pET-gB1.将pET-gB1质粒转化BL(21)宿主菌后,对培养和表达条件进行了优化,实现了DPV gB蛋白主要抗原域的高效表达.免疫印迹试验表明获得的表达产物具有良好的反应原性.应用His·Bind亲和层析柱纯化重组DPV gB蛋白,以纯化的重组gB1蛋白作为检测抗原,初步建立了检测鸭瘟病毒抗体的igB1-ELISA.结果表明,抗原的最佳包被浓度为6.5μg/mL,血清的最佳稀释度为1∶80,阳性标准初步定为:待检血清OD490＞0.4,且待检血清OD490/阴性血清OD490＞2.应用igB1-ELISA对鸭血清样品进行检测,结果表明igB1-ELISA与全病毒包被的iDPV-ELISA符合率达到95.6%.     

Climate controls on C3 vs. C4 productivity in North American grasslands from carbon isotope composition of soil organic matter

We analyzed the δ¹³C of soil organic matter (SOM) and fine roots from 55 native grassland sites widely distributed across the US and Canadian Great Plains to examine the relative production of C₃ vs. C₄ plants (hereafter %C₄) at the continental scale. Our climate vs. %C₄ results agreed well with North American field studies on %C₄, but showed bias with respect to %C₄ from a US vegetation database (statsgo ) and weak agreement with a physiologically based prediction that depends on crossover temperature. Although monthly average temperatures have been used in many studies to predict %C₄, our analysis shows that high temperatures are better predictors of %C₄. In particular, we found that July climate (average of daily high temperature and month's total rainfall) predicted %C₄ better than other months, seasons or annual averages, suggesting that the outcome of competition between C₃ and C₄ plants in North American grasslands was particularly sensitive to climate during this narrow window of time. Root δ¹³C increased about 1‰ between the A and B horizon, suggesting that C₄ roots become relatively more common than C₃ roots with depth. These differences in depth distribution likely contribute to the isotopic enrichment with depth in SOM where both C₃ and C₄ grasses are present.     

Three-dimensional structure of the ciliate Didinium nasutum nucleoli

The nucleolar organization in ciliate Didinium nasutum somatic interphase nuclei was studied using serial ultrathin sections and compared for various physiological states of the cell, namely, fed ciliates, starved ciliates, and dormant cysts. It has been shown that the interphase nucleoli are large structures with a complex architecture: the fibrillar component forms an intricate network in the macronucleus space, while the granular component is located inside this network. The structures looking as individual nucleoli in single sections are actually parts of branched nucleolar networks. The intricate nucleolar networks do not disintegrate after a 30-h starvation; however, the granular component becomes denser and develops numerous cavities filled with fine fibrils of a nonribonucleoprotein nature. In fed D. nasutum, the fibrillar structures on the periphery of nucleoli contain numerous pores (virtually absent in starved cell nucleoli), which can potentially serve for transporting newly synthesized rRNP. Branched nucleolar networks are undetectable in cysts. Their nucleoli are individual structures consisting mainly of the fibrogranular component.     

Combined expression of chitinase and β-1,3-glucanase genes in indica rice (Oryza sativa L.) enhances resistance against Rhizoctonia solani

Agrobacterium-mediated transformation of rice was done using the binary vector pNSP3, harbouring the rice chitinase (chi11) gene under maize ubiquitin promoter and the tobacco β-1,3-glucanase gene under CaMV 35S promoter in the same T-DNA. Four of the six T₀ plants had single copies of complete T-DNAs, while the other two had complex integration patterns. Three of the four single-copy lines showed a 3:1 segregation ratio in the T₁ generation. Northern and western blot analyses of T₁ plants revealed constitutive expression of chitinase and β-1,3-glucanase genes. Homozygous T₂ plants of the single-copy lines CG20, CG27 and CG53 showed 62-, 9.6- and 11-fold higher chitinase activity over the control plants. β-1,3-Glucanase activity was 1.1- to 2.5-fold higher in the transgenic plants. Bioassay of homozygous T₂ plants of the three single-copy transgenic lines against Rhizoctonia solani revealed a 60% reduction in sheath blight Disease Index in the first week. The Disease Index increased from 61.8 in the first week to 90.6 in the third week in control plants, while it remained low (26.8–34.2) in the transgenic T₃ plants in the corresponding period, reflecting the persistence of sheath blight resistance for a longer period.     

Anaerobic degradation of trans-cinnamate and ω-phenylalkane carboxylic acids by the photosynthetic bacterium Rhodopseudomonas palustris: evidence for a β-oxidation mechanism

The mechanism responsible for the initial steps in the anaerobic degradation of trans-cinnamate and -phenylalkane carboxylates by the purple non-sulphur photosynthetic bacterium Rhodopseudomonas palustris was investigated. Phenylacetate did not support growth and there was a marked CO₂ dependence for growth on acids with greater side-chain lengths. Here, CO₂ was presumably acting as a redox sink for the disposal of excess reducing equivalents. Growth on benzoate did not require the addition of exogenous CO₂. Aromatic acids with an odd number of side-chain carbon atoms (3-phenylpropionate, 5-phenylvalerate, 7-phenylheptanoate) gave greater apparent molar growth yields than those with an even number of side-chain carbon atoms (4-phenylbutyrate, 6-phenylhexanoate, 8-phenyloctanoate). HPLC analysis revealed that phenylacetate accumulated and persisted in the culture medium during growth on these latter compounds. Cinnamate and benzoate transiently accumulated in the culture medium during growth on 3-phenylpropionate, and benzoate alone accumulated transiently during the course of trans-cinnamate degradation. The transient accumulation of 4-phenyl-2-butenoic acid occurred during growth on 4-phenylbutyrate, and phenylacetate accumulated to a 1:1 molar stoichiometry with the initial 4-phenylbutyrate concentration. It is proposed that the initial steps in the anaerobic degradation of trans-cinnamate and the group of acids from 3-phenylpropionate to 8-phenyloctanoate involves -oxidation of the side-chain.Abbreviation 3-PP 3-phenylpropionic acid - 4-PB 4-phenylbutyric acid - 5-PV 5-phenylvaleric acid - 6-PH 6-phenylhexanoic acid - 7-PH 7-phenylheptanoic acid - 8-PO 8-phenyloctanoic acid - 4-P2B 4-phenyl-2-butenoic acid - GC/MS Gas chromatography/Mass spectrometry - HPLC High-pressure liquid chromatography     

The ATP-dependent RNA helicase,DDX42 interacts with paxillin and regulates apoptosis and polarization of Ba/F3 cells

Paxillin is a focal adhesion adaptor protein, heavily phosphorylated at multiple tyrosine residues, as well as at serine 273 (S273), and is known to be critical for cytoskeleton rearrangement and cell migration. We previously found that paxillin plays a regulatory role in IL-3-dependent survival of Ba/F3 cells, a mouse pro-B cell line. In this study, by using overexpressed His6 tagged-paxillin as a bait, we found that DDX42, a DEAD-box RNA helicase, interacted with paxillin, inhibited apoptosis, and promoted polarization of Ba/F3 cells. His6 tagged-paxillin was stably overexpressed in Ba/F3 cells, pulled-down from cell lysates with Ni⁺-NTA beads, and analyzed by one-dimensional SDS-PAGE followed by LC–MS. We found that DDX42 co-precipitated with paxillin, as demonstrated by western blotting analysis of His6 tagged-paxillin precipitates with anti-DDX42 antibodies and His6 tagged-DDX42 precipitates with anti-paxillin antibodies. In addition, we observed a preferential interaction of DDX42 with the paxillin mutant, S273A, compared to the S273D mutant. Furthermore, DDX42 overexpression in Ba/F3 cells delayed the apoptosis induced by IL-3 deprivation and promoted restoration of the elongated shape in Ba/F3 cells induced by IL-3 re-supply after a 6?h-deprivation. These results suggested that DDX42 interacts with paxillin and participates in IL-3-dependent cell survival, as well as in the cytoskeletal rearrangements underlying polarization of Ba/F3 cells.     

Radiation dose verification of an X-ray based blood irradiator using EBT3 radiochromic films calibrated using Gamma Knife machine

AimBlood irradiators (BI) initial acceptance testing and routine annual dosimetry checks require radiation dose measurements in order to comply with regulatory requirements.BackgroundTraditionally thermo-luminescence dosimeters (TLD) have been used to measure the dose. The EBT3 film is reported to be a better dosimeter for low energy X-rays than its predecessors EBT2 and EBT. To the best of our knowledge, the use of EBT3 films to perform dosimetry on X-ray based BI has not been reported yet.Materials and methodsWe performed routine radiation dosimetry checks using EBT3 films on a new X-ray based BI and compared the results with TLD dosimetry. Calibration films were irradiated with radiation beam from a Co-60 Gamma Knife (GK) radiosurgery machine and, alternatively, using an Ir-192 high dose rate (HDR) brachytherapy device. The films were calibrated to cover a wide dose range from 1 to 40 Gy. Such a wide dose range has not been reported yet in BI film dosimetry.ResultsWe obtained a relative difference of about 6.6% between doses measured using TLD and those measured using EBT3 films. Both irradiation methods using GK or HDR were found to be adequate for the calibration of the EBT3 Gafchromic films.ConclusionsWe recommend the use of EBT3 films in routine X-ray based BI dosimetry checks. The presented method takes advantage of available radiotherapy equipment that can be efficiently used for EBT3 films calibration. The method is fast, reproducible and saves valuable medical physicist's time.