Microcystin-RR induced apoptosis in tobacco BY-2 suspension cells is mediated by reactive oxygen species and mitochondrial permeability transition pore status

When tobacco BY-2 cells were treated with 60 mu g/mL MC-RR for 5 d, time-dependent effects of MC-RR on the cells were observed. Morphological changes such as abnormal elongation, evident chromatin condensation and margination, fragmentation of nucleus and formation of apoptotic-like bodies suggest that 60 mu g/mL MC-RR induced rapid apoptosis in tobacco BY-2 cells. Moreover, there was a significant and rapid increase of ROS level before the loss of mitochondrial membrane potential (Delta Psi(m)) and the onset of cell apoptosis. Ascorbic acid (AsA), a major primary antioxidant, prevented the increase of ROS generation, blocked the decrease in Delta Psi(m) and subsequent cell apoptosis, indicating a critical role of ROS in serving as an important signaling molecule by causing a reduction of Delta Psi(m) and MC-RR-induced tobacco BY-2 cell apoptosis. In addition, a specific mitochondrial permeability transition pores (PTP) inhibitor, cyclosporin A (CsA), significantly blocked the MC-RR-induced ROS formation, loss of Delta Psi(m), as well as cell apoptosis when the cells were MC-RR stressed for 3 d, suggesting that PTP is involved in 60 mu g/mL MC-RR-induced tobacco cell apoptosis signalling process. Thus, we concluded that the mechanism of MC-RR-induced apoptosis signalling pathways in tobacco BY-2 cells involves not only the excess generation of ROS and oxidative stress, but also the opening of PTP inducing loss of mitochondrial membrane potential. (C) 2007 Published by Elsevier Ltd.

TRAIL in the mandarin fish Siniperca chuatsi: Gene and its apoptotic effect in HeLa cells

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is one of the TNF superfamily members, participating in many biological processes including cell proliferation and apoptotic death. In this study, a TRAIL gene was cloned from a perciform fish, the mandarin fish Siniperca chuatsi, a major cultured fish in China's aquaculture, and is named as SCTRAIL for S. chuatsi TRAIL. The full-length cDNA of SCTRAIL is 1359 bp, encoding a 283-amino-acid protein. This deduced protein contains the CYS231, a 23-mer fragment of transmembrane region, a glycosylation site and a TNF family signature, all of which are conserved among TRAIL members. SCTRAIL gene consists of six exons, with five intervening introns, spaced over approximately 9 kb of genomic sequence. Southern blotting demonstrated that the SCTRAIL gene is present as a single copy in mandarin fish genome. A 620 bp promoter region obtained by genome walking contains a number of putative transcription factor binding sites, such as Oct-1, Sp-1, NF-1, RAP-1, C/EBPaLp, NF-kappa B and AP-1. The SCTRAIL is constitutively expressed in all the analyzed tissues, as revealed by RT-PCR, which is confirmed by Western blotting analysis using polyclonal antibody against bacteria-derived recombinant SCTRAIL protein. As an apoptosis-inducing ligand, the overexpression of SCTRAIL but not the mutant SCTRAIL-C203S in HeLa cells induced changes characteristic of apoptosis, including chromatin condensation, nucleus fragmentation, DNA ladder, and increase of sub-G0/G1 cells in FACS analysis. (c) 2007 Elsevier Ltd. All rights reserved.

Investigation of flame stretch in turbulent lifted jet flame

DNS data of a laboratory-scale turbulent lifted hydrogen jet flame has been analyzed to show that this flame has mixed mode combustion not only at the flame base but also in downstream locations. The mixed mode combustion is observed in instantaneous structures as in earlier studies and in averaged structure, in which the predominant mode is found to be premixed combustion with varying equivalence ratio. The non-premixed combustion in the averaged structure is observed only in a narrow region at the edge of the jet shear layer. The analyzes of flame stretch show large probability for negative flame stretch leading to negative surface averaged flame stretch. The displacement speed-curvature correlation is observed to be negative contributing to the negative flame stretch and partial premixing resulting from jet entrainment acts to reduce the negative correlation. The contribution of turbulent straining to the flame stretch is observed to be negative when the scalar gradient aligns with the most extensive principal strain rate. The physics behind the negative flame stretch resulting from turbulent straining is discussed and elucidated through a simple analysis of the flame surface density transport equation. © 2014 Copyright Taylor and Francis Group, LLC.

Scalar gradient behaviour in MILD combustion

The results of three-dimensional Direct Numerical Simulation (DNS) of Moderate, Intense Low-oxygen Dilution (MILD) and conventional premixed turbulent combustion conducted using a skeletal mechanism including the effects of non-unity Lewis numbers and temperature dependent transport properties are analysed to investigate combustion characteristics using scalar gradient information. The DNS data is also used to synthesise laser induced fluorescence (LIF) signals of OH, CH2O, and CHO. These signals are analysed to verify if they can be used to study turbulent MILD combustion and it has been observed that at least two (OH and CH2O) LIF signals are required since the OH increase across the reaction zone is smaller in MILD combustion compared to premixed combustion. The scalar gradient PDFs conditioned on the reaction rate obtained from the DNS data and synthesised LIF signals suggests a strong gradient in the direction normal to the MILD reaction zone with moderate reaction rate implying flamelet combustion. However, the PDF of the normal gradient is as broad as for the tangential gradient when the reaction rate is high. This suggests a non-flamelet behaviour, which is due to interaction of reaction zones. The analysis of the conditional PDFs for the premixed case confirms the expected behaviour of scalar gradient in flamelet combustion. It has been shown that the LIF signals synthesised using 2D slices of DNS data also provide very similar insights. These results demonstrate that the so-called flameless combustion is not an idealised homogeneous reactive mixture but has common features of conventional combustion while containing distinctive characteristics. © 2013 The Combustion Institute.

Mitochondrion-mediated apoptosis induced by Rana grylio virus infection in fish cells

A fish cell line, fathead minnow (FHM) cell, was used to investigate the alteration of mitochondrial dynamics and the mechanism of apoptosis under Rana grylio virus (RGV) infection. Microscopy observations, flow-cytometry analysis and molecular marker detection revealed the apoptotic fate of the RGV-infected cells. Some typical apoptotic characteristics, such as chromatin condensation, DNA fragmentation and mitochondrial fragmentation, were observed, and significantly morphological changes of mitochondria, including size, shape, internal structure and distribution, were revealed. The mitochondria in RGV-infected cells were aggregated around the viromatrix, and the aggregation could be blocked by colchicine. Moreover, the Delta psi m collapse was induced, and caspase-9 and caspase-3 were activated in the RGV-infected cells. In addition, NF-kappa B activation and intracellular Ca2+ increase were also detected at different times after infection. The data revealed the detailed dynamics of mitochondrion-mediated apoptosis induced by an iridovirus, and provided the first report on mitochondrial fragmentation during virus-induced apoptosis in fish cells.

Auxetic nuclei in embryonic stem cells exiting pluripotency.

Embryonic stem cells (ESCs) self-renew in a state of naïve pluripotency in which they are competent to generate all somatic cells. It has been hypothesized that, before irreversibly committing, ESCs pass through at least one metastable transition state. This transition would represent a gateway for differentiation and reprogramming of somatic cells. Here, we show that during the transition, the nuclei of ESCs are auxetic: they exhibit a cross-sectional expansion when stretched and a cross-sectional contraction when compressed, and their stiffness increases under compression. We also show that the auxetic phenotype of transition ESC nuclei is driven at least in part by global chromatin decondensation. Through the regulation of molecular turnover in the differentiating nucleus by external forces, auxeticity could be a key element in mechanotransduction. Our findings highlight the importance of nuclear structure in the regulation of differentiation and reprogramming.

Microcystin-RR-induced apoptosis in tobacco BY-2 cells

Our previous studies showed that microcystin-RR could induce oxidative damage in plant cells as they do with animal cells. However, whether microcystin can induce plant cell apoptosis is still unknown. In this study, the morphological changes of tobacco BY-2 suspension cells exposed to microcystin-RR were observed under light microscopy and transmission electron microscopy, and apoptosis was clearly distinguished by intense perinuclear chromatin margination, condensation of nuclear chromatin after 6d exposure of 50 mg/L (about 50 mu M) microcystin-RR. We also found that microcystin-RR can induce tobacco cell apoptosis in a dose- and time-dependent manner with flow cytometry analysis. Our study provides the first evidence that microcystins can induce plant cell apoptosis. (c) 2006 Elsevier Ltd. All rights reserved.

Modelling of turbulent lifted jet flames using flamelets: A priori assessment and a posteriori validation

This study focuses on the modelling of turbulent lifted jet flames using flamelets and a presumed Probability Density Function (PDF) approach with interest in both flame lift-off height and flame brush structure. First, flamelet models used to capture contributions from premixed and non-premixed modes of the partially premixed combustion in the lifted jet flame are assessed using a Direct Numerical Simulation (DNS) data for a turbulent lifted hydrogen jet flame. The joint PDFs of mixture fraction Z and progress variable c, including their statistical correlation, are obtained using a copula method, which is also validated using the DNS data. The statistically independent PDFs are found to be generally inadequate to represent the joint PDFs from the DNS data. The effects of Z-c correlation and the contribution from the non-premixed combustion mode on the flame lift-off height are studied systematically by including one effect at a time in the simulations used for a posteriori validation. A simple model including the effects of chemical kinetics and scalar dissipation rate is suggested and used for non-premixed combustion contributions. The results clearly show that both Z-c correlation and non-premixed combustion effects are required in the premixed flamelets approach to get good agreement with the measured flame lift-off heights as a function of jet velocity. The flame brush structure reported in earlier experimental studies is also captured reasonably well for various axial positions. It seems that flame stabilisation is influenced by both premixed and non-premixed combustion modes, and their mutual influences. © 2014 Taylor & Francis.

Induction of apoptosis in a carp leucocyte cell line infected with turbot (Scophthalmus maximus L.) rhabdovirus

A rhabdovirus was observed from the diseased turbot (Scophthalmus maximus L.) with lethal syndrome. In this study, a carp leucocyte (CLC) cell line was used to investigate the infection process and cell death mechanism occurring during the virus infection. Strong cytopathogenic effect (CPE) and the morphological changes, such as extreme chromatin condensation, nucleus fragmentation, and apoptotic body formation, were observed under fluorescence microscopy after DAPI staining in the infected CLC cells. Transmission electron microscopy analysis showed cell shrinkage, plasma membrane blebbing, cytoplasm vacuolization, chromatin condensation, nuclear breakdown and formation of discrete apoptotic bodies. The bullet-shaped nucleocapsids were measured and ranged in size from 110 to 150 nm in length and 40 to 60 nm in diameter. And therefore the virus is called Scophthalmus maximus rhabdovirus (SMRV). Agarose gel electrophoresis analysis of the DNA extracted from infected cells showed typical DNA ladder in the course of SMRV infection. Flow cytometry analysis of SMRV infected CLC cells detected apoptotic peak in the virus infected CLC cells. Virus titre analysis and electron microscopic observation revealed that the virus replication fastigium was earlier than that of the apoptosis occurrence. No apoptosis was observed in the CLC infected with UV-inactivated SMRV. All these supported that SMRV infected CLC cells undergo apoptosis and the virus replication is necessary for apoptosis induction of CLC cells. (C) 2004 Published by Elsevier B.V.

Infection and propagation of lymphocystis virus isolated from the cultured flounder Paralichthys olivaceus in grass carp cell lines

The causative agent of lymphocystis disease that frequently occurs in cultured flounder Paralichthys olivaceus in China is lymphocystis virus (LV). In this study, 13 fish cell lines were tested for their susceptibility to LV. Of these, 2 cell lines derived from the freshwater grass carp Ctenopharyngodon idellus proved susceptible to the LV, and 1 cell line, GCO (grass carp ovary), was therefore used to replicate and propagate the virus. An obvious cytopathic effect (CPE) was first observed in cell monolayers at 1 d post-inoculation, and at 3 d this had extended to about 75% of the cell monolayer. However, no further CPE extension was observed after 4 d. Cytopathic characteristics induced by the LV were detected by Giemsa staining and fluorescence microscopic observation with Hoechst 33258 staining. The propagated virus particles were also observed by electron microscopy. Ultrastructure analysis revealed several distinct cellular changes, such as chromatin compaction and margination, vesicle formation, cell-surface convolution, nuclear fragmentation and the occurrence of characteristic 'blebs' and cell fusion. This study provides a detailed report of LV infection and propagation in a freshwater fish cell line, and presents direct electron microscopy evidence for propagation of the virus in infected cells. A possible process by which the CPEs are controlled is suggested.

Effects of cell-cycle arrest agents on cleavage and development of loach (Misgurnus anguillicaudatus) embryos

The aim of this study was to determine the lowest concentration of nocodazole and colchicine to arrest blastomere division during the cleavage stage of loach embryos and to assess the reversibility and toxicity of the treatments in the treated embryos. Eight-cell loach embryos were incubated for 4, 8, 12, or 16 h in 1/10x Holtfreter supplemented with either nocodazole, an inhibitor of tubulin polymerization, or colchicine, an inhibitor of tubulin assembly. Complete arrest of cell cycle was observed, at a colchicine concentration of 0.996 mM and at a nocodazole concentration of 0.275 muM, respectively (the lowest effective concentration). No major morphological alteration in chromatin was observed. Reversibility and toxicity of both agents were dose and exposure period dependent. For both agents, prolonging cleavage arrest for more than 4 h (at the effective concentrations) is detrimental to development of embryos. Nocodazole treatment was less cytotoxic, whereas the concentrations of colchicine which induce cleavage arrest were detrimental to development beyond the blastula stage. Toxic effects beyond the blastula stage could be minimized for both agents by reducing the period of treatment and concentration.

Nuclear transfer in leach (Paramisgurnus dabryanus Sauvage) by cell-to-cell electrofusion

To study nuclear transfer in the leach (Paramisgurnus dabryanus Sauvage), blastula and gastrula cells were fused with UV-inactivated oocytes by cell-to-cell electrofusion. To facilitate nuclear transfer, blastula and gastrula cells were cultured or incubated at 4 degreesC in different solutions. TC-199 medium supplemented with 20% calf serum was the best culture solution, and effectively retained the totipotence of blastula or gastrula cells for up to 10 days, It was found that gastrula cells incubated at 4 degreesC had the same totipotence as blastula cells, The optimal UV dosage for inactivation of the oocyte chromatin was 180-240 mJ cm(-2). Electrofusion was carried out in a cone-shaped fusion chamber, which permitted the recipient oocyte and the donor blastula cell to contact one another. The electrofusion procedure resulted in a 10% success rate of normal-appearing fish. Genetic analysis indicated that the nuclear material originated from the donor cell (blastomere) and the oocyte pronucleus did not take part in development.

基于直接数值模拟的超/高超声速湍流模型评估与改进

对来流Mach数2.25及6的平板边界层湍流进行了直接数值模拟，并通过与理论、实验及他人计算结果的对比对数值结果进行了验证。基于直接数值模拟得到的湍流数据库，对常用的湍流模型进行了先验评估。评估的湍流模型有k-e模型（包括标准k-e模型、可实现的k-e模型及低Reynolds数k-模型）、SA模型及BL模型。结果显示，可实现的k-e模型的具有较好的预测能力，而标准k-e模型预测的湍流粘性系数偏高。SA模型在边界层内层预测准确度较高，而在外层预测值偏高。对于高Mach数情况，原始的BL模型严重低估了内-外层交界位置，造成湍流粘性系数预测值偏低。作者通过修改模型系数及内-外层交界位置对BL模型进行了修改，修改后模型预测的湍流粘性系数与DNS给出的值吻合较好

ACHT操作引起黑麦(Secale cereale)染色体的畸变和重建

用ACHT处理黑麦萌动种子，对修复前后材料的观察和分析结果表明：1. ACHT操作引起染色体数目变化和染色体断裂损失。在一定 条件和范围内，不同处理引起的这种变化具有显著差异，条件越剧烈，染色体数目变化的范围和频率愈大，断片发生的数量和频率 也愈高，同时修复前后染色体数目的变化范围和频率与断片发生的数量和频率以及它们的修复频率均表现明显的相关性。2. ACHT 操作引起染色体畸变的多样性。经ACHT处理后，胚根细胞染色体有4种断裂方式，包括着丝粒断裂、次溢痕断裂、长臂断裂和短臂 断裂等，其中着丝断裂频率最高；产生6种断片类型，包括有着丝粒和端粒的、有着丝粒而无端粒的、有部分着丝粒和端粒的、有 部分着丝粒而无端粒的、只有端粒的、既无着丝粒也无端粒的断片等。3. ACHT操作引起遗传结构重建的多样性。经ACHT处理后， 对修复24-72小时材料进行核型比较(按Stebbins 和 Levan 标准)和随体分析，处理细胞在染色体数目、大小、形态、位置等方面 均发生显著变化，说明ACHT处理使这些细胞的染色体结构和染色体组型发生了深刻变化。进一步通过Giemsa C— 带分析，观察到 多种重建染色体类型，包括易位型染色体、附加型染色体、无着丝粒染色体、化染色体、增加的m染色体以及某些带型特异的染色 体等。4. RAPD 分析从分子水平上验证了ACHT能有效地引起遗传结构的改变。所用10种引物对处理和对照材料基因组DNA的扩增产 物在条带数目、条带位置及带型特征等方面均有明显差异，其中4种引物出现条带减少，6种引物出现条带增加，后者还包括一个带 位移动。这说明两种材料的基因组DNA具有明显的RAPD反应多态性差异。This paper descripes some results draw on the basis of the observation and analysis on the rye before and after repaired through treating its budding seeds by ACHT in contrast to without ACHT: 1. ACHT manipulation caused the number variation and breakage damage of rye chromosome. Within certain conditions and timits, this phenomenon caused by different treats had signifcant difference: the more the treatment condition is drastie, the more the chageable range and frequence of rye chromosomae number, and so is the produced fragments. Meanwhile, there existed striking relationship among the changeable range and frequence of rye chromosome, the produced number and frequence of fragments and repairing frequence. 2. ACHT manipulafion engendered the diversify of rye chromosomal aberration. Four breakage patterns and six sorts of fragment were observed by watching the chromosome of the rye radicle treated by ACHT, including centric breakage (occuring in the highest frequence), secondary constriction breakage, long arm breakage and short arm breakage to the former, Comprising that with both centromere and telomere, that with centromere and without telomere, that with partial centromere and with telomere, that with partfial ceetromere and without telomere, that only with telomere and that neither with centromere nor with telomere, etc. 3. ACHT manipulation engendered the diversify or rye genetic structs reconstruction. Karureotype analysis(according to Stebbins and Levan) and satellite anaeysis were carried out to rye radicle through 24-72-hour-long recoverage after ACHT manipulation, which showed remarkable change happened on the rye chromosomal number、shape、arm ration and pattern, etc. and also on the satellite number、size、shape and location etc. Those indicated that ACHT manipulation engendered violent changes to rye chromatin structure and chromosome type. Further Giemsa C-banding analysis showed many types of reconstructed chromosome, such as translocation、addition、without centromere、st and other chromosome. 4. RAPD analysis checked the validity of ACHT on changing genetic structure of rye on the level of molecular biology. The treated and recovered rye has different amplifying band pattern by using IO valid arbitary primers selected from 40 comparing with the control.

Production of heavy and superheavy nuclei in massive fusion reactions

Within the framework of a dinuclear system (DNS) model, the evaporation-residue excitation functions and the quasi-fission mass yields in the 48Ca induced fusion reactions are investigated systematically and compared with available experimental data. Maximal production cross sections of superheavy nuclei based on stable actinide targets are obtained. Isotopic trends in the production of the superheavy elements Z = 110, 112–118 based on the actinide isotopic targets are analyzed systematically. Optimal evaporation channels and combinations as well as the corresponding excitation energies are proposed. The possible factors that influencing the isotopic dependence of the production cross sections are analyzed. The formation of the superheavy nuclei based on the isotopes U with different projectiles are also investigated and calculated.