The cell agglutination agent, phytohemagglutinin-L, improves the efficiency of somatic nuclear transfer cloning in cattle (Bos taurus)

One of the several factors that contribute to the low efficiency of mammalian somatic cloning is poor fusion between the small somatic donor cell and the large recipient oocyte. This study was designed to test phytohemagglutinin (PHA) agglutination activity on fusion rate, and subsequent developmental potential of cloned bovine embryos. The toxicity of PHA was established by examining its effects on the development of parthenogenetic bovine oocytes treated with different doses (Experiment 1), and for different durations (Experiment 2). The effective dose and duration of PHA treatment (150 microg/mL, 20 min incubation) was selected and used to compare membrane fusion efficiency and embryo development following somatic cell nuclear transfer (Experiment 3). Cloning with somatic donor fibroblasts versus cumulus cells was also compared, both with and without PHA treatment (150 microg/mL, 20 min). Fusion rate of nuclear donor fibroblasts, after phytohemagglutinin treatment, was increased from 33 to 61% (P < 0.05), and from 59 to 88% (P < 0.05) with cumulus cell nuclear donors. The nuclear transfer (NT) efficiency per oocyte used was improved following PHA treatment, for both fibroblast (13% versus 22%) as well as cumulus cells (17% versus 34%; P < 0.05). The cloned embryos, both with and without PHA treatment, were subjected to vitrification and embryo transfer testing, and resulted in similar survival (approximately 90% hatching) and pregnancy rates (17-25%). Three calves were born following vitrification and embryo transfer of these embryos; two from the PHA-treated group, and one from non-PHA control group. We concluded that PHA treatment significantly improved the fusion efficiency of somatic NT in cattle, and therefore, increased the development of cloned blastocysts. Furthermore, within a determined range of dose and duration, PHA had no detrimental effect on embryo survival post-vitrification, nor on pregnancy or calving rates following embryo transfer.     

Apoptotic effects of over-expressed estrogen receptor-beta on LoVo colon cancer cell is mediated by p53 signalings in a ligand-dependent manner

Epidemiologic studies reported that the prevalence of hereditary non-polyposis colon cancer (HNPCC) in male is about 1.5-fold higher than that in female. Decreases in circulatory estradiol (E2) have been reported to downregulate the expression of E2 receptor (ER) and significantly increase the risk of colorectal cancer. Patients that received E2 replacement therapy were found to have a reduction in the incidence of colon adenoma and carcinoma. Furthermore, significant decreases in the expression of ER have been found in colorectal cancer specimens. These data strongly suggest the protective roles of E2 and ER against colorectal cancer. However, the mechanisms remain unexplored. LoVo cells were transient transfected to overexpress ER-beta, DNA fragmentation and caspase activity assay were performed to evaluate apoptotic effects. Western blotting was used to evaluate protein levels, and luciferase activity assay to measure the TNF-alpha promoter activity. Our data clearly demonstrated that E2 and ER-beta alone could upregulate p21 and p27 proteins, which further activate caspase-8 and caspase-9 to induce apoptosis in LoVo cell, and the ER-beta. effects were enhanced by E2. However, overexpressed ER-beta did not influence the expression and promoter activity of TNF-alpha. In addition, E2 and overexpressed ER-beta downregulated the beta-catenin proteins which cause the downregulation of its target genes, cyclin D1 and Rb, to inhibit the cell cycle and cell proliferation. The results indicate that overexpressed ER-beta may induce LoVo cell apoptosis and anti-proliferation by increasing p53 signaling in a ligand-dependent manner, and without hTNF-alpha involvement. Efforts aiming at enhancing ER-beta expression and/or activity may prove to be an attractive alternative therapy against colorectal cancer.     

A novel feeding strategy for enhanced plasmid stability and protein production in recombinant yeast fedbatch fermentation

A novel feeding strategy in fedbatch recombinant yeast fermentation was developed to achieve high plasmid stability and protein productivity for fermentation using low-cost rich (non-selective) media. In batch fermentations with a recombinant yeast, Saccharomyces cerevisiae, which carried the plasmid pSXR125 for the production of beta-galactosidase, it was found that the fraction of plasmid-carrying cells decreased during the exponential growth phase but increased during the stationary phase. This fraction increase in the stationary phase was attributed to the death rate difference between the plasmid-free and plasmid-carrying cells caused by glucose starvation in the stationary phase. Plasmid-free cells grew faster than plasmid-carrying cells when there were plenty of growth substrate, but they also lysed or died faster upon the depletion of the growth substrate. Thus, pulse additions of the growth substrate (glucose) at appropriate time intervals allowing for significant starvation period between two consecutive feedings during fedbatch fermentation should have positive effects on stabilizing plasmid and enhancing protein production. A selective medium was used to grow cells in the initial batch fermentation, which was then followed with pulse feeding of concentrated non-selective media in fedbatch fermentation. Both experimental data and model simulation show that the periodic glucose starvation feeding strategy can maintain a stable plasmid-carrying cell fraction and a stable specific productivity of the recombinant protein, even with a non-selective medium feed for a long operation period. On the contrary, without glucose starvation, the fraction of plasmid-carrying cells and the specific productivity continue to drop during the fedbatch fermentation, which would greatly reduce the product yield and limit the duration that the fermentation can be effectively operated. The new feeding strategy would allow the economic use of a rich, non-selective medium in high cell density recombinant fedbatch fermentation. This new feeding strategy can be easily implemented with a simple IBM-PC based control system, which monitors either glucose or cell concentration in the fermentation broth.     

Further Daphniphyllum Alkaloids with Insecticidal Activity from the Bark of Daphniphyllum macropodum Miq.

Two new Daphniphyllum alkaloids, macropodumines J and K ( 1 and 2 , resp.), together with six known structurally related alkaloids, 3 – 8 , were isolated from the bark of Daphniphyllum macropodum Miq . The structures of the new compounds 1 and 2 were elucidated on the basis of a comprehensive analysis of their spectroscopic and chemical data. Macropodumine J ( 1 ) contains a CN group which is relatively rare in naturally occurring alkaloids. All isolated compounds were tested for their insecticidal activities against a number of insect species. Daphtenidine C ( 5 ) is the most active compound against Plutella xylostella. This is the first report of insecticidal properties of Daphniphyllum alkaloids.     

Combinatorial signals of activin/nodal and bone morphogenic protein regulate the early lineage segregation of human embryonic stem cells

Cell fate commitment of pre-implantation blastocysts, to either the inner cell mass or trophoblast, is the first step in cell lineage segregation of the developing human embryo. However, the intercellular signals that control fate determination of these cells remain obscure. Human embryonic stem cells (hESCs) provide a unique model for studying human early embryonic development. We have previously shown that Activin/Nodal signaling contributes to maintaining pluripotency of hESCs, which are derivatives of the inner cell mass. Here we further demonstrate that the inhibition of Activin/Nodal signaling results in the loss of hESC pluripotency and trophoblast differentiation, similar to BMP4-induced trophoblast differentiation from hESCs. We also show that the trophoblast induction effect of BMP4 correlates with and depends on the inhibition of Activin/Nodal signaling. However, the activation of BMP signaling is still required for trophoblast differentiation when Activin/Nodal signaling is inhibited. These data reveal that the early lineage segregation of hESCs is determined by the combinatorial signals of Activin/Nodal and BMP.     

Technical note phytoremediation of triazophos by Canna indica Linn. in a hydroponic system

The phytoremediation of triazophos (O, O-diethyl-O-(1-phenyl-1, 2, 4-triazole-3-base) sulfur phosphate, TAP) by Canna indica Linn. in a hydroponic system was studied. After 21 d of exposure, the removal kinetic constant (K) of TAP was 0.0229-0.0339 d(-1) and the removal percentage of TAP was 41-55% in the plant system and the K and removal percentage of TAP were about 0.002 d(-1) and 1%, respectively, in darkness and disinfected control. However, the K and removal percentage of TAP were 0.006 d(-1) and approximately 11%, respectively, in the treatment with eluate from the media of constructed wetland. The contribution of plant to the remediation of TAP was 74% and C. indica played the most important role in the hydroponic system. Under the stress of TAP and without inorganic phosphorus nutrient, the activity of phosphatase in the plant system increased and phytodegradation was observed. The production and release of phosphatase is seen as the key mechanism for C. indica to degrade TAP. C. indica, which showed the potential of phytoremediation of TAP, and is commonly used in constructed wetland, so the technique of phytoremediation of TAP from contaminated water can be developed with the combination of constructed wetland.     

Change in microbial communities in acetate- and glucose-fed microbial fuel cells in the presence of light

Power densities produced by microbial fuel cells (MFCs) in natural systems are changed by exposure to light through the enrichment of photosynthetic microorganisms. When MFCs with brush anodes were exposed to light (4000 lx), power densities increased by 8–10% for glucose-fed reactors, and 34% for acetate-fed reactors. Denaturing gradient gel electrophoresis (DGGE) profiles based on the 16S rRNA gene showed that exposure to high light levels changed the microbial communities on the anodes. Based on 16S rRNA gene clone libraries of light-exposed systems the anode communities using glucose were also significantly different than those fed acetate. Dominant bacteria that are known exoelectrogens were identified in the anode biofilm, including a purple nonsulfur (PNS) photosynthetic bacterium, Rhodopseudomonas palustris, and a dissimilatory iron-reducing bacterium, Geobacter sulfurreducens. Pure culture tests confirmed that PNS photosynthetic bacteria increased power production when exposed to high light intensities (4000 lx). These results demonstrate that power production and community composition are affected by light conditions as well as electron donors in single-chamber air-cathode MFCs.     

北京西山侏罗纪大同锥叶蕨的研究

大同锥叶蕨Coniopteris tatungensis 是侏罗纪植物群常见的分子。由于对该植物特征认识的不同, 对其在分类上的归属有不同意见,或归入膜蕨型锥叶蕨C．hymenophylloides,或归入简单锥叶蕨C. simplex。通过对产自北京西山斋堂地区窑坡组下段的标本的深入研究,特别是用扫描电子显微镜对该 标本的孢子囊群和孢子囊的观察,确认大同锥叶蕨在锥叶蕨属的种级分类上的独立性。文中还初步讨论了其分类和地理分布。     

地上部植食者褐飞虱对不同水稻品种土壤线虫群落的影响

地上和地下部生物群落的交互作用对于调控陆地生态过程具有重要作用。在盆栽条件下利用2×2析因设计研究了褐飞虱(Nilaparvata lugens)取食不同水稻品种后对土壤线虫群落的影响。结果表明, 褐飞虱侵害水稻9 d后, 感虫品种(广四和汕优63)的土壤线虫总数、属数及自生线虫(食细菌线虫、食真菌线虫和捕食性线虫)数量增加, 并且一般达到显著水平(P<0.05); 而上述指标在抗虫品种(汕优559和IR36)土壤中则呈现相反的趋势。植食性线虫数量在强感虫品种广四上显著增加(P<0.05), 而在强抗虫品种IR36上显著减少(P<0.05)。褐飞虱和水稻品种对土壤线虫的生态指数(线虫通道指数、Shannon-Wiener指数、成熟度指数、富集指数和结构指数)没有明显影响, 可能与供试土壤线虫群落组成单一及褐飞虱作用时间较短有关。总之, 褐飞虱强烈影响土壤线虫数量、群落组成和营养结构, 并且作用的方向(促进或抑制)和程度依赖于水稻的品种特性, 揭示出地上部植食者的短期侵害将对稻田土壤生态系统的结构和功能产生深远影响。