Pretreatment of corn stover by aqueous ammonia

Corn stover was pretreated with aqueous ammonia in a flow-through column reactor, a process termed ammonia recycled percolation (ARP). This method was highly effective in delignifying of the biomass, reducing the lignin content by 70-85%. Most lignin removal occurred within the first 20 min of the process. Lignin removal by ARP was further confirmed by FTIR analysis and lignin staining. The ARP process solubilized 40-60% of the hemicellulose but left the cellulose intact. The solubilized carbohydrate existed in oligomeric form. Carbohydrate decomposition during the pretreatment was insignificant. Corn stover treated for 90 min exhibited enzymatic digestibility of 99% with 60 FPU/g of glucan enzyme loading, and 92.5% with 10 FPU/g of glucan. The digestibility of ARP treated corn stover was substantially higher than that of alpha-cellulose. The enzymatic digestibility was related with the removal of lignin and hemicellulose, perhaps due to increased surface area and porosity. The SEM pictures indicated that the biomass structure was deformed and its fibers exposed by the pretreatment. The crystallinity index increased with pretreatment reflecting removal of the amorphous portion of biomass. The crystalline structure of the cellulose in the biomass, however, was not changed by the ARP treatment.     

Production of recombinant hirudin in galactokinase-deficientSaccharomyces cerevisiae by fed-batch fermentation with continuous glucose feeding

The artificial gene coding for anticoagulant hirudin was placed under the control of theGAL10 promoter and expressed in the galactokinase-deficient strain (Δgal1) ofSaccharomyces cereivisiae, which uses galactose only as a gratuitous inducer in order to avoid its consumption. For efficient production of recombinant hirudin, a carbon source other than galactose should be provided in the medium to support growth of the Δgal1 strain. Here we demonstrate the successful use of glucose in the fed-batch fermentation of the Δgal1 strain to achieve efficient production of recombinant hirudin, with a yield of up to 400 mg hirudin/L.     

Egr-1, a new downstream molecule of Epstein-Barr virus latent membrane protein 1

To investigate the effect of Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) on human cancer cells, we sought to identify and analyze potential target genes that were differentially expressed in the presence and absence of LMP1. Our cDNA microarray analysis revealed that expression of early growth response gene-1 (Egr-1) was increased by LMP1 expression in MCF7 and Jurkat cells. An NFkappaB inhibitor (SN50) antagonized LMP1-induced enhancement of Egr-1 expression, indicating that LMP1 induced Egr-1 via NFkappaB. Furthermore, three lines of evidence indicated that Egr-1 was required for LMP1-induced cancer cell survival. First, Egr-1 expression enhanced the survival of doxorubicin-treated MCF7 cells. Second, inhibition of Egr-1 expression by siRNA (siEgr-1) effectively suppressed LMP-1-induced survival of MCF7 cells. Third, Egr-1 knockdown decreased LMP1-induced expression of Bfl-1. Similar relationships among EBV infection, Egr-1 and drug resistance were also observed in tissues of peripheral T-cell lymphoma-unspecified (PTCL-u) patients.     

8-Hydroxydeoxyguanosine induces senescence-like changes in KG-1, human acute myelocytic leukemia cell line

8-Hydroxydeoxyguanosine (oh⁸dG) treatment induced senescence-like changes in KG-1 cells, a human acute myelocytic leukemia cell line. The oh⁸dG-treated cells stained positive for senescence associated β-galactosidase (SA-β-galactosidase) and had enlarged cell shape, both of which are senescence indexes. The oh⁸dG-treated cells were also cell growth inhibited and arrested at G₁ in the cell cycle. The accumulation of cdk (cyclin dependent kinase) inhibitors, such as p16, p21, and p27, also implies that cellular senescence was induced in oh⁸dG-treated cells. However, these changes were not accompanied by cell differentiation or telomerase activity. Taken together, we conclude that oh⁸dG treatment of KG-1 cells induces cellular senescence.     

Enrichment of electrochemically active bacteria using a three-electrode electrochemical cell

Electrochemically active bacteria were successfully enriched in an electrochemical cell using a positively poised working electrode. The positively poised working electrode (+0.7 V vs. Ag/AgCl) was used as an electron acceptor for enrichment and growth of electrochemically active bacteria. When activated sludge and synthetic wastewater were fed to the electrochemical cell, a gradual increase in amperometric current was observed. After a period of time in which the amperometric current was stabilized (generally 8 days), linear correlations between the amperometric signals from the electrochemical cell and added BOD (biochemical oxygen demand) concentrations were established. Cyclic voltammetry of the enriched electrode also showed prominent electrochemical activity. When the enriched electrodes were examined with electron microscopy and confocal scanning laser microscopy, a biofilm on the enriched electrode surface and bacterium-like particles were observed. These experimental results indicate that the electrochemical system in this study is a useful tool for the enrichment of an electrochemically active bacterial consortium and could be used as a novel microbial biosensor.     

Improved fuel cell and electrode designs for producing electricity from microbial degradation

A new one-compartment fuel cell was composed of a rubber bunged bottle with a center-inserted anode and a window-mounted cathode containing an internal, proton-permeable porcelain layer. This fuel cell design was less expensive and more practical than the conventional two-compartment system, which requires aeration and a ferricyanide solution in the cathode compartment. Three new electrodes containing bound electron mediators including a Mn(4+)-graphite anode, a neutral red (NR) covalently linked woven graphite anode, and an Fe(3+)-graphite cathode were developed that greatly enhanced electrical energy production (i.e., microbial electron transfer) over conventional graphite electrodes. The potentials of these electrodes measured by cyclic voltametry at pH 7.0 were (in volts): +0.493 (Fe(3+)-graphite); +0.15 (Mn(4+)-graphite); and -0.53 (NR-woven graphite). The maximal electrical productivities obtained with sewage sludge as the biocatalyst and using a Mn(4+)-graphite anode and a Fe(3+)-graphite cathode were 14 mA current, 0.45 V potential, 1,750 mA/m(2) current density, and 788 mW/m(2) of power density. With Escherichia coli as the biocatalyst and using a Mn(4+)-graphite anode and a Fe(3+)-graphite cathode, the maximal electrical productivities obtained were 2.6 mA current, 0.28 V potential, 325 mA/m(2) current density, and 91 mW/m(2) of power density. These results show that the amount of electrical energy produced by microbial fuel cells can be increased 1,000-fold by incorporating electron mediators into graphite electrodes. These results also imply that sewage sludge may contain unique electrophilic microbes that transfer electrons more readily than E. coli and that microbial fuel cells using the new Mn(4+)-graphite anode and Fe(3+)-graphite cathode may have commercial utility for producing low amounts of electrical power needed in remote locations.     

Monitoring and classification of PAH toxicity using an immobilized bioluminescent bacteria

An immobilized recombinant bioluminescent Escherichia coli strain, harboring a lac::luxCDABE fused plasmid, which shows lower bioluminescence levels when cellular metabolism is inhibited, was used to monitor the cellular toxicity of polycyclic aromatic hydrocarbons (PAHs). PAHs, classified as pericondensed (PCPAHs) or catacondensed (CCPAHs) according to their molecular structures, were differentiable according to the response of this biosensor. Only CCPAHs were found to cause cellular toxicity, resulting in a dose-dependent decrease in the bioluminescent output. The induction of cellular toxicity by CCPAHs and PCPAHs was compared with acute toxicity predictions obtained using the quantitative structure-activity relationship (QSAR) model. A good relationship was obtained between the toxicities determined with the bioluminescent response of the immobilized bacterium GC2 and the QSAR model. It was also found that the present study offers a new method of predicting the cellular toxicities of CCPAHs or PCPAHs using this biosensor.     

The two NK-1 binding sites are distinguished by one radiolabelled substance P analogue

Two non-stoichiometric binding sites had previously been characterized for the NK-1 receptor using two different types of radiolabelled analogues of substance P. However, the question remained on their eventual conformational interconversion induced or not by the ligand. In this study, kinetic, saturation, and competition studies using [3H]propionyl[Pro(9)]SP demonstrate the existence of two independent binding components in CHO cells transfected with the human NK-1 receptor, with K(d) values of 0.040 nM ( approximately 20% of total sites) and 5.9 nM ( approximately 80% of total sites) that correspond to those of the two previously described binding sites. These two binding sites do not seem to interconvert since the minor one can be selectively extinguished in saturation studies in the presence of a SP analogue specific of this binding site.     

Mechanism of activation of human c-KIT kinase by internal tandem duplications of the juxtamembrane domain and point mutations at aspartic acid 816

The patients suffering from acidosis usually sign psychological deficits. The cerebral dysfunction is reportedly caused by an acid-induced functional impairment of GABAergic neurons; however, the role of pyramidal neurons in this process remains unclear. By using electrophysiological method and changing extracellular pH, we investigated the influence of acidic environment on pyramidal neurons in the cortical slices, such as their ability of firing spikes and response to synaptic inputs. A low pH of artificial cerebral spinal fluid elevates the responses of pyramidal neurons to excitatory synaptic inputs and their ability of encoding digital spikes, as well as reduces the signal transmission at GABAergic synapses. The elevated ability of neuronal spiking is associated with the decreases of refractory periods and threshold potentials. Therefore, acidosis deteriorates brain functions through making the activities between cortical pyramidal neurons and GABAergic neurons imbalanced toward the overexcitation of neural networks, a process similar to neural excitotoxicity.     

Antioxidant and anticancer activity of extract from Betula platyphylla var. japonica

The antioxidant and anticancer properties of a medicinal plant, Betula platyphylla var. japonica were investigated. The total methanol extract of B. platyphylla var. japonica had protective effects against hydrogen peroxide (H2O2) in the Chinese hamster lung fibroblast (V79-4) cell line and induced apoptotic cell death in human promyelocytic leukemia (HL-60) cells, a cancer cell line. B. platyphylla var. japonica extract significantly increased cell viability against H2O2. The extract also showed high 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity (IC50 2.4 microg/ml) and lipid peroxidation inhibitory activity (IC50 below 4.0 microg/ml). Furthermore, B. platyphylla var. japonica extract reduced the number of V79-4 cells arrested in G2/M in response to H2O2 treatment and increased the activities of several cellular antioxidant enzymes, including superoxide dismutase, catalase and glutathione peroxidase. Treatment with B. platyphylla var. japonica extract induced cytotoxicity and apoptosis in HL-60 cells, as shown by nucleosomal DNA fragmentation, increases in the subdiploid cell population, and fluorescence microscopy. B. platyphylla var. japonica extract gradually increased the expression of pro-apoptotic Bax and led to the activation of caspase-3 and cleavage of PARP. These findings suggest that B. platyphylla var. japonica exhibits potential antioxidant and anticancer properties.