Inhibition of PKB/Akt activity involved in apigenininduced apoptosis in human gastric carcinoma cells

Apigenin is a flavonoid widely distributed in fruits and vegetables. It possesses growth inhibitory properties against numerous cancer cell lines. However,the molecular mechanism(s) by which api-genin elicits its effects have not been fully elucidated. Here we studied whether apigenin inhibits growth and induces apoptosis in human gastric carcinoma cells. We showed that the flavonoid inhibited growth of the cells and caused apoptosis,as evidenced by DNA Ladder,cleavage of pro-caspase-3 in a time-dependent manner. Induction of apoptosis was dependent on inhibition of the PKB/Akt activity. We found that while apigenin had no effect on the expression of Akt and Bad,it inhibited specific phosphorylation of the two proteins that are associated with pro-survival mechanisms. We propose that this important flavonoid induces apoptosis in gastric cancer cells by inhibiting Akt activity. Since Akt is often activated in cancers,our findings may have clinical implications.     

Effect of HS2 and HS3 elements on erythroid-specific expression in transgenic mice

The expression plasmids CMV/GFP, HS2ALL, HS3ALL and HS23ALL were selected to investigate the effect of HS2 and HS3 element on erythroid-specific expression in transgenic mice. These plasmids were digested with restriction enzymes and purified. And five DNA fragments, CMV/GFP, HS2/GFP, CMV/HS2/GFP, HS23/GFP and HS3/GFP were obtained. After purification, the above DNA fragments were microinjected into the pre-nuclei of the mice fertilized eggs and transgenic mice were generated, with an integration rate of 10.89%. The green fluorescence protein(GFP) expression in many transgenic mouse tissues was determined by FACS analysis. The results showed that the HS2 and 1.7 kb of β-globin gene promoter were sufficient for the erythroid-specific expression of β-globin gene. The GFP expression of different recombinant constructs was also analyzed in blood of all the transgenic mice with FACS. The results indicated that HS2 and HS3 had the same enhancement activity on the regulation of β-globin gene expression. Moreover, these two elements showed a significant synergistic effect on gene expression at the transgenic mouse level, although the GFP expression varied largely among different transgenic mouse litters.     

Depletion of phosphatidyl-glycerol head-group induces changes in oxygen evolution and protein secondary struc-tures of photosystem Ⅱ

The techniques of oxygen electrode polarogra-phy and Fourier transform infrared (FT-IR) spectroscopy were employed to explore the roles of polar head-group of phosphatidylglycerol (PG) molecules in the functional and structural aspects of photosystem Ⅱ (PS Ⅱ) through enzymatic approach. It was shown that the depletion of PG by treatment of phospholipase C (PLC) on PS Ⅱ particles caused the inhibition of oxygen evolving activity in PS Ⅱ. This effect also gave rise to changes in the protein secondary structures of PS Ⅱ, that is, an increase in a-helical conformation which is compensated by the loss of p-strand structures. It revealed that the head-group of PG molecules plays an important structural role in the maintenance of normal structure of PS Ⅱ proteins, which is required to maintain the appropriate physiological activity of the PS Ⅱ complex such as the oxygen evolving activity. It is suggested that there most probably exist hydrogen-bonding interactions between PG molecules and PS Ⅱ proteins.     

Catalytic combustion properties of nanocomplex oxide for natural gas

The catalysts of copper oxide supported on cerium dioxide were prepared by different methods for methane catalytic combustion. The effects of copper content in the catalysts and calcination temperatures of the catalysts on the catalytic activity are investigated. Results show that the complex oxide catalyst exhibits high catalytic activity for methane combustion due to the synergistic effect of CuO and CeO₂. The catalyst prepared by impregnation is more active than that prepared by controlled coprecipitation even if CuO content is the same. When W(CuO)<13%, the light-off temperature and full conversion temperature for the CH₄ reaction decrease with the increasing of CuO content in the catalysts. However, when the copper content is above 13%, the excess CuO has a negative effect on the catalytic activity owing to the formation of bulk CuO particles. A proper calcinations temperature of 650 ℃ can lead to a high dispersion of CuO and accordingly can enhance the catalytic activity of the composites.     

Comparative research on the catalytic activities of different molecular sieves for acetalization and ketalization

Molecular sieves with different acidities and pore-sizes have been applied to catalyze the acetalization and ketalization of carbonyl compounds with glycol. A comparative research on the catalytic activities of the catalysts for certain reactions has been carried out to figure out the relationship between the characters of the catalysts and the activities. The results show that the pore-size of the molecular sieves is one of the most important factors that determine the catalytic activity. Appropriate pore-size is needed for the certain reaction.The acidity is another factor that affects the activities. They also have great effects on the recycled activities of the catalysts.     

Rational design of glycerol dehydratase： Swapping the genes encoding the subunits of glycerol dehydratase to improve enzymatic properties

1,3-propanediol (1,3-PD) is an important material for chemical industry,and there has been always much interest in the production of 1,3-PD using all possible routes. The genes encoding glyc-erol dehydratase (GDHt) from Citrobacter freundii,Klebsiella pneumoniae and metagenome were cloned and expressed in E. coli. All glycerol dehy-dratases but the one from metagenome could be detected to show enzyme activities. In order to im-prove the enzymatic properties of GDHts,the genes encoding α and β-γ subunits were cloned,and the enzyme characteristics were evolved by rational de-sign based on their 3D structures which were con-structed by homology modeling. Six heteroenzymes were obtained by swapping the α subunit genes of these three different-source-derived GDHts. The pH,thermal stability and Vmax of some heteroenzymes were dramatically improved by 2―5 times compared with the wild one (GDHtKP). The GDHt cloned from metagenome,originally proved to be with no enzyme activity,was converted into active enzyme by swap-ping its subunits with other different GDHts. In addi-tion,the effect of subtle 3D structural changes on the properties of the enzyme was also observed.     

A new mechanism of invader success： Exotic plant inhibits natural vegetation restoration by changing soil microbe community

Since the 1950s of the last century, the exotic plant, Eupatorium adenophorum, has spread rapidly across southwest China, damaging native ecosystems and causing great economic losses. We examined the pH, N, P, K, and organic matter concentrations, and the bacterial community character (by Biolog EcoPlateTM) in soils from sites heavily and lightly invaded by this exotic species. Also, soil from the lightly invaded site was treated with a water extract of E.adenophorum roots to examine the effect of the plant on soil properties. We grew three plant species, one native and two exotic, in pot experiment using soil from heavily invaded site to examine the effects of the soil on these plants growth. The soil analysis demonstrated that the pH, organic matter, total N, total P and total K in soils from the heavily invaded site were only slightly different from those of the lightly invaded site, but concentrations of NH4^ , NO3^- and available P and K in the heavily invaded site were greater than those in the lightly invaded site. The catabolic activity of soil bacterial community in the heavily invaded site was different from that in the lightly invaded site. The catabolic activity of bacterial community in soils treated by the water extract of E.adenophorum roots changed and became similar to that in soils from the heavily invaded site. The pot experiment showed that the exotic plants growth in heavily invaded soil were not different from in lightly invaded soil; however, the native plant biomass decreased dramatically when grown in soil from the heavily invaded site as compared to soil from the lightly invaded site; and the same phenomenon was found when any potential allelopathic effects by E. adenophorum were eliminated by added activated carbon to those soils. Difference in soil nutrient availability and allelopathy could not explain this phenomenon of the native plant in the soils from the heavily and lightly invaded sites. Changes observed in the soil bacterial community were obviously related to native plant growth in those tow soils. Those results suggest that changing soil microbial community may be an important part of E. adenophorum invasion process. Since the soil microbial community serves as bridge in connection of exotic and natural plants, the exotic plant could inhibit the natural plant growth and reproduction by changing the soil microbial community in invaded site.     

Depletion of phosphatidylglycerol head-group induces changes in oxygen evolution and protein secondary structures of photosystemⅡ

The techniques of oxygen electrode polarography and Fourier transform infrared (FT-IR) spectroscopy were employed to explore the roles of polar head-group of phosphatidylglycerol (PG) molecules in the functional and structural aspects of photosystemⅡ(PSⅡ) through enzymatic approach. It was shown that the depletion of PG by treatment of phospholipase C (PLC) on PSⅡ particles caused the inhibition of oxygen evolving activity in PSⅡ. This effect also gave rise to changes in the protein secondary structures of PSⅡ, that is, an increase in α-helical conformation which is compensated by the loss of β-strand structures. It revealed that the head-group of PG molecules plays an important structural role in the maintenance of normal structure of PSⅡ proteins, which is required to maintain the appropriate physiological activity of the PSⅡ complex such as the oxygen evolving activity. It is suggested that there most probably exist hydrogen-bonding interactions between PG molecules and PSⅡ proteins.     

Equilibrium dialysis study on the interaction between Cu(Ⅱ) and HSA or BSA

The interaction of Cu(Ⅱ) and human serum albumin (HSA) or bovine serum albumin (BSA) at physiological pH is studied by equilibrium dialysis. The successive stability constants are obtained by non-linear least square methods fitting Bjerrum formula. For both the Cu(Ⅱ)-HSA and Cu(Ⅱ)-BSA systems, the order of magnitude of K 1 and K 2 was found to be ≈10 4 mol -1·dm 3. There are about twenty stoichiometry binding sites found in one HSA or BSA molecule. They can be divided into two or three sets. Results of equilibrium dialysis experiments suggest that there exists one strong metal binding site in both Cu(Ⅱ)-HSA and Cu(Ⅱ)-BSA. It is the imidazol group nitrogen atoms of His 3 that are primarily concerned with copper binding site. After reaching dialysis equilibrium, there is the interaction among the different binding sites, the values of K all deviate from the simple statistical effect except for K-1 and K-2 in both Cu(Ⅱ)-HSA and Cu(Ⅱ)-BSA systems, and the positive cooperative effect is found.     

C-terminal domain of Chk1 regulates its subcellular location and kinase activity for DNA repair

Effector kinase Chk1 is an evolutionarily conserved protein kinase. It is a key mediator linking the mechanisms that monitor DNA integrity to components of the cell cycle engine. In this study, recombinant vectors pEGFP-C1-Chk1/C 288/C 334/C 368 were constructed and transfected into HeLa cells to study the effect of the Chk1 regulatory domain on the regulation of subcellular Chk1 location in response to DNA damage. We found that DNA damage-induced nuclear accumulation is regulated by 34 amino acids (334–368) in the C-terminal regulatory domain. Recombinant vectors pXJ41-Chk1/C 288/C 334/C 368 were co-transfected with reporter plasmid pEGFP-N2 into HeLa cells to study the repair abilities of the different human Chk1 truncation mutants. In addition, recombinant vectors were transfected into HeLa cells to study the effects of the different truncation mutants on the cell cycle. Furthermore, to study the kinase activity of the different truncation mutants, Ser216 phosphorylation of Cdc25C was studied by Western blot analysis. We found that the enzymatic activity of C 368, missing the 108 C-terminal amino acids (368–476), was higher than that of full-length Chk1, and C 368 delayed the cell cycle progression. The enzymatic activity of C 334, missing the 142 C-terminal amino acids (334–476), was equivalent to that of full-length Chk1. C 288, missing the 188 C-terminal amino acids (288–476), had almost no enzymatic activity, suggesting that the regulatory domain contains both inhibitory and regulatory elements. This study provides useful information for further research on Chk1 function.     

Effect of Starch Sources on the Release Rates of Herbicides Encapsulated

The effects of starch sources on the behaviors of starch matrices and on the rates of herbicides released from the matrices were studied for slow release. The starches considered include native corn starch, wheat starch, potato starch and cassava starch. The matrices were prepared through encapsulating 2,4-dichlorophenoxyacetic or 2,4,5 trichlorophenoxyacetic acids as model herbicides with hot-gelatinized starch pastes. The encapsulation was evaluated in terms of herbicide content, swellability, encapsulation efficiency, and release rate. The results show that starch sources play an important role on the matrix behaviors and on release rates. The rate of 2,4-D released follows the order： wheat starch 〈 potaot starch 〈 corn starch 〈 cassava starch. And for the rate of 2,4,5-T, this order is nearly the same only with an exception that the late two kinds of starch are similar. It is also demonstrated that herbicides with different water solubility show different release rates, no matter what type of starch is used as the matrices.     

Kinetics of Inhibition of Polyphenol Oxidase Obtained from Tobacco Nicotiana Tobacum

In this study, the kinetics of inhibition of polyphenol oxidase by L-cysteine has been investigated. The inhibition of tobacco polyphenol oxidase was studied using the progress-of-substrate-reaction method proposed by Tsou, following the substrate reaction during irreversible inhibition of the enzyme activity. Analysis of the inhibition kinetics shows that inhibition occurs by an irreversible and non-complexation reaction. The microscopic rate constants were determined for reaction of the inhibitor both with the free enzyme and with the enzyme-substrate complex. The results show that the presence of the substrate has a significant protective effect of the enzyme against inactivation by L-cysteine.     

Effect of plastic deformation on diffusion-rolling bonding of steel sandwich plates

Diffusion bonding is one of the most important techniques for composite materials, while bonding temperature, holding time,and rolling reduction are the key parameters that affect the bonding strength of sandwich plates. To study the effect of plastic deformation on the bonding strength, laboratory experiments were carried on a Gleeble Thermal Simulator to imitate the diffusion-rolling bonding under different reductions for steel sandwich plates. The bonding strength and interlayer film thickness were measured, and the element diffusion was analyzed using line scanning. The relationship between the bonding strength and “diffused interlayer” thickness was investigated. It has been found that the bonding strength increases with reduction, whereas the interlayer film thickness decreases gradually as the reduction increases. The diffusion under plastic deformation is obviously enhanced in comparison with that of nil reduction. The mechanism of plastic deformation effect on the diffusion bonding and related models have been discussed.     

Experimental investigation of the influence of electric field on frost layer growth under natural convection condition

The influence of direct current (DC) electric field on the thickness and mass of frost on a cold vertical plate was investigated. The photos of frost layer growth were taken with and without the presence of electric field, and results showed that the electric field has a strong influence on the frost thickness. The influences of cold plate temperature and ambient temperature on frost thickness and frost mass were also investigated under the natural convection condition with electric field. Experimental results demonstrated that the cold plate temperature has very strong effect on the frost layer thickness, but its influence on frost mass is minor; the influence of ambient temperature on the frost mass is more obvious than that on the frost thickness.     

Effect of Mo content on the thermal stability of Ti-Mo-bearing ferritic steel

The effects of tempering holding time at 700℃on the morphology,mechanical properties,and behavior of nanoparticles in Ti-Mo ferritic steel with different Mo contents were analyzed using scanning electron microscopy and transmission electron microscopy.The equilibrium solid solution amounts of Mo,Ti,and C in ferritic steel at various temperatures were calculated,and changes in the sizes of nanoparticles over time at different Mo contents were analyzed.The experimental results and theoretical calculations were in good agreement with each other and showed that the size of nanoparticles in middle Mo content nano-ferrite(MNF)steel changed the least during aging.High Mo contents inhibited the maturation and growth of nanoparticles,but no obvious inhibitory effect was observed when the Mo content exceeded 0.37wt%.The tensile strength and yield strength continuously decreased with the tempering time.Analysis of the strengthening and toughening mechanisms showed that the different mechanical properties among the three different Mo content experiment steels were mainly determined by grain refinement strengthening(the difference range was 30-40 MPa)and precipitation strengthening(the difference range was 78-127 MPa).MNF steel displayed an ideal chemical ratio and the highest thermodynamic stability,whereas low Mo content nano-ferrite(LNF)steel and high Mo content nano-ferrite(HNF)steel displayed relatively similar thermodynamic stabilities.     

Effect of Thrombin on the Apoptosis of Hippocampal Neurons in vitro

Hippocampal neurons were treated by thrombin and thrombin receptor activating peptides (TRAP). Cell survival rate was decreased in a dose-dependent manner by MTT assay. The numbers of apoptotic cell and apoptotic rate of hippocampal neurons treated by different concentrations of thrombin were increased in a dose-dependent manner by terminal deoxynucleotidyl transferase (TdT) mediated dUTP-biotin nick end-labeling (TUNEL) method and Flow Cytometry. When the concentration of thrombin is 40 U/mL, TUNEL positive cells and apoptotic rate of hippocampal neurons reached peak value, were 27. 3 4.0 and (29.333 4.633 ) %, respectively. Immunocytochemistry assay show that Bcl-2 protein expression was down- regulated and Bax protein expression was up-regulated with the concentration of thrombin increased. TRAP can mimic the effect of thrombin to induce apoptosis on hippocampal neurons. These data demonstrated that thrombin induced hippocampal neuron apoptosis in a dose-dependent manner through activating protease-activated protein-1 (PAR-1). The change in expression of Bcl-2 and Bax was related with the effect of high concentration thrombin induced apoptosis on hippocampal neurons.     

Molecular mapping of QTLs for root response to phosphorus deficiency at seedling stage in wheat (Triticum aestivum L.)

Phosphorus (P) deficiency in the soil is one of the major abiotic stresses that limit plant growth and crop productivity throughout the world. Development of cultivars with improved P-deficiency tolerance is an efficient strategy for sustainable agriculture. Plant roots play an important role in crop growth and development, especially in nutrient uptake and improvement of P-efficiency. Mapping quantitative trait loci (QTLs) for root traits and their response to low P stress at seedling stage will facilitate the development of P-efficient wheat cultivars. In this study, 30 QTLs (LOD>2.0) were mapped for the three root traits, such as root length, root number and root dry matter under different P supply conditions and their response to P-stress. These QTLs were distributed on 14 chromosomes, with each of the 5 QTLs explaining more than 10% phenotype variance. Analyses showed that root traits and their response to P-deficiency were controlled by different QTLs. In addition, alleles with positive effects were separated on both parents, and wheat cultivars with improved P-efficiency could be developed by accumulating these positive effect alleles together.     

Effect of Mo content on the thermal stability of Ti–Mo-bearing ferritic steel

The effects of tempering holding time at 700°C on the morphology, mechanical properties, and behavior of nanoparticles in Ti–Mo ferritic steel with different Mo contents were analyzed using scanning electron microscopy and transmission electron microscopy. The equilibrium solid solution amounts of Mo, Ti, and C in ferritic steel at various temperatures were calculated, and changes in the sizes of nanoparticles over time at different Mo contents were analyzed. The experimental results and theoretical calculations were in good agreement with each other and showed that the size of nanoparticles in middle Mo content nano-ferrite(MNF) steel changed the least during aging. High Mo contents inhibited the maturation and growth of nanoparticles, but no obvious inhibitory effect was observed when the Mo content exceeded 0.37 wt%.The tensile strength and yield strength continuously decreased with the tempering time. Analysis of the strengthening and toughening mechanisms showed that the different mechanical properties among the three different Mo content experiment steels were mainly determined by grain refinement strengthening(the difference range was 30–40 MPa) and precipitation strengthening(the difference range was 78–127 MPa). MNF steel displayed an ideal chemical ratio and the highest thermodynamic stability, whereas low Mo content nano-ferrite(LNF) steel and high Mo content nano-ferrite(HNF) steel displayed relatively similar thermodynamic stabilities.     

Characterization of the separate kinase domain of chicken liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase

The bifunctional enzvme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase consists of two dis tinct domains which catalyze the synthesis and hydrolysis of fructose-2, 6-bisphosphate, respectively. In this work the properties of the separate 6-phosphofructo-2-kinase domain were investigated. Purification of the expressed separate do main or isolation of this domain from purified glutathione S-transferase (GST) fusion protein with thrombin cleavage led to the loss of its kinase activity. Thus the domain in the GST-tagged form was characterized. The two forms of the do main with different lengths (amino acids 1 ～ 249 and 1 ～ 286) were very similar in kinetic property and could catalyze the formation of fructose-2,6-bisphosphate with a kcat 4-fold lower than that of the full-length enzyme. In addition, the domain was much more sensitive to guanidine inactivation and heat denaturation, and less stable at pH values below 7 than the full-length enzyme. The results suggest that the separate kinase domain of the bifunctional enzyme is far less perfect in structure in the absence of the bisphosphatase domain, though it still possesses the 6-phosphofructo-2-kinase activity.