Expression of human clotting factor Ⅸ mediated by recom- binant lentiviral vector in cultured cells and hemophilia B mice

To explore the expression of human clotting factor Ⅸ (hFⅨ) cDNA in vitro and the feasibility of gene therapy for hemophilia B mice mediated by recombinant lentiviral vector, a recombinant hFⅨ lentiviral vector driven by ubiquitin-C promoter, FUXW, and by ABP liver specific promoter, FAXW, was constructed respectively. Recombinant lentivirus was harvested from 293T cells by calcium phosphate-mediated transient cotransfection of three plasmids (transgene vector, CMV腞8.2, VSV-G). hFⅨ expression was detected in supernatant of 293T, BHK and L-02 cells infected with FUXW virus, whereas higher expression of hFⅨ levels (630 ng/106 cells/48 h) was detected only in L-02 cells infected with FAXW virus. Serum hFⅨ antigen was detected in all hemophilia B mice treated with FAXW virus by tail vein injection, an efficiency level of hFⅨ was observed (45 ng/mL, approximately 1% of normal human levels), the expression lasted for more than 60 d. The results indicated that HIV-based lentiviral vectors offer a promising approach to the gene therapy of hemophilia B.     

Efficient transfer and expression of human clotting factor IX cDNA in neonatal hemophilia B mice mediated by VSV-G pseudotyped retrovirus

The feasibility of in vivo gene therapy for hemophilia B by VSV-G pseudotyped retroviral vector was introduced. The novel packaging cell line 293GPG was used to produce VSV-G/G1NaBAIX pseudotyped virus with the highest titers up to 8.5×10⁸ cfu·mL^-1. In contrast to the conventional retrovirus, VSV-G pseudotyped virus was more resistant to inactivation by serum complements (P＜0.001). Our results also demonstrated that VSV-G pseudotyped virus was more stable in neonatal mice serum than in adult mice serum (P＜0.01). After intraperitoneal injection of different doses of virus, hFIX antigen was detected and lasted for more than 120 d, the highest level reached (72.5±6.1) ng·mL^-1. Moreover, the functional activity was improved to some extent in all hFIX-treated mice, the most remarkable improvement was observed in the mice treated with higher dose of virus whose clotting activity increased to (3.4±1.5)% and APTT (activated partial thromboplastin time) reduced to (43.2±7.2) s. The anti-hFIX antibody was not detected by the method of Bethesda, no germ line transmission and any side effects associated with gene transfer were found. Our results indicated that neonatal gene therapy for hemophilia B mice by VSV-G pseudotyped retrovirus is promising.     

The high-resolution climate recorded in the δ18O ofPorites lutea from the Nansha Islands of China

A Porites lutea core from Yongshu Reef of Nansha Islands covering 50 years growth history was analyzed for oxygen isotopic composition with monthly and seasonally resolution. The calibration of the δ ¹⁸O with the instrumental temperature indicated that the coral δ ¹⁸O is a good indicator for sea surface temperature (SST) and air temperature ( t ). It can be used to reconstruct the SST and air temperature of the Yongshu Reef sea area. In addition, the coral δ ¹⁸O provides signatures for the intensity of the East Asia monsoon and it is a record for the activities of El Niño events. With the calibrated SST and air temperature formulas, the most recent fifty years SST and air temperature were reconstructed based on the coral δ ¹⁸O, thus back up the understanding of the climate of Nansha Islands to 1950, far beyond the limit of the instrumental recording since September 1988. It was found that, in general, increasing 1℃ air temperature results in 0.24‰ decrease in skeletal δ ¹⁸O.     

Expression of biologically active human clotting factor Ⅸ(hFⅨ) in the mammary gland of transgenic mice

The DNA of human factor Ⅸ (hFⅨ) gene vector pMCⅨm, which had been proven to be able to express in in vitro and living cells, was introduced into 586 zygotes of Kunming White Mice by positive pressure microinjection technique with manual operation. The 499 survival embryos after microinjection were then transferred into pseudopregnant recipient mice and 216 F 0 pups were born. The analysis of PCR and Southern blot hybridization showed that, of the 216, 6 (2 females and 4 males) were integrated with foreign DNA in their genomes, giving an integration frequency of 3% (6/216). Two F\-0 female transgenic mice could express hFⅨ protein in their milk and the content was over 100 ng/mL as measured with ELISA. The biological activities of hFⅨ in the milk of two F\-0 mice were 44 67% and 79 43%, respectively.     

Temperature dependent spectral response characteristic of III-V compound tandem cell

The GaInP/GaAs/Ge triple-junction tandem cells with a conversion efficiency of 27.1% were fabricated using metalorganic chemical vapor deposition (MOCVD) technique. Temperature dependence of the spectral response measurements of the GaInP/GaAs/Ge tandem cell was performed by a quantum effi-ciency system at temperatures ranging from 25℃ to 160℃. The red-shift phenomena of the absorption limit for all subcells were observed with increasing temperature, which is dued to the energy gap nar-rowing with temperature. The short-circuit current densities (J_sc)of GaInP, GaAs and Ge subcells at room temperature calculated based on the spectral response data were 12.9, 13.7 and 17 mA/cm², re-spectively. The temperature coefficient of J_sc for the tandem cell was determined to be 8.9 mA/(cm²·℃), and the corresponding temperature coefficient of the open-circuit voltage deduced from the se-ries-connected model was -6.27 mV/℃.     

Adsorption and reduction of palladium (Pd<Superscript>2+</Superscript>) by <Emphasis Type="Italic">Bacillus licheniformis</Emphasis> R08

Preliminary study on the mechanism of Pd²⁺ biosorption by resting cells of Bacillus licheniformis R08 biomass has been carried out by means of chemical kinetics and AAS, TEM, XRD and FTIR methods. The results showed that at 30℃ and pH 3.5, when dry R08 biomass powder (800 mg/L) was mixed with Pd²⁺ (100 mg/L) for 45 min, the rate constant k of biosorption of Pd²⁺ attained a maximum of 5.97×10^-2 min^-1 and the half life period of the reaction reached 12 min. The part of Pd²⁺ adsorbed by R08 biomass was reduced to elemental, cell-bound Pd⁰ at the same condition. The cell wall of R08 biomass was the primary location for accumulating Pd²⁺ , and aldoses, i. e. hydrolysate of a part of polysaccharides on the peptidoglycan layer in the acidic medium, serving as the electron donor, in situ reduced the Pd²⁺ to Pd⁰.     

Kinetic studies of the decomposition reaction of adducts of dinuclear Fe(Ⅱ)/O2

Kinetic studies of the decomposition reaction of dinuclear Fe(Ⅱ) adducts [Fe₂(N-Et-HPTB){O₂P(OPh)₂}](Cl- O₄)₂ (1) and [Fe₂(N-Et-HPTB) {O₂P(Ph)₂}] (ClO₄)₂ (2) with O₂ have been carried out at low temperature using UV-vis spectra. The decomposition reaction of Fe(Ⅱ)/O₂ adducts was first-order in the experimental conditions, and the activation parameters were obtained. ?H^¹ = 85.62 kJ·mol^-1, ?S¹ = 19.43 J·mol^-1·K^-1 for compound (1) and ?H^¹ = 97.97 kJ·mol^-1, ?S^¹ = 55.68 J·mol^-1·K^-1 for compound (2). These results are similar to those of dioxygen adducts of other metals complexes and natural enzymes such as methane mono- oxygenase (MMOH).     

Effect of Mn cluster on the formation of superoxide radicals in photoinhibition of photosystem Ⅱ

To further realize the action of superoxide radicals (O₂^{− .}) in photoinhibition of photosystem Ⅱ (PSⅡ), we employed 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) as a spin trap, associated with EPR spectroscopy, to study the effect of illumination time on O₂^{− .} formation during high light photoinhibition in PSⅡ membranes and Mn-depleted PSⅡ membranes. Results indicated that the removal of Mn cluster from PS
membranes has a strong influence on the dynamics of superoxide formation. The relative mechanism was also discussed. These novel findings may further promote the studies of the structure and function of PSⅡ and the mechanism of photoinhibition.     

Building of hFⅨ transgenic mice by spermatogenic cells

Human FⅨ expression vector pCMVⅨ was packaged by effectene^TM reagent and injected into mice seminiferous tubules with glass pipettes.The expressional frame of pCMVⅨ was examined by PCR and Southern blotting among 41 progenies.There were 2(4%) mice being integrated with hFⅨ gene into chromosomes.4.6ng/mL of hFⅨ protein was expressed in plasma of one mouse,which was tested by ELISA.We demonstrated that building of transgenic animals by spermatogonial stem cells is an efficient method.Meanwhile,it has also been proved to be an alternative choice for mammary gland bioreactor.     

Characteristics of δ 18O in precipitation over Eastern Monsoon China and the water vapor sources

Monsoon circulation is an important carrier of water vapor transport, and it impacts the precipitation of the monsoonal regions through the constraints and controls of large-scale water vapor transport and distributions as well as the water vapor balance. An overall research on stable Hydrogen and Oxygen isotopes in precipitation over Eastern Monsoon China could benefit a compre-hensive understanding of the monsoonal precipitation mechanism. Seventeen field stations of the Chinese Network of Isotopes in Precipitation (CHNIP) have been selected to collect monthly composite precipitation samples during the years 2005―2006. Components of δD and δ¹⁸O have been analyzed to achieve the spatiotemporal distributions. The established Local Meteoric Water Line δD=7.46 δ¹⁸O+0.90 based on the 274 obtained monthly samples could be treated as isotope input functions across the region, due to basically reflecting the specific regional meteorological conditions over Eastern Monsoon China. The δ-value de-pleted from coastal to inner area. In Southern China and Northeastern China there were typical periodic patterns of δ¹⁸8O. Differ-ent dominant affecting metrological factors have been raised with different regions. From south to north, the temperature effect of δ¹⁸18O enhanced, while the amount effect changed from existing at an all-year-scale in Southern China to being only remarkable during the main rainy seasons in North China and Northeastern China. Main geographical controls varied from altitude in South-ern China and North China to latitude in Northeastern China. Furthermore, δ¹⁸O had an implication of advance and retreat of the monsoon as well as rainfall belt transfer δ¹⁸O was also a tracer for the movement path of typhoon and tropical storms.     

Preparation of rAAV/hFⅨ by HSV/AAV hybrid helper virusand evaluation of its safety

Hemophilia B is a hemorrhagic disease resulting from Factor Ⅸ gene (hFⅨ) mutation as an X-linked recessive inherited trait. The incidence of this disease is 1 in 30000. Clinical treatments depend mainly upon blood transfusions or administration of prothrombin complex so that patients are at the risk of infections with the HIV and hepatitis viruses. Gene therapy offers an attractive alternative in the treatment of hemophilia B by eliminating those risks. In 1991, our lab conducted clinica…     

Formate and acetate as recorded in a mid－latitude glacier in west China

Formic (HCOOH) and acetic (CH3COOH) acids are the chief carboxylic compounds in the troposphere[1]. Although weak acids, they contribute significantly to the free acidity, ionic balance of precipitation and the chemical reactions in the atmosphere, especially in the remote areas[2—4]. General sources for formate and acetate have been identified in the past 3 decades although a satisfactory source attribution is lacking to explain atmospheric observations[5]. Possible sources include veget…     

Antibacterial activity and structure-activity relationships of Schiff bases on <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> by microcalorimetry

The growth of Staphylococcus aureus under the action of six kinds of Schiff bases(A,B,C,D,E,F) was studied by means of microcalorimetry. Growth constant k and inhibition ratio I were calculated. Also,the antibacterial activity,the action characteristics and the structure-activity relationships of the compounds on S. aureus were analyzed. Results indicate that B,F show good antibacterial activity(IC50:293.6 mg · L-1 and 307.8 mg · L-1) ,D,E show moderate antibacterial activity(IC50:966.3 mg · L-1 and 1 126.7...     

The contribution of root respiration to soil CO2 efflux in Puccinellia tenuiflora dominated community in a semi-arid meadow steppe

Rising atmospheric CO2 and temperature are altering ecosystem carbon cycling. Grasslands play an important role in regional climate change and global carbon cycle. Below-ground processes play a key role in the grassland carbon cycle because they regulate …     

Transformation character of ferrite formation by a ledge mechanism under a mixed-control model

A mixed-control model was developed to study the transformation character of ferrite formation by a ledge mechanism. A numerical two-dimensional diffusion-field model was combined to describe the evolution of the diffusion field ahead of the migrating austenite/ferrite interface. The calculation results show that the bulk diffusion-controlled model leads to a deviation from experimental results under large solute supersaturation. In the mixed-control model, solute supersaturation and a parameter Z together determine the transformation character, which is quantified by the normalized concentration of carbon in austenite at the austenite/ferrite interface. By comparing with experimental data, the pre-exponential factor of interface mobility, M₀, is estimated within the range from 0.10 to 0.60 mol·m·J-1·s-1 for the alloys with 0.11wt%-0.49wt% C at 700-740℃. For a certain Fe-C alloy, the trend of the transformation character relies on the magnitude of M 0 as the transformation temperature decreases.     

Fluorescence Determination of Artemisinin Using Hemoglobin as Catalyst and Pyronine B as Substrate

0 IntroductionMaalnadria siusb atr ompajicosr .he aAltrthe pmrisoibnlienmi (n qtihneghtraoopsiucs,QHS,Fig.1) is a sesquiterpene endoperoxide isola-ted fromArtemisia annuaL., an ancient Chineseherbal medicine usedfor treatment of fever and ma-laria.Studies of the structure and activity relation-ship have shownthat endoperoxide groupis essentialfor anti malarial activity of QHS and absence of thismoiety lead to completely loss in activity of thedrug. Many techniques have been developed to de…     

Nitric oxide produced by cumulus cells stimulates maturation of mouse oocytes

The effect of sodium nitroprusside (SNP), a donor of nitric oxide, on meiotic maturation of mouse oocytes was studied by injecting N^w-nitro-L-arginine methyl ester (L-NAME) intra-peritoneal (ip), a nitric oxide synthase inhibitor, or culturing oocytes in the medium supplemented with L-NAME or hypoxanthine (HX) to arrest the spontaneous oocyte maturation in vitro. The results showed that the inhibitory effect of L-NAME by injecting 10 mg/kg ip on extrusion of the first polar body only could be reversed by injecting 2.5 mg/kg SNP with L-NAME simultaneously (P < 0.05). Half an hour later ten mice died when given 10 mg/kg SNP ip. The treatment of some concentrations of SNP (10^–7, 10^–6, 10^–5mol/L) significantly stimulated meiotic maturation to metaphase Ⅱ stages in cumulus enclosed oocytes in the presence of HX. However, other concentrations of SNP (10^–8, 10^–4, 10^–3 mol/L) had no effect on HX-arrested oocyte meiotic maturation. The optimal concentration of SNP on CEOs had no effect on DOs. The dose of 10^–3 mol/LL-NAME demonstrated a significant suppression in formation of PB₁, but not in GVBD. This inhibition was reversed by the addition of SNP. These results indicated that the physiological levels of NO produced by cumulus cells could stimulate meiotic maturation of mouse oocytes both in vivo and in vitro.     

Trace determination of zirconium using anodic adsorptive voltammetry at a carbon paste electrode modified with multi-walled carbon nanotubes

A carbon paste electrode modified with multi-walled carbon nanotubes （MWCNT） was prepared and the determination of ultra trace amount of zirconium based on the anodic adsorptive voltammetry of the zirconium-calcium-alizarin red S mix-polynuclear complex is described in this paper for the first time. The results showed that the sensitivity and the selectivity of the method are excellent. The second derivative linear scan voltammograms of the complex were recorded by polarographic analyzer from 200 to 1200 mV （vs. SCE） and it was found that the complex can be adsorbed on the surface of the electrode, yielding a peak at about 840 mV, corresponding to the oxidation of ARS in the complex. The peak current increases linearly with Zr （IV） concentration in the range of 6.0×10^-12--6.0×10^-11 mol. L^-1 （accumulation time 120 s）, 6.0×10^-11--2.0×10^-9 mol. L-1 （accumulation time 90 s） and 2.0×10^-9--1.0×10^-7 mol. L^-1 （accumulation time 60 s） and the detection limit （S/N = 3） is 2.0×10^-12 mol. L^-1 （accumulation time 180 s）. The procedure has been successfully applied to the determination of zirconium in the ore samples.