Expression of Porcine Reproductive and Respiratory Syndrome Virus ORF7 Gene and Purification and Immunological Activity Analysis of the Recombinant Protein

[Objective] The aim of this study was to realize efficient expression of the porcine reproductive and respiratory syndrome virus （PRRSV） ORF7 gene in genetic engineering bacteria and analYze the immunological activity of the recombinant protein after purification. [ Method] The constructed recombinant expression vector pET-ORF7 was transformed into Escherichia co1BL21 （DE3） and induced by IPTG under the optimal condition. After analysis of SDS-PAGE and Western Blot, the expression products were purified by Ni-NTA His · Bind Resin chrom- atographic column under denaturing condition and renatured by gradient dialysis. Subsequently, the immunological activity of the renatured recombinant protein was detected by Westem Blot and indirect ELISA. [ Result] The recombinant plasmid pET-ORF7 expressed in E. coli successfully, and the fusion protein was in the form of inclusion body. By SDS-PAGE detection, the molecular weight of the expression protein was approximate 33 kD, according with the expectation. Analysis by Bandscan software showed that the expressed fusion protein was about 50% of total bacterial protein of BL21 （DE3）. Wastem Blot and indirect ELISA detection showed that the renatured protein could react with PRRSV positive serum specifically, indicating its good immunological activity. [ Conclusion] This study lays a foundation for the preparation of PRRSV monoclonal antibody and diagnostic kit.

Synthesis and antitumor activity of 7-azaindirubin

Twenty 7-azaindirubin derivatives were designed and synthesized. Their antitumor activities were evaluated in vitro against DU145 cell line. The pharmacological results showed that most of the prepared compounds displayed the excellent activity. Compound 18 exhibited the most potent antitumor activity among the tested compounds.

Potential bone-inducing activity in vitro of recombinant human bone morphogenetic protein-7 from a CHO expression system

Objective: To express the recombinant human bone morphogenetic protein-7(rhBMP-7) in Chinese hamster ovary(CHO) cells, and to establish the in vitro biological activity assay of rhBMP-7.Methods: Human BMP-7 cDNA was subcloned into p114 mammalian expression vector and transfected to CHO cells by using the Lipofectamine 2000 transfection method. CHO cell supernatants were harvested and analyzed to identify the molecule mass of secreted rhBMP-7 and examine its biological activity in vitro to stimulate the synthesis of alkaline phophatase(ALP), a characteristic of osteoblast phenotypes. Results: rhBMP-7 was produced stably in CHO cells, as a processed mature disulfide-linked homodimer, with an apparent molecular mass of 36 000. Examination of the rhBMP-7 biological activity showed that rhBMP-7 specifically stimulated the production of ALP(4-fold increase at 100 ng of rhBMP-7/ml). Conclusion: The rhBMP-7 from CHO expression system has significant biological activity in induction of osteoblast phenotype, which demonstrates rhBMP-7 has the potential bone regeneration activity.

Sawtooth Activity in Ohmically Heated Plasma on HT-7 Tokamak

Sawtooth activity on HT-7 tokamak has been investigated experimentally mainly by using soft x-ray diode array and magnetic probes. Their behaviors and occurrences are correlated closely to the discharge conditions: the electron density Ne, the electron temperature Te, the safety factor qa on plasma boundary and wall condition etc. When central line-averaged electron density Ne(0) is over 2.0×1013cm-3 , major sawtooth activity emerges with a period of up to 6.5 ms and a fluctuation amplitude of up to 2-30 % of SXR radiation signal. In some cases such as the safety factor between 4.2-4.7 and Zeff=3.0-6.0, a monster sawtooth activity often emerges without apparent deterioration of plasma confinement and without major disruption. During these events, abundant MHD phenomena are observed including partial sawtooth oscillations. In this paper, the observed sawtooth behaviors and their dependence on the and their dependence density Ne and wall condition in ohmically heated plasma are introduced, the results are discussed and presented.

The Study of Natural Solution Protective Activity on H7N9 Virus

Background: One of potentially dangerous problems for a human organism is the new strain of a virus of bird flu-A/H7N9. As it is regular mutation of bird flu virus, it obvious, that of antibacterial preparations is not efficient. Efficiency decreases when the number of agents with multiple stability to antimicrobic remedy vastly increases, the part of associate infections enlarges, and aggression of opportunistic pathogenic flora rises. This reduces the role of the preparations in prevention of epidemics. Therefore, the optimization of only etiotropic therapies does not fully solve the problem. In this connection natural preparations seem extremely promising which strengthen the functional condition of immune system and, thereby, activate protective forces of macroorganism. Objectives: One of such preparations is BAE Synergy Liquid, a natural mineral water which was underwent subtle energetic changes at the natural energetic deposit. Design: An estimation of protective efficiency of naturally modified mineral BAE SL water was performed on white outbred mice-males in models of H7N9 virus. The animals were monitored during 16 days after infection, and survived and fallen mice were counted daily. Results: The results revealed significant effect of the investigated preparation as possible prophylactic care and medical remedy to the mentioned virus. This means that one can be considered as potential effective remedy for human. Conclusions: As significant effect of the immune system resistance was revealed, the experimental model with studied naturally modified mineral water is potentially generalizable.

In Vitro Assessment of Probiotic Potential of Lactobacilus acidophilus and Antagonistic Activity Against Escherichia coli O157:H7

The antagonistic activity of Lactobacillus acidophilus KLDS1.0901, KLDS1.0902, KLDSI.1003 and NCFM against Escherichia coli O157 ： H7 were investigated in this study. The culture supematants of all the L. acidophilus stains showed high bacteriostatic activities against E. coli O 157 ： H7 and the bacteriostatic substances of their Cell-Free Supernatants （CFS） were preliminarily determined from organic acids. The bacteriostatic activity from CFS or viable L. acidophilus against E. coli O157 ： H7 was also assessed by using co-incubation methods, CFS had high bactericidal activity against E. coli O157 ： H7, no viable E. coli O157 ： H7 was detected when 5×10^7 CfU ofE. coli O157 ： H7 was added to 5 mL of CFS and incubated at 37℃ for 2 h. However, L. acidophilus themselves had no bacteriostatic activity after directly contacted with E. coli O157 ： H7. The inhibition E. coli O157 ： H7 adhesion and colonization of L. acidophilus were also investigated based on competition, exclusion and displacement assays. L. acidophilus KLDS1.0901, KLDSI.1003 and NCFM strains were effective to displace E. coli O157 ： H7 from a Caco-2 cell layer in competition and exclusion assays. However, in displacement assay, all of the strains showed no significant antagonistic activities. Meanwhile, the probiotic potential of L. acidophilus strains was investigated based on adhesion assay to Caco-2 cells and anti- inflammatory effects by IL-8 produced in Caco-2 cells. The adhesion ability and anti-inflammatory effects of L. acidophilus strains showed a strain-dependent manner. In general, L. acidophilus KLDS 1.0901 and NCFM showed better probiotic potential than KLDS1.0902 and KLDSI.1003. Thus, the use ofL. acidophilus KLDS1.0901 and NCFM to prevent or treat of diseases associated induced E. coli O157 ： H7 in vivo was suggested.

Cytotoxic Activity of <i>Thelesperma megapotamicum</i>Organic Fractions against MCF-7 Human Breast Cancer Cell Line

Thelesperma megapotamicum (Asteraceae) is commonly used in Argentine to treat various diseases (renal, digestive affections, and as anaesthesia). The present study showed the mechanisms involved “in vitro” cytotoxicity of T. megapotamicum Fractions. Five Fractions (F1 - F5) were separated by column chromatography (Silica gel) using hexane:diethyl ether as eluents. Viability was evaluated in Human breast carcinoma cell line (MCF-7) by staining with crystal violet. With respect to F1 Fraction treatment, the cell survival was 49.14% ± 8.87%, while the F2 and F3 ones exhibited a strong reduction of cell viability to only 26.35% ± 1.63% and 23.3%1 ± 0.53% of the control cell at 50 μg/ml, respectively. Apoptotic effect of these Fractions was detected using FITC-labeled Annexin V and propidium iodide binding assays and was confirmed by a higher proportion of apoptotic cells due to F2 and F3 treatments. T. megapotamicum active Fractions could facilitate the tumoral cells death by decreasing the activity of the enzyme Gamma-glutamyltranspeptidase and causing alteration in cell membrane sialoglycoconjugates and others involved anticancer mechanisms including apoptosis.

Monster Sawtooth Activity in Ohmically Heated HT-7 Plasma

Experimental properties of monster sawtooth activities in ohmic HT-7 plasma are presented in this paper. The monster sawtooth activities belong to global fluctuations and are characterized with a series of large core collapses on SXR intensity traces with a long period, a large amplitude fluctuation and a large inversion radius. However, they emerge without apparent deterioration of plasma confinement and without major plasma disruption. During the events, long partial sawtooth collapses and abundant MHD phenomena are also observed.

Studies on Semisynthesis and Anibacterial Activity of 7 β-(5-Methyl-1-Aryl-1H-1,2,3-Triazoly-4-Carboxamido) Cephalosporins

Nine new derivatives of 7 β - (5- methyl- 1- aryl- 1H-1, 2, 3- triazoly- 4- carboxamido) cephalosporins were synthesized by acylction of 7 β -amino group of 7-ACA, 7-ADCA and 7-ACT with 5 -methsyl- 1 -aryl- 1 H- 1,2,3-triazoly-4-formyl chloride. The structure of the compounds were confirmed by elementray analysis IR, HNMR and FAB-MS. Some of them showed significant antibacterial activity

Antioxidant Activity of the Natural Flavonoid 7-Hydroxy-5,6,4’-trimethoxyflavone Isolated from the Leaves of Lippia rugosa A. Chev

Phytochemical studies and antioxidant activities were carried out on n-hexane, ethyl acetate, ethanol and methanol extracts of the leaves of Lippia rugosa A. Chev (Verbenaceae), a medicinal plant used traditionally in the Cameroonian savannah’s region to protect foodstuffs and to cure degenerative diseases. Results indicated that theses extracts contain terpenoids, phenolic and flavonoid compounds. Except the n-hexane extract, all of the obtained extracts exhibit antioxidant activities with the ethanol extract being the most effective with an inhibition percentage of 85.668% ± 1.233% and an inhibition concentration (IC50) of 58 μg/ml (R2 = 0.987, P < 0.01) at a concentration of 100 mg/ml. Chromatographic separation on silica gel of the ethanol extract led to the isolation of a pure organic compound which was characterized as 7-hydroxy-5,6,4'-trimethox- yflavone by extensive 1D and 2D NMR spectroscopy, a flavonoid exhibiting antioxidative activity with an inhibitory percentage of 25.506% ± 0.205% and inhibition concentration (IC50) of 221 μg/ml (R2 = 0.966, P < 0.01). This is the first time that 7-hydroxy-5,6,4'-trimethoxyflavone is being isolated from L. rugosa and its antioxidant activity evaluated.

Synthesis, Crystal Structure and Antitumor Activity of (E)-1-(7-Methoxy-2,2-dimethyl-2,3-dihydrobenzofuran-5- yl)-3-(2-methoxyphenyl)-2-(1H-1,2,4-triazol-1-yl)propenol

The title compound, （E）-1-（7-methoxy-2,2-dimethyl-2,3-dihydrobenzofuran-5-yl）- 3-（2-methoxyphenyl）-2-（1H-1,2,4-triazol-1-yl）propenol （3a）, was synthesized by the Aldol con- densation reaction of 1-（7-methoxy-2,2-dimethyl-2,3-dihydrobenzofuran-5-yl）-2-（1,2,4-triazol- 1-yl）ethanone with 2-methoxybenzaldehyde and then reduced with NaBH4, and its crystal structure was determined by single-crystal X-ray diffraction： monoclinic system, space group P21 with a = 6.2002（3）, b = 12.8452（7）, c = 13.2257（7） ？, Z = 2, V = 1031.23（9） ？3, Mr = 407.46, Dc = 1.312 Mg/m3, S = 1.054, μ = 0.091 mm-1, F（000） = 432, the final R = 0.0353 and wR = 0.0769 for 3161 observed reflections （I 〉 2σ（I））. X-ray analysis displays that the title compound adopts an E configuration for the C（7）=C（8） double bond and S configuration for the chirality center with the specific rotation of –63.75°. Furthermore, the stability of the crystal was maintained through the intermolecular hydrogen bond O（1）–H？？？N（3）. The antitumor assay exhibits that the title compound 3a （E configuration） has a good antitumor activity against the Hela cell line with the IC50 value of 36.9 μM, which is better than that of 3b （Z configuration）.

Analysis of Seismic Activity Characteristics before the Yutian M_S7.3 Earthquake,Xinjiang on February 12,2014

This paper introduces the geological structure background around the 2014 Yutian Ms7. 3 earthquake area, investigates and analyzes the regime of small earthquake activity and the characteristics of regional seismicity pattern in Xinjiang before the earthquake, and compares the characteristics of the regional seismic activity with the 2008 Yutian Ms7.3 earthquake. The results show： ① 2 ~ 3 years before the 2014 Yutian Ms7. 3 earthquake, Xinjiang was in a seismic active state with strong earthquake occurring successively, and before the 2008 Ms 7. 3 earthquake, Xinjiang was in the quiet state of moderate-small earthquakes with M3. 0 ~ 4. 0. ② Before this Yutian Ms7. 3 earthquake, the regional seismic activity showed a short-term anomaly feature, that is, seismicity of M ≥ 5. 0 earthquakes significantly increased on the Altun seismic zone and in the source area three years before the Ms7.3 earthquake, while a five year long quiescence of seismicity of M ≥4. 0 earthquakes appeared on the east of the source area in a range of about 440kin. Six months before this M7. 3 earthquake, there existed seismic gap of M3. 0 ~ 4. 0 earthquakes and near-conjugate seismic belt magnitude 3. 0 and 4. 0 in the source area. ③ The state of strong earthquake activity and the seismicity pattern of small earthquakes before this Yutian Ms7. 3 earthquake were significantly different to that before the 2008 Yutian Ms7. 3 earthquake, and this may be related to the different seismogenic environments of the two Ms7. 3 earthquakes.

Construction of Cox7a2 fluorescent vector and its effect on cytochrome C oxidase activity in mouse Sertoli cell line TM4

Objective To construct Cox7a2 fluorescent vector and study its effect on cytochrome C oxidase ( COX) activity in mouse Sertoli cell line TM4. Methods The coding region of CoxTa2 was amplified from mouse Sertoli cell line TM4 by RT-PCR. PCR product was

生防链霉菌17-7菌株发酵配方优化及发酵滤液稳定性研究

为了提升链霉菌17-7菌株的抑菌活性,合理利用菌株的代谢产物,本研究以橡胶树褐根病菌为靶标菌,采用正交试验和单因子试验相结合,在原培养基的基础上对链霉菌17-7菌株的发酵培养基配方进一步优化,获得该菌株最适宜产活性物质的配方为:大豆粉25 g/L、玉米粉10 g/L、葡萄糖10 g/L、酵母粉2 g/L、K_(2)HPO_(4)1 g/L、NaCl 0.5 g/L、CaCO_(3)0.5 g/L。发酵滤液稀释500倍后对橡胶树褐根病菌的相对抑菌活性为51.37%,较优化前提高了17.66%,并且具有广谱的抑菌活性。在此基础上测定了温度、紫外线辐射、蛋白酶、酸碱度和贮存条件对17-7菌株发酵滤液抗真菌物质稳定性的影响。结果表明:17-7菌株发酵滤液对高温不敏感;紫外线照射1 h后发酵滤液的抑菌活性开始显著下降,长时间紫外照射并不会丧失抑菌活性;17-7菌株发酵滤液还具有较好的酸碱耐受性,pH为3~10时其发酵滤液均能发挥较好的抑菌活性;并且对胰蛋白酶和胃蛋白酶不敏感;菌株发酵滤液在低温(4℃)和室温(25℃)条件下贮存120 d仍能保持较强生物活性。对17-7菌株无菌发酵滤液稳定性和贮存条件的研究可为后续该菌株的开发利用提供理论依据和数据支持。

Resveratrol elongates the lifespan and improves antioxidant activity in the silkworm Bombyx mori

A number of research has shown that the plant polyphenol resveratrol,one of the most prominent small molecules,has beneficial protective effects in multiple organisms,including worms,flies,and killifish.To understand the effects of resveratrol on lifespan,we evaluated its effects in the silkworm Bombyx mori.In this study,we found that lifespan was significantly prolonged in both female and male silkworms treated with resveratrol.Silkworm larval weight was significantly increased from day 3 of the 5th larval instar(L5D3)to day 7 of the 5th larval instar(L5D7).However,the weight of the pupa,cocoon,and total cocoon was not significantly different in female silkworms with resveratrol treatment than that in controls.Meanwhile,resveratrol significantly improved the thermotolerance of the silkworms,which enhanced their survival rate.Moreover,antioxidant activity was increased by resveratrol in both female and male silkworms.Furthermore,an antioxidant-related signalling pathway,SIRT7-FoxO-GST,was activated in silkworms with resveratrol treatment.Collectively,these results help us to understand the molecular pathways underlying resveratrol induced pro-longevity effects and indicate that silkworm is a promising animal model for evaluating the effects of lifespan-extending drugs.

Regeneration of Acid Orange 7 Exhausted Granular Activated Carbon Using Pulsed Discharge Plasmas

In this paper, a pulsed discharge plasma （PDP） system with a multi-needle-to-plate electrodes geometry was set up to investigate the regeneration of acid orange 7 （AO7） exhausted granular activated carbon （GAC）. Regeneration of GAC was studied under different conditions of peak pulse discharge voltage and water pH, as well as the modification effect of GAC by the pulse discharge process, to figure out the regeneration efficiency and the change of the GAC structure by the PDP treatment. The obtained results showed that there was an appropriate peak pulse voltage and an optimal initial pH value of the solution for GAC regeneration. Analyses of scanning electron microscope （SEM）, Boehm titration, Brunauer-Emmett-Teller （BET）, Horvath-Kawazoe （HK）, and X-ray Diffraction （XRD） showed that there were more mesopore and macropore in the regenerated GAC and the structure turned smoother with the increase of discharge voltage; the amount of acidic functional groups on the GAC surface increased while the amount of basic functional groups decreased after the regeneration process. From the result of the XRD analysis, there were no new substances produced on the GAC after PDP treatment.

Syntheses and Vasodilatory Activities of New Pyrazolo[4,3-d]pyrimidin-7-ones

A series of novel pyrazolo [ 4,3-d ] pyrimidin-7-ones was designed and synthesized in order to find new potent phosphodiesterase 5 inhibitors（PDE5）. The structures of all the compounds were confirmed by IR, ^1H NMR and MS; their in vitro vasodilatory activities were tested on isolated rabbit aortic spiral strips. Among them, compound X3 displays a significant vasodilatory activity, which is more potent than verapamil.

Tetramethylpyrazine analogue T-006 promotes clearance of alpha-synuclein by enhancing proteasome activity in Parkinson disease models

OBJECTIVE To investigate the effects of T-006(tetramethylpyrazine derivative)in promotingα-Synuclein(α-Syn)degradation and evaluated the neuroprotective effects in cellular and animalα-Syn model of Parkinson disease(PD).METHODS The inducible PC12 cells overexpressingα-syn and the homozygous transgenic(Tg)mice expressing A53T humanα-syn were used to evaluate the neuroprotective effects of T-006.For cellular study,MTT,Western blotting,proteasomal activity assay and qRT-PCR were applied to analyze the pharmacological effects and underlying mecha⁃nisms.The gene knock-down and overexpression approaches were used to dissect the molecular signaling pathways.For animal study,ten-month-old homozygousα-Syn Tg mice were treated with T-006(3 mg·kg-1)daily by gavage for four weeks.The Western blotting,immunohistochemistry and behavioral tests were applied to determine the neuropatho⁃logical changes.RESULTS T-006 promoted the degradation of WT and mutantα-Syn in PC12α-Syn inducible cells via an ubiquitin-proteasome system(UPS)dependent and autophagy-lysosome pathway independent manner.The mecha⁃nism of action involved the upregulation of 20S proteasome subunit LMP7 expression,which leads to activation of the chymotrypsin-like proteasomal activity for protein degradation.Mechanistically,we demonstrated that T-006 activated PKA/Akt/mTOR pathway upstream for LMP 7 up-regulation and UPS activation.Finally,we illustrated that T-006 promoted both Triton-soluble and-insoluble forms ofα-syn and protected againstα-Syn-induced neurotoxicity in A53Tα-Syn Tg mice.CONCLUSION T-006 is a potent UPS activator which promotes the degradation of pathogenic proteinα-Syn in cellular and animal PD models.Our study thus high-lights the therapeutic potential of small molecular UPS activator like T-006 in the treatment of PD and related conditions.

Synthesis,Crystal Structure and Cytotoxic Activities of 1-(Prop-2-yn-1-yl)-7,8-dihydro-1H-benzo[d][1,3]-thiazine-2,5(4H,6H)-dione Derivatives

The important synthetic precursor（Ⅲ）, 1-（prop-2-yn-1-yl）-7,8-dihydro-1Hbenzo[d][1,3]thiazine-2,5（4H,6H）-dione（C（11）H（11）NO2S）, was prepared through a three-component reaction, which was further transferred into cytotoxic triazoles by alkylation and ＂click＂ synthesis in satisfactory yields of 87%^95%. Their structures were characterized by IR, H-RESI-MS and NMR analysis. Meanwhile, the crystal of Ⅲ was obtained and determined by X-ray single-crystal diffraction. Crystal data： orthorhombic system, space group P212121, a = 5.189（4）, b = 8.661（6）, c = 23.498（17） A, V = 1056.2（13） A^3, Z = 4, F（000） = 464, Dc = 1.392 g/cm^3, μ =0.284 mm^-1, R = 0.0637 and wR = 0.1668 for 8182 independent reflections（R（int） = 0.1580） and 2166 observed ones（I 〉 2σ（I））.

DNA Binding and Antitumor Activity of a New Fluores cent ProbeEu(Ⅲ)-Complex Containing 4-Methyl-Coumarin Moiety

A new europium(Ⅲ)complex containing(4-Methyl-2-oxo-2 H-chromen-7-yloxy)-acetic acid moiety(CMMC)was synthesized,characterized,and confirmed as antitumor agent and fluorescent probe.The spectroscopic measurements of Eu(Ⅲ)in the presence of CMMC were obtained in different solvents.The results show that the strongest Eu(Ⅲ)emission bands were monitored in iso-propyl alcohol while the weakest Eu(Ⅲ)emission band was observed in acetonitrile.The interaction of Eu(Ⅲ)-(CMMC)2 complex with DNA was monitored using absorption and emission techniques.From fluorescence titration measurements,the binding constants of DNA with Eu(Ⅲ)-(CMMC)_2 complex were found to be 1.04×10~5 L·mol^(-1) in Tris-HCl and 1.17×10~7 L·mol^(-1) in DMSO-Tris-HCl buffer(9∶1 V/V).Hypochromism was observed from the absorption titration experiment which indicates the intercalation of Eu(Ⅲ)-complex between the base pair of DNA.This result further confirmed by fluorescent Ethidium bromide displacement assay.The fluorescence calibration curve was used for the determination of DNA with LOD of 1.2 ng in DMSO-Tris-HCl buffer(9∶1 V/V)and 5 ng in Tris-HCl buffer.The preliminary antitumor investigation shows promising cytotoxicity against MDA-MB-231,MCF-7(mammary cancer),and PC-3(prostate carcinoma)cell lines with IC50 values of 40.63,25.42 and 30.25μmol·L^(-1),respectively.