In the present study, the regulation of Vitreoscilla hemoglobin (VHb) on astragaloside IV biosynthesis was investigated. An intermediate expression vector consisting of the CaMV35S promoter fused to the vgb and nopaline synthase terminator was transferred into Astragalus membranaceus via Agrobacterium rhizogenes. The transgenic hairy roots were confirmed by PCR amplification and Southern blot hybridization. The expression of vgb in transgenic hairy roots was confirmed by RT-PCR. After 15 days cultivation, the dry weight and growth rate of transgenic hairy roots were higher than that of the non-transgenic hairy root. ELSD-HPLC analysis showed that astragaloside IV content of transgenic hairy roots was 5 to 6 times of non-transgenic hairy root control and 10 to 12 times of Radix Astragali from Shanxi Province. These results suggested that the expression of vgb promoted the growth of transgenic hairy roots, and increased the content of astragaloside IV.

Objective To evaluate systematically the effect of ventilator circuit changes on incidence of ventilator-associated pneumonia(VAP).Methods Literatures about the effect of ventilator circuit changes on occurrence of VAP were searched from PubMed,Cochrane Library,CNKI,VIP,and Wanfang database,incidence of VAP in pa-tients who received circuit changes at different intervals was compared,odds ratio (OR)and 95%CI were as effect index of the study,RevMan 5.2 software was used for data analysis.Results A total of 12 literatures were included,including 18003 patients using ventilators.According to the interval of ventilator circuit changes,patients were divided into 1-day group (n=191),2-day group(n=8932),3-day group(n=228),and 7-day group(n=8652).Meta analysis showed that circuit changed every 7 days had a lower risk of VAP than that changed once ev-ery day (OR,3.72[95%CI,1.50,9.23]),there was no significant difference in risk of VAP between 7-day group and 2-day group (OR,1.17[95%CI,1.00,1.37])as well as 7-day group and 3-day group(OR,0.77 [95%CI, 0.49,1 .23]).Conclusion Patients who received circuit changes every 7 days have a lower risk of developing VAP than those who received circuit changes once per day,so change the ventilator circuit in patients with mechanical ventilation every 7 days is more reasonable.

Objective: To compare the immunomodulatory effects of spore polysaccharides (Gl-SP) and broken spore polysaccharides (Gl-BSP) isolated from Ganoderma lucidum(Leyss et Fr.) Karst. on murine splenic lymphocytes and peritoneal macrophages in vitro. Methods: Mixed lymphocyte culture reaction (MLR), lymphocyte proliferation in the presence or absence of mitogen, and the cytotoxic activity of splenic natural killer (NK) cells were detected with MTT assay in vitro. The percentage of phagocytosis of neutral red (NR) by mouse peritoneal macrophages was detected by colorimetric assay. Splenic T-lymphocyte subpopulations were measured with flow cytometry(FCM). IL-2, IFN-? and TNF-? in the culture supernatants were detected by ELISA and biological assay. Nitric oxide (NO) production was examined by Griess reaction. Results: At the concentration range of 0.2-12.8 mg/L, Gl-SP and Gl-BSP were shown to increase lymphocyte proliferation in the presence or absence of mitogen, enhance NK cytotoxic activity, augment the production of TNF-? and NO in Gl-SP-or Gl-BSP-activated macrophages, as well the percentage of phagocytosis of NR by macrophages in vitro. Both Gl-SP and Gl-BSP could promote MLR, however, at the dose of 12.8mg/L, Gl-BSP showed higher activity than Gl-SP in the proliferation of lymphocytes. These two kinds of polysaccharide could significantly increase the secretion of IL-2 and IFN-? in double-way MLR at the concentrations of 0.2-12.8 mg/L, but Gl-BSP had stronger effects than Gl-SP at the same concentrations. Both Gl-SP and Gl-BSP could increase the ratio of T-lymphocyte subpopulations in double-way MLR. At the concentrations of 0.2-12.8 mg/L or 3.2-12.8 mg/L, Gl-BSP demonstrated more significant activity in increasing the percentage of the CD4+ or CD8+ subset than Gl-SP. At the concentrations of 0.2-0.8 mg/L, the ratio of the CD4+ and CD8+ subset in the Gl-BSP treated group was higher than that of the Gl-SP treated group. Conclusion:Gl-SP and Gl-BSP have sim-ilar immunomodulatory effects in vitro, as though the immunomodulatory effects of Gl-BSP are stronger than that of Gl-SP.

Objective To explore whether the neural stem cells(NSCs) can act directly as a gene target cell which can be infected by the recombinant retrovirus and express the products of exogenous genes after infection. Methods The NSCs were cultured with supernatant containing the recombinant retroviruses with the genes of NGF or GDNF for two days.After screened with G418,the infected NSC were expanded at the present of bFGF in culture.The PC12 cells and the neurons of ventral midbrain of rat were cultured by the medium from the infected NSC,which were called as GDNF\|containing conditioned medium NGF or GDNF\|containing conditioned medium the morphological changes of the dopamine neurons of the ventral midbrain and expression of exogenous genes of the infected NSCs were detected by immunohistochemistry staining. Results It was estimated that about fifty percent of NSCs via retrovirus\|mediated NGF or GDNF gene transduction were G418\|resistant.These infected NSCs began to differentiate.Long and radical processes reached out from the sphere of proliferation and the cells migrated towards outside along the processes.The NSC infected with gene of NGF showed an astroid\|shape with larger body and processes.The NSC infected with gene of GDNF showed a shuttle\|shape with a smaller body and long processes.The PC12 cells increased in the NGF\|containing conditioned medium and stretched out long neurites.The dopamine neuron of the ventral midbrain which were immunoreactive for TH also showed a larger body and longer processes in the GDNF\|containing conditioned medium.Most of G418\|resistant NSCs were immunoreactive for NGF or GDNF. Conclusion NSC can act directly as a gene target cell which not only be infected by the recombinant retrovirus,but also express and secrete the products of exogenous genes.

Objective To study the structural features and anticoagulant activity of the polysaccharide isolated from Aeodes orbitosa.Methods Sugar composition analysis,methylation analysis and IR were used to characterize the structural features.The anticoagulant activity of the polysaccharide was evaluated by cutting tail and capillary methods.Results The polysaccharide was composed of 3,6-anhydrous-galactose,6-methyl-galactose,2-methyl-galactose,galactose,xylose and glucose in the molar ratios of 6.4∶0.9∶5.8∶1.6∶84.1∶1.2.The content of Sulfate group was 37.5%. The main linkage model of the polysaccharide were 1,3 and 1,4 linkages,branch point located at O-2 and 6 of galactose residues.Sulfate group located at 2 and 6 position of 1,4 linked galactose residue and 2 position of 1,3 linked galactose residue.The polysaccharide showed significant activities to extend bleeding time and coagulation time in mice.Conclusion The polysaccharide from Aeodes orbitosa was a sulfate galactan with strong anticoagulant activity.

Estrogen participates in many life activities through combination with estrogen receptor alpha (ERalpha) or estrogen receptor beta (ERbeta) in the body. In order to establish an in vitro estrogen-like compound screening model, the coding region of human ERalpha and ERbeta was separately constructed into pET32-ERalpha and pET43-ERbeta prokaryotic expression vector and water-soluble recombinant ERalpha and ERbeta proteins were expressed in Escherichia coli strain BL21. Western blotting revealed that both recombinant proteins have estrogen receptor binding sites. The proteins were purified using S-Tag affinity Purification Kit and digested with enterokinase to get the ERalpha and ERbeta proteins. About 0.90 mg of ERalpha and 0.65 mg of ERbeta were obtained at the concentration of 0.181 and 0.131 mg x mL(-1), respectively.

Danshen (Salvia miltiorrhiza Bunge) hairy roots were obtained by infecting Danshen leaves with Agrobacterium rhizogenes 9402. Besides rosmarinic acid (RA) and salvianolic acid B (SAB), the hairy root could also produce salvianolic acid K (SAK), salvianolic acid L, ethyl salvianolic acid B (ESAB), methyl salvianolic acid B (MSAB), and a compound with a molecular weight of 538 (compound 538) identified by using LC-MS. Effects of methyl jasmonate (MeJA) and yeast elicitor (YE) on the accumulation of these compounds had been investigated. MeJA increased the accumulation of SAB, RA, SAK, and compound 538 from 4.21%, 2.48%, 0.29%, and 0.01% of dry weight to 7.11%, 3.38%, 0.68%, and 0.04%, respectively. YE stimulated the biosynthesis of RA from 2.83% to 5.71%, but depressed the synthesis of SAB, SAK and compound 538. It was indicated in all the results that these Danshen hairy roots could be used as alternative resources to produce salvianolic acids. Analysis of the content variation of these compounds after elicitation suggested that SAK and compound 538 might be the intermediates in the biosynthesis from RA to SAB in Danshen hairy roots.

Epicatechin content in Uncaria macrophylla Wall. was determined by RP-HPLC. As aresult, the epicatechin contents were 0.38％ in the hook and stem, and 0. 820％ in the leaf. Thus it seemedto be more worthwhile to produce epicatechin from the leaf rather than from the hook and stem.

Metabolic profile of bile acids was used to evaluate hepatotoxicity of mice caused by ethanol extraction of Dioscorea bulbifera L. (ethanol extraction, ET) and diosbulbin B (DB), separately. Ultra-performance liquid chromatography coupled with quadrupole mass spectrometry (UPLC-MS) was applied to determine the contents of all kinds of endogenous bile acids including free bile acids, taurine conjugates and glycine conjugates. Obvious liver injuries could be observed in mice after administrated with ET and DB. Based on the analysis using principle components analysis (PCA), toxic groups could be distinguished from their control groups, which suggested that the variance of the contents of bile acids could evaluate hepatotoxicity caused by ET and DB. Meanwhile, ET and DB toxic groups were classified in the same trends comparing to control groups in the loading plot, and difference between the two toxic groups could also be observed. DB proved to be one of the toxic components in Dioscorea bulbifera L. Bile acids of tauroursodeoxycholic acid (TUDCA), taurochenodeoxycholic acid (TCDCA), taurocholic acid (TCA), taurodeoxycholic acid (TDCA), cholic acid (CA) and others proved to be important corresponds to ET and DB induced liver injury according to analysis of partial least square-discriminant analysis (PLS-DA) and the statistical analysis showed that there were significant differences between the control groups and toxic groups (P < 0.01). Furthermore, good correlation could be revealed between the foregoing bile acids and ALT, AST. It indicated that taurine conjugated bile acids as TUDCA, TCDCA, TCA and TDCA along with CA could be considered as sensitive biomarkers of ET and DB induced liver injury. This work can provide the base for the further research on the evaluation and mechanism of hepatotoxicity caused by Dioscorea bulbifera L.