Objective To isolate and identify the components from the stem barks of Morus yunnanensis.Methods The compounds were isolated and purified by silic gel column,Sephadex LH-20,and RP-18 chromatography.Their chemical structures were elucidated on the basis of physicochemical properties and spectral data.Results Eleven compounds were isolated and identified as:mulberroside C(1),oxyresveratrol(2),2',4',7-trihydroxy-(2S)-flavone(3),norartocarpetin(4),moracin P(5),betulinic acid(6),sitosteryl 3?-glucoside 6'-O-palmitate(7),lupeol(8),betulinic acid(9),?-daucosterol(10),and ?-sitosterol(11).Conclusion All the compounds are isolated from the plant for the first time,and compound 9 shows cytotoxic activities.

Objective To investigate the effect and clinical valve of target-controlled infusion of remifentanil for conscious intubation of patients under fibrobronchoscopy.Methods Forty patients were randomly divided into two groups(F and R),each group contained 20 cases.All of the patients were intubated on regional anaesthesia,as the first step,then the patients in F group were given fentanyl 2?g/kg by vein.In remifentanil target-controlled infusion group(R group),the patients were kept infusing remifentanil 2ng/ml.Results (1)Haemodynamics:in R group,SBP and HR kept steadily;In F group,while the two indices increased significantly.(2)Static pulmonary function:in R group,RR,VT deceased remarkablely.There were no difference between two groups in SpO_2 and P_ ET CO_2.(3)Tracheal intubating condition and Ramsay score in R group were better than in F group;Cardiovascular response in R group was lower.Conclusion Target-controlled infusion of remifentanil for conscious intubation can achieve excellent result,with less cardiovascular response.

Objective To investigate the role of Huang-qi in balance of TH1/TH2 in asthma on dendritic cells level. Methods DCs from human peripheral blood mononuclear cells(PBMC) were induced by rhGM-CSF and rhIL-4, and then were identified. The level of IL-12 and IL-10 produced by DCs were de-tected by ELISA assay. After autoreactive T cells, mRNA of T-bet and GATA-3 was measured by RT-PCR. Simultaneously, IL-4 and IFN-γ were determined by flow cytometer. Results After 7 days culture, IL-12 was significantly decreased in asthma group compared to control group (P ＜ 0.01), whereas IL-10 on the opposite. At the 7th day of co-culture with T cells derived from floating cells, the IFN-γ/and T-bet mRNA level in asthma group were significantly decreased than that in control group, whereas IL-4, GATA-3 mRNA level, the ratio of IL-4/IFN-γ and GATA-3/T-bet were apparently increased in asthma group than that in control group(P＜0.01). After Huang-qi treatment, the IFN-γ/and T-bet mRNA level were significantly in-creased, whereas the ratio of IL-4/IFN-γ and GATA-3/T-bet, and the IL-10 level were apparently de-creased, but the level of IL-12, IL-4 and GATA-3 mRNA were not changed significantly. Conclusion DCs in asthma regulated the balance of TH1/TH2 by means of secreting decreased IL-12 and increased IL-10, that made TH2 playing a dominance role which is the key factor in initiating asthma. Huang-qi regulated DCs through decreasing the level of IL-10, and thus decreased the ability of inhibiting the differentiation of TH1 from TH0, that is also inhibiting the differentiation of TH2 from TH0 directly.

BACKGROUND:Tissue engineering provides new ideas and approaches for repair of cartilage defects.
OBJECTIVE:To develop a complete set of solutions for construction of tissue engineered cartilagein vitro, with chondrocytes as seed cels and cross-linked sodium hyaluronate as scaffold materials.
METHODS: New Zealand rabbit articular chondrocytes were isolated, counted, and then cultured and passaged to prepare cellsuspension. Toluidine blue staining, RT-PCR and immunocytochemistry were exerted to evaluate the cultured cels. Chondrocytes were seeded and co-cultured with cross-linked sodium hyaluronate scaffold for 21 days. Then, RNA was isolated for RT-PCR, and frozen sections were prepared for morphological observation and immunohistochemistry study.
RESULTS AND CONCLUSION:The chondrocytes could adhere to the cross-linked sodium hyaluronate scaffold and aggregate, growing between fibers or adhering to the scaffold in a monolayer manner. The transcripts of cartilage specific aggrecan gene and colagen type II alpha 1 gene and cartilage specific protein colagen type II were expressed in cel-scaffold complexes to maintain the phenotype of chondrocytes. Cel-scaffold complexes co-cultured in vitro can form cartilage extracelular matrix, by which tissue engineered cartilage is expected to be obtained.

Objective To establish a method of HPLC assay for determining stilbene glucoside in Zishen Ningshen Pills(ZNP),and to study the influence factors on the content of stilbene glucoside in the process of preparation.Methods HPLC was used for the determination of stilbene glucoside in ZNP.Through simulation the process of preparation,the stilbene glucoside content in the intermediate products was determined by HPLC,and its retention rate and metastasis rate were also investigated.Results The resolution and the linearity of stilbene glucoside were fine,the average recoveries being 98 % ～ 102 %.The retention rate of stilbene glucoside in the drying powder was 60.3 %,lower than that in the original medicinal powder.Conclusion The quantitative method for determining the ingredients in ZNP is simple,feasible and reproducible,and is beneficial for quality control of ZNP.The drying process under normal pressure is the main influence factors of the decrease of stilbene glucoside content,and the decompression drying can be taken into account to take the place of the atmospheric drying.

Objective To retrospectively study the high risk factors for biliary complication (BC) and the application of the Clavien system to classify BC in a large cohorts of subjects undergoing liver transplantations (LT).Methods The clinical data of 181 patients who received LT from Jan.2004 to Dec.2008 were studied.BC was classified using the Clavien system.The risk factors of biliary complication were evaluated by using a binary forward stepwise logistic regression analysis.Results 14.4％ (26/181) recipients developed BC (BC group).In 84.6％ (22/26) patients the BC was above the Clavien Ⅲ b.Regression analysis of BC revealed that the placement of a T tube (P =0.0090,OR=31.177),RIld (P=0.0094,OR＜0.001).RI1w (P=0.0013,OR＞999.999) were significantly associated with the development of BC.Regression analysis of BC above Clavien Ⅲ b revealed that RIld (P=0.0065,OR＜0.001,RI1w (P=0.0022,OR＞999.999) were significantly associated with the development of BC above Clavien Ⅲ b.Conclusions The Clavien classification system was useful to classify BC.The placement of a T tube was an independent risk factor to predict BC,it was not a factor for BC above Clavien Ⅲ b.Hepatic arterial insufficiency (HAI) was an independent risk factor for BC and BC above Clavien Ⅲ b.

Objective To construct a eukaryotic expression vector in Pichia pastoris containing human fibrinogen gene, in order to achieve high level secretory expression in extracellular.Methods Expression plasmid,pGAPZαA-FGB-FGG-FGA-AOX1,was constructed by inserting the synthesized sequence encoding human fibrinogen(FGA, FGB,FGG) and then introduced into Pichia pastoris SMD1168H by electroporation.Transformants were availably screened by Zeocin resistance,the expression of recombinant protein was identified by SDS-PAGE and Western blot analysis, the protein yield was tested by ELISA assay.After ultrafiltration and purification, the biological activity of protein was detected.Results The crude yield of human fibrinogen in Pichia pastoris supernatant reached 15 mg/L in flask and the biological aggregation activity was determined.Conclusion The human fibrinogen gene was obtained and successfully expressed in Pichia pastoris and the active products were secreted into the medium.

Aim To investigate the modeling of breast cancer in zebrafish embryos and its related protein expression. Methods 48hpf wild type AB/ TG(Transgenic) zebrafishs were micro-in-jected with breast cancer cell line: MCF-7,T-47D, MDA-MB-231 respectively, the relationship between the number of tumor and model application was investigated, and the number of sub-intestinal veins(SIVs) was detected under confocal microscope, as well as the metastasis of tumor cells in embryos; then the ze-brafish xenografts of MB-231 were co-cultured with tofacitinib/ptk787 for 48 h, optical density(OD) of the cell survival and subintestinal veins(SIVs) were evaluated under confocal micro-scope, and Western blot(WB) analysis was used to test micro-circumstances related protein. Results When the number of in-oculated cells was more than 200 per embryo, xenograft model rate woule be more than 0. 90;MB-231 xenografts showed metas-tasis feature in zebrafish, which could be inhibited by tofacitinib (P < 0. 01), while the number of xenograft MB-231 cells was reduced significantly(P < 0. 01); in another zebrafish xenografts SIVs assay, the tumor could promote the proliferation of SIVs, and 4 mg·L - 1 PTK787 showed inhibiton effect( P < 0. 01). Western blot showed 4d T-47D xenograft zebrafish got more HER2 expression than AB embryos; VEGFa expression in ze-brafish MB-231 model group was higher, and model zebrafish P53 expressi was higher after treated by tofacitinib. Conclusion A zebrafish xenograft model of human brest cancer can be es-tablished, which demonstrates applicability for screening com-pounds in drug discovery studies.

Objective To construct a prokaryotic expression vector in BL21 to secretorily expressα-Cyclodextrin Glycosyltransferase(α-CGTase). Methods α-CGT gene was amplified from Bacillus macerens genome by PCR.pET26b and α-CGT gene were connected after digested with Nco I, Xho I respectivly, and then transformed into Escherichia coli BL21 strain.α-CGTase was expressed in fermentation culture medium and AA-2G was prepared by using α-CGTase, VC and starch.Results α-CGTase was expressed secretorily and the enzyme activity was up to 120 U/mL.AA-2G was prepared by the biotransformation of VC and starch using α-CGTase which proved to be correct by HPLC.Conclusion AA-2G was prepared by using self-madeα-CGTase, after optimized the preparation conditions the yield of AA-2G was 17.46 g/L, and the conversion rate reached 58.2%(mg/mg).

Aim To study the effect of rhein on renal damage induced by aristolochic acid. Methods Ze-brafish model of aristolochic acid nephropathy, genera-ted by treating zebrafish larvae with aristolochic acid for 24 h, was treated with rhein simultaneously . Mor-pholigical changes were observed and the creatinine level in larvae tissue was measured. And mRNA ex-pression levels of inflammatory factor cox2 a and fibrosis factor TGF-β1 in larvae tissue were detected using qPCR. Results Some larvae show periocular edema and circulation system defection e. g. weak heart beat, narrow cardiac vesicle, decreased blood flow and even blockage , with a dose-response relationship after expo-sure to aristolochic acid for 24 h. The creatinine level in larvae tissue of the treated group was significantly higher than that of the control larvae. And the expres-sion levels of cox2 a and TGF-β1 in larvae tissue of the treated group were also significantly increased. Per-centage of abnormal larvae and creatinine level in lar-vae tissue were decreased when treated with rhein sim-ultaneously. And the expression levels of cox2a was down-regulated by rhein compared with the aristolochic acid treated group. But rhein had no effect on TGF-β1 expression. Conclusion To some extent rhein can protect renal from damage induced by aristolochic acid.