A possible ?-Galactosidase gene(pwtsA) was discovered from soil metagenomic DNA of Taishan Mountain.PwtsA gene was inserted into the expression vector pET30a and transferred into E.coli BL21(DE3).Recombinant protein PWTSA was expressed as a soluble form at high level through IPTG induction,with a molecular mass of 57 kD analyzed by SDS-PAGE.PWTSA can produce o-nitrophenol from o-nitrophenol-?-D-galactopyranoside(ONPG),and its specific activity was determined as 13.6 U/mg.The enzymatic studies demonstrated that the recombinant protein PWTSA was a thermostable ?-Galactosidase,its optimum temperature and pH were 85?C～95?C and 6.5 respectively.In standard assays,the Km for ONPG was 0.83 mmol/L.

Objective To investigate the expression and clinical significance of BRMS1 ( breast cancer metastasis suppressorl ) protein in colon cancer.Methods The expression of BRMS1 protein was detected by using EliVision immunohistochemical techniques in 46 cases of colon cancer,and adjacent non cancerous colon tissues.The clinical significance with histopathologic records was aralyzed.Results The expression levels of BRMS1 ( 34.8％ ) in the colon cancer tissues was significantly lower than those of adjacent non cancerous colon tissues( x2 =23.92,P ＜0.01 ).The expression of BRMSl was significantly correlated with tumor size,clinical stage,and lymph node status (x2 =6.02,4.28,4.35,all P＜0.01) ;BRMS1 had no correlation with age,pathological type.Conclusion BRMS1 might synergistically promote the metastasis of colon cancer.Detection of the expression of BRMS1 may be hdpful in determineing the prognosis of colon cancer.

In the pathophysiology experiment teaching,by combining the teaching of prob-lem-based learning with local area network(LAN) teaching,students first carry out a simple ex-periment to ask a question on the observed phenomenon,and then put forward a hypothesis,de-sign experiments to answer questions,and implement the experiment,and finally present experi-mental results.Such experimental design teaching is not only a great way to mobilize the stu-dents’interest in scientific research and learning initiative,but also greatly enhances the effi-ciency of the experiment.Students preliminarily master the basic scientific research program and methods.

OBJECTIVETo research the effect of icariin (ICA) on the proliferation and differentiation of bone mesenchymal stem cells (BMSCs) and to study its influence on the expressions of transforming growth factor-beta1 (TGF-beta1) and bone morphogenetic protein-2 (BMP-2) in the progress of BMSCs differentiating into osteoblast, for providing an experimental evidence to explain the mechanism of ICA, also for exploring the feasibility of establishing a platform upon TGF-beta, and BMP-2 to screen out the medicine in preventing and treating osteoporosis.

METHODSAfter the most effective concentration of ICA for promoting the differentiation of BMSCs into osteoblast was judged with the indices like alkaline phosphatase (ALP), etc., a grouped experiment was conducted for the sake of studying the effect and mechanism of ICA in its process of inducing BMSCs differentiation into osteoblast through detecting expression of ALP and calcium nodes, as well as the expressions of TGF-beta1, and BMP-2 with ELISA.

RESULTSThe most effective concentration of the ICA on the BMSCs differentiation was judged as 1 x 10(-9) mol/L, ICA of that concentration showed effects in enhancing the expressions of osteoblast-indices and increasing the secretion of TGF-beta1, and BMP-2. Besides, the increase of TGF-beta1, and BMP-2 was revealed in all the groups, in which ICA showed its influence visibly.

CONCLUSIONICA could promote the differentiation of BMSCs into osteoblast; the up-regulation of TGF-beta1, and BMP-2 expressions is possibly one of the action mechanisms of various interventional drugs in their differentiation promoting progress.

Animals ; Bone Marrow Cells ; cytology ; drug effects ; Bone Morphogenetic Protein 2 ; metabolism ; Cell Differentiation ; drug effects ; Cells, Cultured ; Flavonoids ; pharmacology ; Mesenchymal Stromal Cells ; cytology ; drug effects ; Osteoblasts ; cytology ; Rabbits ; Transforming Growth Factor beta1 ; metabolism ; Up-Regulation

As the main medicinal powder for drawing out pus and removing necrotic tissue in external therapies of traditional Chinese surgery, Sheng Powder has made great contributions to the treatment of inflammatory wounds and has the unique bactericidal and decay-discharging function that can not be replaced by antibiotics. However, Sheng Powder has toxicity because it contains mercury. So far, there is no clinical research on the standards of dose and usage of Sheng Powder and there is a lack of objective and quantitative criteria for operating standards and monitoring of toxicity and side effects. Therefore, the authors choose Jiuyi Powder, one of the most commonly used Sheng Powder, to evaluate the safety of its external use, and form a standardization program for clinical implementation.

OBJECTIVETo construct plant expression pCAMBIA1301-PMI by substituting PMI for hygromycin resistance gene in pCAMBIA1301 and obtain transgenic Salvia miltiorrhiza f. alba using PMI-mannose selection system.

METHODThe 6-phosphomannose isomerase gene (PMI) of Escherichia coli was amplified by PCR. Sequence analysis showed that it shared 100% amino acids identities with the sequences of PMI genes isolates reported in the NCBI. Based on pCAMBIA1305, the plant expression pCAMBIA1305-PMI was constructed successfully by substituting PMI for hygromycin resistance gene in pCAMBIA1305. pCAMBIA1305-PMI was transformed into Agrobacterium tumefaciens LBA4404, and then the leaves of S. miltiorrhiza f. alba were inoculated in LBA4404 with pCAMBIA1305-PMI.

RESULTPlant expression pCAMBIA1301-PMI was successfully constructed and the leaves of S. miltiorrhiza f. alba inoculated in LBA4404 with pCAMBIA1305-PMI were selected on medium supplemented with a combination of 20 g x L(-1) mannose and 10 g x L(-1) sucrose as a carbon source. The transformation efficiency rate was 23.7%.

CONCLUSIONGenetic transformation was confirmed by PCR, indicating that a new method for obtaining transgenic S. miltiorrhiza f. alba plants was developed using PMI-mannose selection system.

Anti-Bacterial Agents ; pharmacology ; Biomarkers ; Cinnamates ; pharmacology ; Escherichia coli ; enzymology ; genetics ; Escherichia coli Proteins ; genetics ; metabolism ; Gene Expression ; Genetic Vectors ; genetics ; metabolism ; Hygromycin B ; analogs & derivatives ; pharmacology ; Mannose-6-Phosphate Isomerase ; genetics ; metabolism ; Plants, Genetically Modified ; drug effects ; genetics ; metabolism ; Salvia miltiorrhiza ; drug effects ; genetics ; metabolism ; Transformation, Genetic

OBJECTIVETo evaluate the effect of neuronal differentiation induced by nerve growth factor (NGF) on the tolerance-dosage of ultraviolet radiation of PC12 Cells.

METHODSNeuron-differentiated PC12 cells and untreated PC12 cells were exposed to different ultraviolet radiation dosage of 10, 30, 60, 80, 100, and 200 mJ/cm2. Cell survival rates were determined by MTT assay.

RESULTSNeuron-differentiated PC12 cells had increased tolerance dose to ultraviolet radiation with noticeable apoptosis at the radiation dose of 100 mJ/cm2 in contrast to 30 mJ/cm2 for normal PC12 cells.

CONCLUSIONNeuronal differentiation exerts the effect of increasing the tolerance dose of PC12 cells to ultraviolet radiation.

Animals ; Cell Differentiation ; Cell Transformation, Neoplastic ; drug effects ; radiation effects ; Dose-Response Relationship, Radiation ; Nerve Growth Factor ; pharmacology ; Neurons ; cytology ; PC12 Cells ; Rats ; Ultraviolet Rays

OBJECTIVETo study the application of degrading multi-enzymes from Ganoderma lucidum in extracting effective constituents from fibrous roots of Salvia miltiorrhiza.

METHODEffective constituents were extracted from fibrous roots by degrading multi-enzymes of wood fiber. The enzymatic parameters were optimized by the orthogonal design.

RESULTThe extraction efficiencies of total tanshinones and total salvianolic acids in the extracts of fibrous roots of S. miltiorrhiza was obtained using optimum enzymolysis process reached 11.923%, 12.465%, respectively, which were 62.794%, 56.086% more than that by conventional non-enzymatic hydrolysis.

CONCLUSIONDegrading multi-enzymes of wood fiber can be used to fully extract effective constituents from fibrous roots of S. miltiorrhiza, which provides a new approach for recycling wastes of traditional Chinese medicines.

Alkenes ; isolation & purification ; metabolism ; Diterpenes, Abietane ; isolation & purification ; metabolism ; Drugs, Chinese Herbal ; isolation & purification ; metabolism ; Hydrogen-Ion Concentration ; Plant Roots ; chemistry ; Polyphenols ; isolation & purification ; metabolism ; Reishi ; enzymology ; Salvia miltiorrhiza ; chemistry ; Temperature ; Wood ; enzymology

Objective To discuss the technique and technique-related issues of percutaneous kyphoplasty(PKP).Methods The study involved 69 vertebrae in 51 cases of painful osteoporotic vertebral compressive fractures.Under X-ray fluoroscopy monitoring,the fractured vertebral bodies were treated by kyphonplasty with inflatable balloon.The preoperative and postoperative vertebral height and Cobb angle in radiography were measured and analyzed.Results All patients tolerated the procedure well with dramatic pain relief within 72 hours after the procedure.No clinical complication was found.The loss heights of the anterior and mid portion of the vertebral body reduced from 15?4mm and 11?4mm preoperatively to 10? 4mm and 6?3mm postoperatively,respectively.Cobb angle corrected averagely from 22??6? preoperatively to 12??4?.There was significant difference between preoperative and postoperative measures (P

OBJECTIVETo establish a rapid, relatively quantitative method of detecting acetylated proteins.

METHODSThe proteins of Jurkat cells were acetylated by Trichostatin A (TSA) at different concentrations, then enriched and purified by anti-acetylated lysine antibodies affinity chromatography colum. The components eluted by acid were fixed on the microplate, the levels of acetylated proteins were tested by ELISA, and their components were identified by MALDI-TOF-TOF mass spectrometry. Also the above-mentioned methods were applied to the other three agents (gallic acid, emodin and monoacetylated emodin A).

RESULTSThat 4 × 10(5) Jurkat cells treated with 1 µmol/L TSA produced the optimal acetylated effect, up to 22 acetylated proteins were identified by MALDI-TOF-TOF, of them 15 were acetylated histones. The other three agents also induced acetylation, the relative values of acetylated proteins of Jurkat cells treated with 35.09 µmol/L and 17.54 µmol/L gallic acid were 4.3% and 14.2% respectively; those as of 28.7% and 11.5% treated with 1.47 µmol/L and 2.94 µmol/L emodin; those as of 22.0% and 3.6% treated with 152.91 µmol/L and 30.58 µmol/L monoacetylated emodin A.

CONCLUSIONThe method based on affinity chromatography colum may be useful for the detection of acetylated proteins, and could be used to screen agents which target to histone deacetylase.

Acetylation ; Chromatography, Affinity ; Histones ; analysis ; Humans ; Hydroxamic Acids ; pharmacology ; Jurkat Cells ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization