Objective To construct baculovirus expression system for the human plasminogen kringle 5(hK5) gene,and detect its expression in Spodoptera frugiperda 21(sf21). Methods Baculovirus Bacmid-rhK5 was prepared with bac-to-bac baculovirus expression system,and sf21 cells were infected.The expression of rhK5 in sf21 cells was determined by Western blot.The activity of purified protein was detected by vascular endothelial cell inhibition test. Results The pFastBac HTB-rhK5 was successfully constructed,and sf21 cells were transfected with the constructed vector.The output of rhK5 obtained was 90 ug/L,and the protein possessed the inhibitory activity of endothelial cell proliferation.The median effective dose(ED50) was 4 ug/mL. Conclusion The rhK5 baculovirus expression vector is highly expressed in sf21 cells,which lays a foundation for the the expression of rhK5 protein with biologic activities.

Adenoma, Pleomorphic ; metabolism ; pathology ; surgery ; Aged ; Carcinoma ; metabolism ; pathology ; surgery ; Cell Transformation, Neoplastic ; Humans ; Lung Neoplasms ; metabolism ; pathology ; surgery ; Male ; Mucin-1 ; metabolism ; S100 Proteins ; metabolism ; Vimentin ; metabolism

Objective To construct a recombinant lentiviral expression vector containing NIS and EGFP gene,and to explore the feasibility of NIS gene for monitoring the bone marrow derived mesenchymal stem cells (BMSCs) migration to the intracranial glioma.Methods The NIS and EGFP gene fragments were subcloned into lentiviral vector pLVX-puro,then packaged and amplified in HEK293T cells to obtain recombinant lentivirus pLVX-CMV-NIS-EGFP.pLVX-CMV-0-EGFP was constructed as control.BMSCs were isolated,cultured,and transfected by lentivirus.The antibiotic-resistant transfected BMSCs (BMSCs-NIS-EGFP and BMSCs-EGFP) were selected.The expression of NIS gene was examined by Western blot.Functional NIS activity was confirmed by the uptake of 125I and the inhibition effect of NaClO4.The nude mice intracranial glioma models were established.MicroSPECT was performed at 24 h post BMSCs-NIS-EGFP injection via the tail vein.Results pLVX-CMV-NIS-EGFP and pLVX-CMV-0-EGFP were successfully constructed and packaged.BMSCs were successfully isolated and cultured.Stable cell lines BMSCs-NIS-EGFP and BMSCs-EGFP were constructed after lentivirus transfection and puromycin selection.The expression of NIS gene was detected by Western blot in BMSCs-NIS-EGFP,but not in BMSCs-EGFP.BMSCs-NIS-EGFP showed significantly more uptake of 125I (nearly 10 times than the uptake in BMSCs-EGFP) and the uptake could be significantly inhibited by NaClO4.The nude mice intracranial glioma models were successfully established and the BMSCs-NIS-EGFP in glioma foci could be visualized by microSPECT imaging at 24 h post injection.Conclusions A recombinant lentivirus containing NIS gene could be successfully constructed for monitoring BMSCs migration towards intracranial glioma.It might provide evidence on the research of BMSCs and NIS gene mediated therapy for glioma.

Animals ; Cyanides ; toxicity ; Mice ; Random Allocation ; Toxicity Tests, Chronic

Glossectomy ; methods ; Humans ; In Situ Hybridization, Fluorescence ; Keratins ; metabolism ; Male ; Middle Aged ; Oncogene Proteins, Fusion ; genetics ; Sarcoma, Synovial ; genetics ; metabolism ; pathology ; surgery ; Tongue Neoplasms ; genetics ; metabolism ; pathology ; surgery ; Translocation, Genetic ; Vimentin ; metabolism

Paracetamol was used as a model drug in this study to investigate the synergetic effects of lipid coating and beta-cyclodextrin (beta-CD) inclusion for masking the bitter taste of poorly soluble drugs. To control the concentration as low as possible of the free drug which produced a bitter taste, a kinetic model was established to calculate the drug distribution theoretically among the free drug in medium, lipid coated particles and molecular inclusion on the basis of the preparation and characterization of the lipid microspheres, so as to select the proper amount of beta-CD. Finally, the synergetic drug delivery systems were prepared and characterized by 1H nuclear magnetic resonance (1H NMR), molecular simulation and the electronic tongue. As a result, the drug release rate constant (k) of the lipid microspheres coated with octadecanol was determined as 0.001 270 s(-1). Then, the synergetic drug delivery systems were prepared with the ratio of 6.74 : 1 (w/w) for beta-CD and paracetamol. The chemical shift values for the fingerprint peaks of paracetamol all increased and hydrogen bonds were formed between the oxygen on the phenolic hydroxyl group, the nitrogen on the imino in paracetamol and the hydrogens on the hydroxyl groups in beta-CD. The results tested by the electronic tongue indicated that the paracetamol, lipid microspheres, beta-CD inclusion and their mixture showed different taste characteristics, with the bitterness order of the synergetic drug delivery systems approximately lipid microspheres < beta-CD inclusion < paracetamol, which confirmed the synergetic taste masking effects of lipid coating and beta-CD molecular inclusion. In summary, the synergetic taste masking was jointly achieved through the retard of the drug release by the lipid coating and the inclusion of the free paracetamol by beta-CD through hydrogen bonds.
Acetaminophen ; administration & dosage ; chemistry ; Administration, Oral ; Drug Delivery Systems ; Electrical Equipment and Supplies ; Electrochemical Techniques ; instrumentation ; methods ; Hydrogen Bonding ; Kinetics ; Lipids ; chemistry ; Microspheres ; Solubility ; Taste ; drug effects ; beta-Cyclodextrins ; chemistry

The zinc-regulated transporters (ZRT), iron-regulated transporter (IRT)-like protein (ZIP) plays an important role in the growth and development of plant. In this study, a full length cDNA of ZIP encoding gene, designed as DoZIP1 (GenBank accession KJ946203), was identified from Dendrobium officinale using RT-PCR and RACE. Bioinformatics analysis showed that DoZIP1 consisted of a 1,056 bp open reading frame (ORF) encoded a 351-aa protein with a molecular weight of 37.57 kDa and an isoelectric point (pI) of 6.09. The deduced DoZIP1 protein contained the conserved ZIP domain, and its secondary structure was composed of 50.71% alpha helix, 11.11% extended strand, 36.18% random coil, and beta turn 1.99%. DoZIP1 protein exhibited a signal peptide and eight transmembrane domains, presumably locating in cell membrane. The amino acid sequence had high homology with ZIP proteins from Arabidopsis, alfalfa and rice. A phylogenetic tree analysis demonstrated that DoZIP1 was closely related to AtZIP10 and OsZIP3, and they were clustered into one clade. Real time quantitative PCR analysis demonstrated that the transcription level of DoZIP1 in D. officinale roots was the highest (4.19 fold higher than that of stems), followed by that of leaves (1.12 fold). Molecular characters of DoZIP1 will be useful for further functional determination of the gene involving in the growth and development of D. officinale.
Amino Acid Sequence ; Cloning, Molecular ; Dendrobium ; chemistry ; classification ; genetics ; metabolism ; Gene Expression Regulation, Plant ; Iron ; metabolism ; Membrane Transport Proteins ; chemistry ; genetics ; metabolism ; Molecular Sequence Data ; Phylogeny ; Plant Proteins ; chemistry ; genetics ; metabolism ; Plants ; chemistry ; classification ; genetics ; Sequence Alignment ; Zinc ; metabolism

AIM:To investigate the affecting factors of detecting platelet activation by flow cytometry (FCM). METHODS:Using decoagulant of natrium citricum,anticoagnlated peripheral venous bloods from 6 healthy donors were labeled with the method of three-colour immunofluorescence assay. Platelet activation markers fibrinogen receptor (Fib-R,PAC-1) and P-selectin (CD62P) were measured. In the same time,the reproducibility of FCM was assessed.RESULTS:The platelet activation markers PAC-1 and CD62P at each time point showed significant difference(P

Objective To observe the structural characteristic of Marshall ligament and coronary sinus of 40 dog hearts. Methods The Marshall ligament and coronary sinus of 40 dog hearts fixed with 10% formaldehyde were observed in gross anatomy. Results The Marshall ligament begins at the right external side edge of left atrium appendage, via the left inferior pulmonary vein to the inner surface of pericardium. The length and the width of Marshall ligament of dog hearts were (2.61?0.95) cm, (0.18?0.04) cm respectively. The angle between Marshall ligament and the coronary sinus was (116?11), and the shortest length between Marshall ligament and left atrium was (0.58?0.52) cm. The length and the width of coronary sinus was (2.84?0.85) cm, (0.28?0.07) cm respectively. There are lots of cardiac fibers of ring (47.5%), vertical (37.5%) and diagonal (15%) shape on the surface of coronary sinus. Conclusion A lot of muscle fibers attaching to the surface of Marshall ligament and coronary sinus in dog hearts are abnormal atrial channels. Dog is the favorable animal for research of focal atrial fibrillation.

Objective To retrospectively investigate the disease spectrum of inpatients aged over 65 year and cost constitution in Sichuan Provincial People's Hospital from 2010 to 2014,so as to provide baseline data for further study.Methods The inpatients'disease spectrum and costs were collected from hospital information system.The diseases were classified according to the International Classification of Diseases(ICD-10).The data were analyzed using SPSS 18.0 software.Results The total number of old inpatients was 111,935,and male (55.2 ％) was more than female (44.8 ％).The average age was (74.5 ±6.8)years.The top four systematic diseases of primary diagnosis were circulatory system disease (21.0 ％),respiratory system disease (13.7 ％),digestive system disease (12.7％)and neoplasms (12.1％).The total number of male inpatients was more than the female inpatients.The average cost per capita was increased from (￥)18,778.1 yuan to (￥)23,391.9 yuan since 2010.The proportion of all costs accounted for by drugs in elderly inpatients was decreased from 45.5％ to 38.9％ since 2010.Nosocomial infection was decreased from 3.19％ to 0.16％ since 2010.Conclusions The number of elderly inpatients are more in male than in female in Sichuan Provincial People's Hospital from 2010 to 2014.The most common systematic disease is circulatory system diseases,and followed by respiratory system diseases,digestive system diseases and neoplasms.The proportion of all expenditures accounted for by drugs was declined in five years,while the average cost per capita is increased.