OBJECTIVEAn accurate and reliable analytical method for-simultaneous determination of six active components (scopolin, chlorogenic acid, scopoletin, isochlorogenic acid A, isochlorogenic acid B and isochlorogenic acid C) in plants of Erycibe was developed.

METHODScopolin, chlorogenic acid, scopoletin, isochlorogenic acid A, isochlorogenic acid B and isochlorogenic acid C in the samples were well separated in analytical HPLC by gradual elution with methanol-0.1% formic acid solution. The chromatographic condictions: Agilent Poroshell 120 EC-C18 column, flowing rate being 1 mL x min(-1), detecting wavelength at 345 nm.

RESULTGood linearities of scopolin, chlorogenic acid, scopoletin, isochlorogenic acid A, isochlorogenic acid B and isochlorogenic acid C were in the range of 0.026 8-2.68, 0.027 0-2.70, 0.008 1-0.81, 0.018 8-1.88, 0.017 6-1.76, 0.019 6-1.96 μg, respectively (r > 0.999 6). The average recoveries of the six components were 98.1%, 98.7%, 100.8%, 100.4%, 99.7%, 101.1%; the relative standard deviations were 2.67%, 2.86%, 2.62%, 1.98%, 2.76%, 2.19%.

CONCLUSIONThe method is simple, feasible and reproducible and can be used for the quality control of plants of Erycibe.

China ; Chlorogenic Acid ; analogs & derivatives ; analysis ; Chromatography, High Pressure Liquid ; methods ; Convolvulaceae ; chemistry ; Coumarins ; analysis ; Drugs, Chinese Herbal ; analysis ; Glucosides ; analysis ; Scopoletin ; analysis

Objective To analyze the epidemiologic characteristics and risk factors for mortality in non-(human immunodeficiency virus,HIV) infected children with pneumocystis carinii pneumonia(PCP). Methods The data of non-HIV infected children with PCP diagnosed in Beijing Children′s Hospital from January 1,2006 to December 31,2012 were collected. They were divided into survival and non-survival group according to the prognosis. The epidemiologic characteristics and risk factors for mortality were analyzed. Results Sixteen patients were enrolled in this study. Ten of them survived and 6 of them were non-survived. The basic diseases included malignant tumor in 5 patients and non-malignancy diseases in 11 of them. Com-pared with the survival group,the non-survival group had a higher average age [(12. 00 ± 2. 00) years vs. (6. 65 ± 4. 32)years,P=0. 01],higher ratio to need mechanical ventilation (6/6 vs. 4/10,P=0. 04),lower PaO2/FiO2[(73. 88 ±26. 95) mmHg vs. (167. 50 ± 97. 17) mmHg,1 mmHg=0. 133 kPa,P=0. 01] and lower pediatric critical illness score(75. 67 ± 5. 72 vs. 86. 40 ± 8. 88,P=0. 02). There were no differences on sex ratio,kinds of basic diseases,whether with co-infections,the time of immunosuppressant administration, the time from onset to diagnosis,the time from onset to beginning trimethoprim-sulfamethoxazole therapy, PaCO2 ,white blood cell counts,lymphocyte counts,CD4+ cell counts,C-reactive protein,and hemoglobin con-centrations between the survival and non-survival group. Conclusion A higher age, need for mechanical ventilation,lower PaO2/FiO2 and lower pediatric critical illness score were risk factors for mortality in non-HIV infected children with PCP.

italic>Artemisia argyi (A. argyi) is a Chinese herbal medicine in China. The main active components are volatile oils, flavonoids, and other compounds, which have various pharmacological activities. Methoxylated flavonoids are the main active ingredients in A. argyi. Flavonoid O-methyltransferase (FOMT) is a key enzyme in the O-methylation of flavonoids. In order to further understand the function and characteristics of FOMT proteins, this paper carried out the whole genome mining and identification of FOMT genes in A. argyi and performed phylogenetic, chromosomal localization, gene sequence characterization, subcellular localization prediction, protein structure, gene structure analysis, and expression pattern analysis. The results showed that a total of 83 FOMT genes were identified in the genome of A. argyi. The phylogenetic tree shows that FOMT genes are divided into two subgroups, CCoAOMT (caffeoyl CoA O-methyltransferase) subfamily (32 genes) and COMT (caffeic acid O-methyltransferase) subfamily (51 genes). Gene sequence analysis showed that the number of amino acids encoded by FOMT was 70-734 aa, the molecular weight was 25 296.55-34 241.3 Da, and the isoelectric point was 4.51-9.99. Compared with 32 members of the CCoAOMT subfamily, nearly 1/3 of the 51 members of the COMT subfamily were hydrophobic proteins and 2/3 were hydrophilic proteins. Subcellular localization prediction showed that more than 80% of CCoAOMT subfamily members were located in the cytoplasm, and 96% of COMT subfamily members were located in the chloroplast. COMT subfamily members have more motifs than CCoAOMT subfamily members. The N-terminal motifs of COMT subfamily proteins are relatively variable, while the C-terminal motifs are relatively conserved. Expression pattern analysis showed that CCoAOMT subfamily members were mainly expressed in roots, while COMT members were mainly expressed in leaves. Some FOMTs showed the tissue expression specificity by real-time quantitative PCR analysis, especially in leaves. In this study, we identified and analyzed the FOMT gene family in A. argyi, and provided a theoretical basis for further research on the function of FOMTs and the biosynthesis of methylated flavonoids in A. argyi.

Methods: The clinical data of 26 patients diagnosed with irreducible atlantoaxial dislocation complicated by atlantodental bony obstruction were analyzed retrospectively. All patients underwent anterior retropharyngeal atlantodentoplasty, followed by posterior occipitocervical fusion. Details including surgical duration and blood loss volume were recorded. Radiographic data such as the anterior atlantodental interval, O–C2 angle, space available for the cord, clivus–canal angle, and cervical medullary angle, and clinical data including the Japanese Orthopedic Association (JOA) score were assessed. The fusion time of the grafted bone and the development of complications were examined. Results: In patients undergoing anterior retropharyngeal atlantodentoplasty, the surgical duration and blood loss volume were 120.1±16.4 minutes and 100.6±33.5 mL, respectively. The anterior atlantodental interval decreased significantly after the surgery (p <0.001). The O–C2 angle, space available for the cord, clivus–canal angle, and cervical medullary angle increased significantly after the surgery (p <0.001). The JOA score during the latest follow-up significantly increased compared with that before the surgery (p <0.001). The improvement rate of the JOA score was 80.8%±18.1%. The fusion time of the grafted bone was 3–8 months, with an average of 5.7±1.5 months. In total, 11 patients presented with postoperative dysphagia and three with irritating cough. However, none of them exhibited other major complications. Conclusions Anterior retropharyngeal atlantodentoplasty can anatomically reduce the atlantoaxial joint with a satisfactory clinical outcome in patients with irreducible atlantoaxial dislocation with atlantodental bony obstruction.

Methods: The clinical data of 26 patients diagnosed with irreducible atlantoaxial dislocation complicated by atlantodental bony obstruction were analyzed retrospectively. All patients underwent anterior retropharyngeal atlantodentoplasty, followed by posterior occipitocervical fusion. Details including surgical duration and blood loss volume were recorded. Radiographic data such as the anterior atlantodental interval, O–C2 angle, space available for the cord, clivus–canal angle, and cervical medullary angle, and clinical data including the Japanese Orthopedic Association (JOA) score were assessed. The fusion time of the grafted bone and the development of complications were examined. Results: In patients undergoing anterior retropharyngeal atlantodentoplasty, the surgical duration and blood loss volume were 120.1±16.4 minutes and 100.6±33.5 mL, respectively. The anterior atlantodental interval decreased significantly after the surgery (p <0.001). The O–C2 angle, space available for the cord, clivus–canal angle, and cervical medullary angle increased significantly after the surgery (p <0.001). The JOA score during the latest follow-up significantly increased compared with that before the surgery (p <0.001). The improvement rate of the JOA score was 80.8%±18.1%. The fusion time of the grafted bone was 3–8 months, with an average of 5.7±1.5 months. In total, 11 patients presented with postoperative dysphagia and three with irritating cough. However, none of them exhibited other major complications. Conclusions Anterior retropharyngeal atlantodentoplasty can anatomically reduce the atlantoaxial joint with a satisfactory clinical outcome in patients with irreducible atlantoaxial dislocation with atlantodental bony obstruction.

Methods: The clinical data of 26 patients diagnosed with irreducible atlantoaxial dislocation complicated by atlantodental bony obstruction were analyzed retrospectively. All patients underwent anterior retropharyngeal atlantodentoplasty, followed by posterior occipitocervical fusion. Details including surgical duration and blood loss volume were recorded. Radiographic data such as the anterior atlantodental interval, O–C2 angle, space available for the cord, clivus–canal angle, and cervical medullary angle, and clinical data including the Japanese Orthopedic Association (JOA) score were assessed. The fusion time of the grafted bone and the development of complications were examined. Results: In patients undergoing anterior retropharyngeal atlantodentoplasty, the surgical duration and blood loss volume were 120.1±16.4 minutes and 100.6±33.5 mL, respectively. The anterior atlantodental interval decreased significantly after the surgery (p <0.001). The O–C2 angle, space available for the cord, clivus–canal angle, and cervical medullary angle increased significantly after the surgery (p <0.001). The JOA score during the latest follow-up significantly increased compared with that before the surgery (p <0.001). The improvement rate of the JOA score was 80.8%±18.1%. The fusion time of the grafted bone was 3–8 months, with an average of 5.7±1.5 months. In total, 11 patients presented with postoperative dysphagia and three with irritating cough. However, none of them exhibited other major complications. Conclusions Anterior retropharyngeal atlantodentoplasty can anatomically reduce the atlantoaxial joint with a satisfactory clinical outcome in patients with irreducible atlantoaxial dislocation with atlantodental bony obstruction.

OBJECTIVETo investigate the culture method of skin-derived precursors (SKPs) and to explore a new cell source for cell transplantation of central nervous system.

METHODSCells from skins of juvenile and adult mice were isolated and cultured in serum-free medium. A mechanical method was chosen to passage these cells and they were identified by the immunocytochemistry assay.

RESULTSSKPs could be isolated from adult and neonatal skins. They could be maintained in vitro for long periods with stable proliferation, and expanded as undifferentiated cells in culture for more than 12 passages. About 50% of SKPs expressed nestin and majority of these cells expressed fibronectin when they were plated on polyornithine and laminin coated plates. About 5% cells showed neuronal differentiation and expressed neurofilament-M (NF-M) and NSE when SKPs were plated in serum-containing medium, and these cells could also differentiate into adipocytes and fibroblast-like cells.

CONCLUSIONSThe data support the hypothesis that adult skin contains stem cells capable of differentiating into neurons, adipocytes, and fibroblast-like cells. They may represent an alternative autologous stem cell source for CNS cell transplantation.

Adipocytes ; Animals ; Cell Differentiation ; physiology ; Cells, Cultured ; Immunohistochemistry ; Mice ; Mice, Inbred BALB C ; Neurons ; Skin ; cytology ; Stem Cell Transplantation

OBJECTIVETo explore the culture conditions of human neural stem cells and to investigate the ultrastructure of neurospheres.

METHODSThe cells from the embryonic human cortices were mechanically dissociated. N2 medium was adapted to culture and expand the cells. The cells were identified by immunocytochemistry and EM was applied to examine the ultrastructure of neurospheres.

RESULTSThe neural stem cells from human embryonic brains were successfully cultured and formed typical neurospheres in suspension, and most of the cells expressed vimentin, which was a marker for neural progenitor cells, and the cells could differentiate into neurons, astrocytes and oligodendrocytes. In vitro myelin formation in neurospheres were observed at an early stage of culture.

CONCLUSIONSHuman neural stem cells can be cultured from embryonic brains, can form the typical neurospheres in suspension in vitro and have the ability of myelinating, and may be potential source for transplantation in treating myelin disorders.

Brain ; cytology ; Cells, Cultured ; Culture Media ; Female ; Humans ; Immunohistochemistry ; Male ; Microscopy, Electron ; Myelin Sheath ; pathology ; ultrastructure ; Neurons ; cytology ; pathology ; ultrastructure ; Sensitivity and Specificity ; Stem Cell Transplantation ; Stem Cells ; physiology ; ultrastructure

OBJECTIVETo investigate the differentiative capability of adult human bone marrow mesenchymal stem cells (BMSCs) into Schwann-like cells.

METHODSBone marrows were aspirated from healthy donors and mononuclear cells were separated by Percoll lymphocytes separation liquid (1.073 g/ml) with centrifugation, cells were cultured in DMEM/F12 (1:1) medium containing 10% fetal bovine serum (FBS), 20 ng/ml epidermal growth factor (EGF) and 20 ng/ml basic fibroblast growth factor (bFGF). Cells of passage 1 were identified with immunocytochemistry.

RESULTSMononuclear cells separated by Percoll's were passaged 10 times by trypsin/ethylenediaminetetraacetic acid (EDTA) digestion in 40 days, and BMSCs increased about 6x10(7) times in this short period. Immunohistochemistry identified that BMSCs were CD34- and CD31-, but they expressed neuron specific enolase; 0.01%-0.02% of total cells expressed nestin, the marker for neural progenitor cells; 40%-50% cells stained heavily by neurofilament 200; and no glial fibrillary acidic protein (GFAP) positive cells were identified; S100 expression was detected among 0.1%-0.2% cells.

CONCLUSIONSBone marrow contains the stem cells with the ability of differentiating into Schwann-like cells, which may represent an alternative stem cell sources for neural transplantation.

Adult ; Bone Marrow Cells ; cytology ; metabolism ; Cell Differentiation ; physiology ; Cell Proliferation ; Humans ; Immunohistochemistry ; Intermediate Filament Proteins ; metabolism ; Mesenchymal Stromal Cells ; cytology ; metabolism ; Nerve Tissue Proteins ; metabolism ; Nestin ; Neurofilament Proteins ; metabolism ; Phosphopyruvate Hydratase ; metabolism ; S100 Proteins ; metabolism ; Schwann Cells ; cytology

OBJECTIVETo construct a phosphatidylinositol 4-kinase beta (PI4K-beta) mutant with the 325th to 373rd amino acid codons deleted, and try to develop a simple method for constructing middle fragment deletion mutant.

METHODSIn line with the mechanism of gene splicing by overlap extension(SOE), an additional PCR was used to get the PI4K-beta mutant in which the 325th to 373rd amino acid codons were deleted. Then the mutated gene was cloned into pCMV-Tag4A mammalian expression vector.

RESULTSA mutant with the 325th to 373rd amino acid codons deleted was constructed successfully.

CONCLUSIONThe improved SOE is a very effective and reliable method to construct middle fragment deletion mutant. It is worthy to be popularized.

1-Phosphatidylinositol 4-Kinase ; genetics ; Base Sequence ; Genetic Vectors ; genetics ; Molecular Sequence Data ; Mutagenesis, Site-Directed ; Mutation ; genetics ; Polymerase Chain Reaction ; methods ; Protein Engineering ; methods ; Recombinant Proteins ; genetics ; Sequence Deletion