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ABSTRACT: BmK AngM1 is an analgesic peptide from the venom of Buthus martensii Karsch (BmK). The synthetic gene encoding BmK AngM1 was optimized on the basis of its cDNA sequence and the codon usage preference
of Pichia pastoris. The codon-optimized gene was cloned into pPIC9K and then transformed into P. pastoris. SDS-PAGE and Western blot analysis showed that the recombinant BmK AngM1 (rBmK AngM1) was expressed by the addition of methanol
to the medium, and its maximum production reached above 500mg/l. The purified rBmK AngM1 could be obtained efficiently by
Nickel affinity chromatography. Analgesic bioassay, by the mouse-twisting model, showed that rBmK AngM1 had evident analgesic
effect with an ED50 of 0.5mg/kg.
World Journal of Microbiology and Biotechnology 04/2012; 25(11):2053-2056. · 1.53 Impact Factor
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ABSTRACT: To explore the capability of human periodontal ligament stem cells (PDLSCs) differentiating into adipose cells in vitro and to determine their changes in cell morphology, structure and function during differentiation.
PDLSCs isolated by magnetic-activated cell selection were treated continuously with adipogenic medium for 21 d. Then the cell morphology, ultrastructure, adipose specific markers of low density lipoprotein (LPL) and peroxisome proliferator activated receptor-gamma (PPAR-gamma) were analyzed by inverted contrast microscope, trans mission electron microscope (TEM), flow cytometry, immunofluorescence, RT-PCR and Western blot, respectively. These adipose-like cells were also identified by oil red O staining to determine the formation of lipid droplet, and the non-induced cells were used as control.
After continuous induction, the treated cells differentiated into adipose-like cells with round shape, and large amount of lipid drop in cytoplasm. 96.54% of the PDLSCs were found to differentiate into adipose cells as showed by flow cytometry, the specific markers of LPL mRNA and PPAR-gamma mRNA, and oil red O staining, respectively. Further, PPAR-gamma protein was detected in the induced cells in a time-dependent manner.
Human PDLSCs have the potential of differentiating into adipose cells under appropriate condition, and the differentiated cells exhibited characteristics of adipose cells both from cell morphology and from their functions.
Hua xi kou qiang yi xue za zhi = Huaxi kouqiang yixue zazhi = West China journal of stomatology 04/2010; 28(2):203-7.
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ABSTRACT: To isolate, culture and identify a dog periodontal ligament stem cells (PDLSC) line in vitro.
The adult dog periodontal ligament cells were isolated by limited dilution of culture cell for single cell clone. Cells originated from one of these clones were assessed through colony-forming efficiency and immunocytochemistry assay and alkaline phosphatase stain was used to identify the source of adult dog periodontal stem cells, at the same time, PDLSC were induced with mineralizatin solution and was found to have long protrude like an osteoblast. Differentiation of PDLSC were assessed. Mineralized potential was studied by Von-Kossa staining.
The dog PDLSC expressed STRO-1, which was the marker of mesenchymal stem cells. Also Vimentin, osteoblast-like marker alkaline phosphatase and Collagen-I expressed weakly. Cells were clonegenic, highly proliferative cells and capable of differentiating into osteoblasts/cementoblasts.
The evidence suggests that the cultured cells were stem cells from adult dog periodontal ligament.
Hua xi kou qiang yi xue za zhi = Huaxi kouqiang yixue zazhi = West China journal of stomatology 03/2009; 27(1):79-83.
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ABSTRACT: The technology of liquid fermentation for producing the recombinant analgesic peptide BmK AngM1 from Buthus martensii Karsch in Pichia pastoris was studied by single-factor and orthogonal test. The results showed that the optimal culture conditions were as follows: 1.2% methanol, 0.6% casamino acids, initial pH 6.0, and three times of basal inoculation volume. Under the above culture conditions, the expression level of recombinant BmK AngM1 in Pichia pastoris was above 500 mg x L(-1), which was more than three times of the control. The study has laid a foundation for the large-scale preparation of BmK AngM1 to meet the needs of theoretical research of BmK AngM1 and development of new medicines.
Yao xue xue bao = Acta pharmaceutica Sinica 02/2009; 44(1):91-4.
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ABSTRACT: Phylogenetic relationship between Paecilomyces hepiali and Cordyceps sinensis was studied by analyzing the sequence of rDNA-ITS. The samples of C. sinensis were collected from Hualong County in Qinghai Province and Kangding County in Sichuan Province in May and June, respectively. The rDNA-ITS fragments were obtained by PCR amplification with the template genomic DNA of the fresh stroma or caterpillar body of the collected samples and the cultured mycelium of P. hepiali, with the universal fungal primers ITS1/ITS4. The amplified fragments were cloned into pMD18-T Vector and sequenced. Phylogenetic analysis was performed with these sequences and those from GenBank. The result showed that all of the 46 clones randomly chosen from the amplification of C. sinensis shared identical or almost identical rDNA-ITS regions and had over 99% identity with some rDNA-ITS sequences of Hirsutella sinensis and C. sinensis registered in GenBank, but all of them had only about 72% identity with that of P. hepiali. Two pairs of specific primers were designed based on the rDNA-ITS sequence of P. hepiali, then PCR and Nest-PCR were performed with the template genomic DNA of the stroma or caterpillar body of C. sinensis samples mentioned above. The apparent bands amplified by Nest-PCR were obtained from all of the samples, and the sequences showed 100% identity with the rDNA-ITS sequence of P. hepiali. In addition, another pair of specific primers were designed based on the rDNA-ITS sequence registered in GenBank as the marker of C. sinensis (accession no. AB067740) but the latter only shared 87.3% identity with that of H. sinensis (accession no. AJ309353). This pair of primers was used to amplify the C. sinensis samples by PCR, and the amplified sequence showed 100% identity with that of AB067740. The result indicated that H. sinensis is the main body of C. sinensis, while some other endoparasitic fungi such as P. hepiali commonly exist in the natural C. sinensis.
Yao xue xue bao = Acta pharmaceutica Sinica 05/2008; 43(4):421-6.
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ABSTRACT: To explore the multi-differentiated capability of human dental pulp stem cells (hDPSCs) obtained by cell-clone culture approach and to determine the appropriate induced medium.
The cloned isolation and expansion of hDPSCs were preinduced for 24 h, and were subsequently replaced with neural-inductive medium containing certain concentration of dimethylsulfoxide (DMSO), butylated hydroxyanisode (BHA), forskolin, P-mercaptoethanol (p-ME) and hydrocortisone for 4 days. Then induced cells were analyzed by morphological observation, immnocytochemical staining for non-specific esterase (NSE) and glial fibrillary acidic protein (GFAP) expression, RT-PCR for GFAP mRNA. Meanwhile, the uninduced hDPSCs were used as negative control.
The morphology of induced cells changed at the initial 12 h, and displayed a typical neuron-like cells at 24 h. There was a gradual increase in the number of these neuronal differentiated cells with continuous induction. Furthermore, immnocytochemical staining showed that the induced cell expressed NSE and GFAP, two marked enzymes of neuron cell. The GFAP mRNA was also detected in induced cells by RT-PCR assay. In contrast, the uninduced cells maintained its original appearance and had no expression on them.
hDPSCs may possess potential of multiple-differentiation and may differentiate into neuron-like cells on certain inductive condition.
Hua xi kou qiang yi xue za zhi = Huaxi kouqiang yixue zazhi = West China journal of stomatology 09/2007; 25(4):331-4.
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ABSTRACT: 1-Hydroxy-2,3,5-trimethoxyxanthone (1), one of the major xanthone derivatives isolated from Halenia elliptica, was biotransformed by two fungi, Trichothecium roseum and Paecilomyces marquandii. Transformation of 1 by T. roseumgave 1,5-dihydroxy-2,3-dimethoxyxanthone (2), 5-O-sulfate-1-hydroxy-2,3-dimethoxyxanthone (3), 5-O-sulfate-1-hydroxy-2,3,7-trimethoxyxanthone (4), 5-O-beta-ribofuranosyl-1-hydroxy-2,3-dimethoxyxanthone (5), and 1,5,6-trihydroxy-2,3-dimethoxyxanthone (6). Compound 2 was also formed by P. marquandii. The structures of the isolated compounds were elucidated by spectroscopic analyses. Among the five microbial-converted compounds, 3, 4, 5, and 6 are new compounds.
Journal of Natural Products 06/2006; 69(5):811-4. · 3.13 Impact Factor
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ABSTRACT: To select a proper Ganoderma luciderm strain for the fruiting body production.
The strains were cultivated on the agar media and in the liquid media, respectively. Then the strains were inoculated onto the solid medium made from agricultural products (such as wheat bran, corn powder, wood meal, etc.) and cultured for a certain period.
Strains, which were easier to produce polyporic tissues at the vegetative growth stage, would be more quickly to form fruiting body with high quality and yield of the spores.
Appearance of the polyporic tissues at the mycelium vegetative growth stage could be used as a marker for the strain selection for the G. luciderm substituted cultivation.
Zhongguo Zhong yao za zhi = Zhongguo zhongyao zazhi = China journal of Chinese materia medica 02/2006; 31(1):21-3.
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ABSTRACT: Anisodus tanguticus, one of the indigenous Chinese ethnological medicinal plants of the Solanaceae, produces anticholinergic alkaloids such as hyoscyamine, 6 beta-hydroxyhyoscyamine and scopolamine. Hyoscyamine 6 beta-hydroxylase (H6H), a key enzyme in the biosynthetic pathway of scopolamine, catalyzes the hydroxylation of hyoscyamine and epoxide formation from 6 beta-hydroxyhyoscyamine to generate scopolamine. A full-length cDNA of H6H has been isolated from A. tanguticus hairy roots. Nucleotide sequence analysis of the cloned cDNA revealed an open reading frame of 1035 bp encoding 344 amino acids with high homology to other known H6Hs. The equivalent amino acid sequence shows a typical motif of 2-oxoglutarate-dependent dioxygenase. The A. tanguticus H6H was expressed in Escherichia coli and purified for enzyme function analysis. This study characterized the recombinant AtH6H and showed it could generate scopolamine from hyoscyamine.
Planta Medica 04/2005; 71(3):249-53. · 2.15 Impact Factor
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ABSTRACT: To modify the structure of dehydroepiandrosterone (DHEA).
Using hairy root cultures of Anisodus tanguticus to perform biotransformation of DHEA, using chromatographic and spectral techniques to isolate and identify the products.
(1) The MS medium without plant hormone was suitable for the growth of the hairy root. (2) DHEA was converted into five products: androst-4-ene-3, 17-dione (I); 6alpha-hydroxyandrost-4-ene-3, 17-dione (II); 6alpha, 17beta-dihydroxyandrost-4-ene-3-one (III); androst-4-ene-3, 6, 17-trione (IV) and 17beta-hydroxyandrost-4-ene-3-one (V).
It is the first time to use hairy root cultures of Anisodus tanguticus for the biotransformation of DHEA and five DHEA-related compounds were obtained.
Yao xue xue bao = Acta pharmaceutica Sinica 07/2004; 39(6):445-8.
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ABSTRACT: OBJECTIVE: To investigate the relations between child anxiety disorder with different family characte ristics. METHODS Family characteristics were measured by family environment scale. 144 mothers of child with anxiety disorder and 100 mothers of health children were invited to fill out questionnaires. RESULTS There were obviously different family characteristics between health children and those with anxiety disorder except phobic anxiety disorder. The scores of cohesion, independence, achievement orientation and active recreational orientation in children with anxiety disorder were significantly lower than those in health children P<0.01). In 4 groups of children with anxiety disorder cohesion showed correlation with intellectual- cultural orientation r=0.9219, 0.8348, 0.8935, 0.9550 respectively, P<0.001). CONCLUSION: The importance of family characteristics must be emphasized for children with anxiety disorder.
Zhejiang da xue xue bao. Yi xue ban = Journal of Zhejiang University. Medical sciences 08/2002; 31(5):371-374.
