Twenty-nine antagonistic bacillus strains, isolated from some Chinese traditional medicine and fermented food , inhibit the growth of Fusarium oxysporum f. sp. Vasinfectum and Verticillium dahliae Kleb. And twelve of them are able to produce antagonistic proteins. Among the twelve strains, five (H110, H184, H216, B316 and B382) showed higher antibacterial activity. Furthermore, H110 and H184 were identified as Bacillus subtilis, and H216, B316 and B382 as Bacillus licheniformis based on physiological and biochemistry experiments. The antagonistic proteins of five strains were all thermostable, resistant to proteinase K and trypsin, while H184 and H216 partially sensitive to pepsin.

0.05).Conclusions The results by GH exercise test is identified with GH provocative test.Because the GH exercise test is safe and simple,it is appropriate to use the GH exercisetest to screen for growth hormone deficiency during childhood.

Objective To evaluate the image quality of 64-multi detector computed tomography (MDCT)and the clinical accuracy in detecting coronary artery lesions.Methods One hundred and five patients were studied by MDCT.The results were compared with invasive coronary angiography(ICA). Patients were excluded for atrial fibrillation,but not for high heart rate,coronary calcification,or obesity. MDCT was analyzed with regard to image quality and presence of coronary artery lesions.Results The data evaluation of the image quality was based on a total of 1365 segments(13 coronary segments for each patient),of which 1144 segments were considered to have diagnostic image quality,but 221 segments (16.2%)could not be sufficiently evaluated because of severe calcifications(153 segments)and motion artifacts(68 segments).The median calcium score[Agatston score equivalent(ASE)]was 154(range 0—1983).87 of the 105 patients had an ASE of less than 1,000[median 105(range 0—994)],and 18 patients had an ASE greater than 1000[median 1477(range 1115—1983)].For detecting lesions with 50% or greater narrowing(without any exclusion criteria),the overall sensitivity,specificity,positive predictive value,and negative predictive value were 85.7%,97.9%,93.0%,and 95.5%,respectively. When limiting the number of patients to those with a calcium score of less than 1000 ASE,the threshold- corrected sensitivity for lesions with 50% or greater narrowing was 96.0%;specificity,98.9%;positive predictive value,95.3%;and negative predictive value,99.0%.Conclusion Our results indicate high quantitative and qualitative diagnostic accuracy of 64-slice MSCT in comparison to QCA in a broad spectrum of patients.

OBJECTIVETo construct the acid-sensitive potassium hannel-3(TASK3) eukaryotic expression plasmid and to establish a stable SH-SY5Y cell line expressing enhanced green fluorescent protein (EGFP)-tagged TASK3.

METHODSTASK3 coding region was subcloned into pEGFP-N1 plasmid to construct a recombinant vector alled pEGFP-TASK3. The correct recombinant expressing plasmid was transfected with X-feet transfection reagent to SH-SY5Y cells. The cell line stably expressiing EGFP tagged-TASK3 gene was established by screening with antibiotic G418 and fluorescence microscope. The expression and localization of the EGFP tagged-TASK3 fusion protein was detected by Western blot and confocal microscope. Exposure of the SH-SY5Y cell line expressing stably TASK3-eGFP fusion proteins was exposed to different pH media (7.0, 6.7, 6.4, 6.1) for 24 h, the cell viability was assessed with cell counting Kit-8 (CCK-8).

RESULTSAll the results of identification by PCR, digestion with restriction endonuclease and sequencing indicated that the recombinant eukaryotic expression plasmid pEGFP-TASK3 was constructed correctly. The stable SH-SY5Y cell line expressing EGFP tagged-TASK3 fusion protein was successfully established. Exposure of the wild type SH-SY5Y cells and the stable SH-SY5Y-GFP tag-TASK3 cell line to different pH media (7.0, 6.7, 6.4, 6.1) for 24 h, the cell viability of two group cells significantly reduced with pH declining, and the difference was statistically significant (P < 0.05). Compared with wild type SH-SY5Y cells, the cell viability of stable SH-SYSY-GFP tag-TASK3 cell line increased significantly with the same pH media, and the difference was statistically significant (P < 0.05).

CONCLUSIONThe eukaryotic expression vector pEGFP-TASK3 is successfully constructed and the cell line stably expressing TASK3-eGFP fusion is established which is important for their fundamental research and potential applications.

Blotting, Western ; Cell Line ; Gene Expression ; Genetic Vectors ; Green Fluorescent Proteins ; genetics ; Humans ; Plasmids ; Polymerase Chain Reaction ; Potassium Channels, Tandem Pore Domain ; genetics ; Transfection

To study the effect of pulchinenoside (PULC) on the Frizzled (FZD) expression of adjuvant arthritis ( AA) rats. AA rats were prepared through the toe injection with complete Freund's adjuvant to culture fibroblast-like synoviocytes (FLS). The effect of the oral administration with PULC on the FZD8 expression was detected by the real time qPCR. The effect of FZD8 knockout on the expressions of IL-1, IL-6, IL-8 were detected by MTT and ELISA. The role of miR-375 in the abnomal expression of FZD8 was detected by the real time qPCR. The results showed signfiicant decrease in the FZD8 expression among AA rats, FLS proliferation ater FZD8 knockout and IL-1, IL-6, IL-8 expressions and notable increase in miR-375 expression after the oral administration with PULC. The up-regulated miR-375 expression can inhibit the FZD8 expression. PULC may inhibit the FZD8 expression by up-regulating the miR-375 expression.
Animals ; Arthritis, Experimental ; drug therapy ; genetics ; metabolism ; Disease Models, Animal ; Drugs, Chinese Herbal ; administration & dosage ; Humans ; Male ; Rats ; Rats, Sprague-Dawley ; Receptors, Cell Surface ; genetics ; metabolism ; Saponins ; administration & dosage

The role of pulchinenoside (PULC) in the regulation of MeCP2 expression was investigated in RA model rats. Adjuvant arthritis rats were used as RA model rats, and fibroblast-like synoviocytes (FLS) from the RA model rats were cultured. The effect of 100 mg x kg(-1) PULC gavage treatment on the MeCP2 expression and the effect of MeCP2 siRNA on the expression of SFRP2 and β-catenin were detected by real time qPCR and Western blotting. The role of PULC in the FLS proliferation was detected by MTT. The results showed that the MeCP2 expression was down-regulated, the SFRP2 expression was up-regulated and the FLS proliferation was inhibited in FLS after therapy. MeCP2 siRNA significantly inhibited the MeCP2 expression, up-regulated the SFRP2 expression and inhibited the β-catenin expression in FLS from RA model rats. PULC may increase the SFRP2 expression, inhibit the Wnt signaling and inhibit the FLS proliferation in FLS from the RA model rats by inhibiting the MeCP2 expression.
Animals ; Arthritis, Rheumatoid ; drug therapy ; genetics ; metabolism ; Disease Models, Animal ; Drugs, Chinese Herbal ; administration & dosage ; Fibroblasts ; drug effects ; metabolism ; Gene Expression Regulation ; drug effects ; Humans ; Male ; Methyl-CpG-Binding Protein 2 ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Synovial Membrane ; cytology ; drug effects ; metabolism ; Wnt Signaling Pathway ; drug effects ; beta Catenin ; genetics ; metabolism

OBJECTIVE: To develop a specific, sensitive, reproducible SPE-HPLC method for the determination of 37 drugs in whole blood. METHODS: With the doxapram as internal standard, Oasis column was used to extract drugs from whole blood. Two kinds of mobile phases were used in this study. Separations were achieved by a LiChrospher 100 RP-C18 (250 mm x 4.0 mm x 5 microm) column kept at 50 degrees C, the DAD detector was set at 230 nm and 250 nm. RESULTS: The limit of detection were 1-30 ng/mL. The method showed excellent linearity and the linear correlation coefficient was > or =0.997 98. The relative standard deviation for between-day and within-day assay were <10%. CONCLUSION The method is effective, simple, reliable and has been used in real cases.
Chromatography, High Pressure Liquid/methods* ; Doxapram/isolation & purification* ; Doxepin/isolation & purification* ; Estazolam/isolation & purification* ; Forensic Medicine ; Humans ; Morphine/isolation & purification* ; Papaverine/isolation & purification* ; Pharmaceutical Preparations/isolation & purification* ; Prazosin/isolation & purification* ; Procaine/isolation & purification* ; Reproducibility of Results ; Sensitivity and Specificity ; Solid Phase Extraction/methods* ; Solvents/chemistry*

AIMTo construct short hairpin RNA (shRNA) eukaryotic expression vectors targeting TGF-beta1 for further research on the effects of TGF-beta1 on vasculogenesis and angiogenesis.

METHODSThree pairs of siRNA target sequences coding from the mRNA of TGF-beta1 gene were designed and three pairs of nucleotides were synthesized. After annealing, the double-strand DNA products were ligated into the pEN_mH1c entry vector, and in turn into the shRNA eukaryotic expression vector pDS_hpEy labled by GFP through the LR recombination reaction. After sequencing successfully, the three resulting TGF-beta1 shRNA expression vectors were transfected into the mouse fibroblast cell line (NIH/3T3), and then cell clones stably expressing TGF-beta1 shRNA were screened. Reverse Transcript-Polymerase Chain Reaction (RT-PCR) and Western blot were used to detect the mRNA and protein expression.

RESULTSRT-PCR and Western blot showed that one of the TGF-beta1 shRNA expression vectors pDS_Tc downregulated TGF-pl mRNA and protein expression markedly in NIH/3T3 cells.

CONCLUSIONShRNA eukaryotic expression vectors targeting TGF-beta1 are successfully constructed which can be used for further investigation on the mechanism through which TGF-beta1 regulates vasculogenesis and angiogenesis.

Animals ; Genetic Vectors ; Green Fluorescent Proteins ; genetics ; metabolism ; Mice ; NIH 3T3 Cells ; Neovascularization, Physiologic ; RNA Interference ; RNA, Messenger ; genetics ; metabolism ; RNA, Small Interfering ; genetics ; Transfection ; Transforming Growth Factor beta1 ; genetics ; metabolism

In order to discover light quality's effects on growth, photosynthesis and effective components content of Panax notoginseng, a pot experiment using 7 light qualities (red, orange, yellow, green, cyan, violet, and blue) was conducted. The growth, photosynthesis and content change of effective components were measured during plant growth. The results showed that light qualities had significant effect on plant growth, red light increased the plant height, while cyan, yellow, violet, and blue lights promoted accumulation of biomass underground, blue and yellow lights increased the photosynthesis, cyan light increased accumulation of ginsenoside Rd, yellow and cyan lights increased total effective components of individual plant.
Light ; Panax notoginseng ; growth & development ; metabolism ; radiation effects ; Photosynthesis ; radiation effects ; Plant Extracts ; analysis ; metabolism

OBJECTIVETo elucidate the possible mechanism of oral carcinogenesis and to explore the value of clinical application of the detection of cytokeratin (CK) 19 for oral squamous cell carcinoma (OSCC) patients.

METHODSThe cancerous tissues, para-cancerous tissues and excised lymph nodes were collected from 20 operated patients with OSCC. The patients didn't receive radiotherapy and chemotherapy before hospitalization. The relative expression of CK19 mRNA in those tissues was detected by fluorescent quantitative polymerase chain reaction (FQ-PCR).

RESULTSThe expression of CK19 mRNA in the cancerous tissues was 1.85 and 1.66 times higher than that in normal oral mucosa and in para-cancerous tissues, respectively. The expression of CK19 mRNA in lymph nodes from 9 patients with OSCC was positive and the positive rate was 45% (9/20). The positive rate of CK19 mRNA in all lymph nodes from 9 patients with OSCC was 81.8% (18/22), and the positive rate of CK19 mRNA in all lymph nodes from 20 patients with OSCC was 41.9%(18/43). CK19 mRNA level in the cancerous tissues relative to para-cancerous tissues and normal oral mucosa of the patients whose CK19 mRNA expression was positive was lower than that of the patients whose CK19 mRNA expression was negative in lymph nodes, respectively.

CONCLUSIONThe possible reason that the expression of CK19 mRNA in the cancerous tissues was higher than that in para-cancerous tissues and normal oral mucosa was that the CK19 synthesis in cancerous tissues increased obviously. The detection of CK19 mRNA in lymph nodes was regarded probably as one of the markers for detecting OSCC micrometastasis in lymph nodes. The detection of CK19 mRNA in lymph nodes by FQ-PCR was more sensitive than hematoxylin-eosin staining in diagnosing OSCC micrometastasis.

Carcinoma, Squamous Cell ; Female ; Humans ; Keratin-19 ; Male ; Middle Aged ; Mouth Mucosa ; Mouth Neoplasms ; RNA, Messenger