Adult ; Arsenic ; pharmacology ; Chromosome Aberrations ; chemically induced ; Dose-Response Relationship, Drug ; Female ; Humans ; Lymphocytes ; drug effects ; metabolism ; Male ; Middle Aged

The total RNA was extracted from ginseng leaves of Panax ginseng. The Cu/Zn-SOD gene was amplified via RT-PCR and the pET-28(a)-Cu/Zn-SOD expression vector was constructed. The pET-28 (a)-Cu/Zn-SOD recombinant plasmid was transformed into Escherichia coli BL21 (DE3) competent cells and was induced by IPTG in order to select optimal induction of expression conditions. The target protein was purified by the nickel ions (Ni ) affinity chromatography and the target protein enzyme activity was determinated by the xanthine oxidase method. The similarity of the Cu/Zn-SOD gene sequences and the Cu/Zn-SOD gene sequences of Korean ginseng in NCBI was 99. 00%. The target protein expression level was about 44.42%, and the molecular weight was 16.30 kDa after the pET-28(a)-Cu/Zn-SOD recombinants were induced by IPTG. The purified Cu/Zn-SOD protease activity reached 10,596.69 U x mg(-1). The P. ginseng pET-28(a)-Cu/Zn-SOD prokaryotic expression vector was built by the method of molecular biology, which provided the foundation for studying the Cu/Zn-SOD biology function.
Cloning, Molecular ; Escherichia coli ; genetics ; Gene Expression ; Genetic Engineering ; methods ; Genetic Vectors ; genetics ; Panax ; enzymology ; genetics ; Sequence Analysis ; Superoxide Dismutase ; genetics ; isolation & purification ; metabolism

In this study, a rapid and sensitive analytical method was developed for the determination of 10 major compounds (procyanidin B1, catechin, procyanidin B2, rutin, isoquercitrin, kaempferol-3-O-rutinoside, astragalin, quercitrin, quercetin, and kaempferol) in Tetrastigma hemsleyanum by using ultra-performance liquid chromatography coupled with triple-quadrupole tandem mass spectrometry (UPLC-MS/MS) in multiple-reaction monitoring (MRM) mode. UPLC-MS/MS assay with negative ion mode was performed on a Waters CORTECS C18 (2.1 mm x 100 mm, 1.6 μm) with the mobile phase consisting of acetonitrile (A) and 0.1% aqueous formic acid (B) in gradient elution at a flow rate of 0.25 mL · min(-1) and the column temperature was set at 45 °C. Under the optimized chromatographic conditions, good separation for 10 target compounds were obtained including chiral isomer procyanidins B1 and B2 were completely separated within 8.5 min. Satisfactory linearity was achieved with wide linear range and fine determination coefficient (r > 0.996 6), the overall recoveries were ranged from 95.44%-110.40% with the RSD ranging from 2.37%-8.69%. It is the first report about simultaneous analysis of 10 major flavonoids components in Tetrastigma hemsleyanum by using UPLC-MS/MS method, which affords highly sensitive, specific, speedy and efficient method for quality control of Tetrastigma hemsleyanum
Acetonitriles ; Chromatography, High Pressure Liquid ; Flavonoids ; chemistry ; Kaempferols ; Quercetin ; analogs & derivatives ; Rutin ; Tandem Mass Spectrometry ; Vitaceae ; chemistry

?AIM:To observe the values and changing rules of angle Kappa in corneal refractive surgery under light and dark conditions.?METHODS:Two hundred and thirty-four eyes of 118 patients for corneal refractive surgery were enrolled for this study.Pupil diameters and angle Kappa values under light and dark conditions were measured by Keratron Scout corneal topography.?RESULTS: There were significant differences in pupil diameters between light and dark conditions (P<0.01). More angle Kappa of both eyes distributed in the superior nasal quadrant under light conditions, and more angle Kappa distributed in the superior temporal quadrant under dark conditions.The differences of horizontal and vertical offsets of angle Kappa under two conditions were statistically significant (P<0.01).?CONCLUSION: The changes of pupil diameters in light and dark conditions could affect angle Kappa and then affect the accuracy of corneal refractive surgery centered on angle Kappa.

OBJECTIVETo investigate the effects of nucleoside analogues on hepatitis B virus (HBV) in hepatic lymph nodes of hepatitis B related liver transplantation recipients who were hepatitis B surface antigen (HBsAg) positive but negative for serum HBV DNA.

METHODSFrom June 2010 to March 2011, thirty-six cases of hepatitis B related liver transplantation recipients [32 males, 4 females, average age (54 ± 7) years] were divided into drug treatment group and non-drug treatment group according to the utility of nucleoside analogues. Drug treatment group was divided into two subgroups: drug treatment > 3 months group and drug treatment ≤ 3 months group. The hepatic lymph nodes in the hepatoduodenal ligament were taken during the operation of liver transplant. Using nested or semi-nested PCR, HBV DNA and the replicative form HBV cccDNA in hepatic lymph nodes were detected. Data were analyzed by Fisher's exact test.

RESULTSThe positive rate of HBV DNA: the difference was not statistically significant between drug treatment group (72.7%, 16/22) and non-drug treatment group (14/14) (P = 0.062), the difference was not statistically significant between drug treatment > 3 months group (10/14) and drug treatment ≤ 3 months group (6/8) in the subgroups of drug treatment group (P = 1.000). The positive rate of HBV cccDNA: drug treatment group (22.7%, 5/22) was significantly lower than the non-drug treatment (12/14) (P = 0.000), drug treatment > 3 months group (1/14) was significantly lower than drug treatment ≤ 3 months group (4/8) in the subgroups of drug treatment group (P = 0.039).

CONCLUSIONSHepatic lymph nodes maybe one of the extrahepatic HBV reservoirs. Treating with nucleoside analogues more than 3 months can significantly decrease the replication of HBV in hepatic lymph nodes of HBV associated liver transplantation recipients.

Adult ; Aged ; DNA, Viral ; analysis ; Female ; Hepatitis B ; drug therapy ; Hepatitis B virus ; drug effects ; physiology ; Humans ; Liver Transplantation ; Lymph Nodes ; virology ; Male ; Middle Aged ; Nucleosides ; therapeutic use ; Preoperative Care ; Virus Replication

A qualitative analytical method of liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry (HPLC-Q-TOF-MS) was developed for identification of multi-constituents and an analytical method was developed for simultaneously determining 4 major compounds (rutin, isoquercitrin, kaempferol-3-0-rutinoside, and astragalin) in Tetrastigma hemsleyanum Diels et Gilg. The HPLC-Q-TOF-MS assay was performed on a Welch Ultimate XB-C18 column (4.6 mm x 150 mm, 5 microm) with the mobile phase consisting of acetonitrile (A) and water containing 0.1% Formic acid (B) in gradient mode at a flow rate of 0.8 mL x min(-1). The column temperature was at 30 degrees C, and negative ion mode was used for TOF-MS. The UPLC-QqQ-MS assay was performed on a Waters CORTECS C18 (2.1 mm x 100 mm, 1.6 microm) with the mobile phase consisting of acetonitrile (A) and water containing 0.1% formic acid (B) in gradient mode at a flow rate of 0.25 mL x min(-1). The column temperature was at 45 degrees C, and MRM mode was used for QqQ-MS. Based on the retention time and MS spectra, 24 compounds were identified or tentatively characterized by comparing with reference substances or literatures. For quantitative the linear range of 4 detected compounds were good (r > 0.9966), and the overall recoveries ranged from 98.27% to 101.58%, with the RSD ranging from 3.15% to 5.88%. The results indicated that new approach conbined HPLC-Q-TOF-MS and UPLC-QqQ-MS was applicable in qualitative and quantitative quality control of Tetrastigma hemsleyanum.
Acetonitriles ; chemistry ; Chromatography, High Pressure Liquid ; methods ; Formates ; chemistry ; Tandem Mass Spectrometry ; methods ; Vitaceae ; chemistry ; Water ; chemistry

OBJECTIVETo improve computer-assisted imaging analysis for quantitatively measuring brain slice volume of rats and mice in comparison with conventional measuring methods,and to evaluate its usefulness in assessment of focal cerebral ischemia.

METHODSThe accurate volumes of rat and mouse brain slices were measured by weight and special gravity measuring. The areas of brain slices were measured by imaging analysis, then the slice volumes of right and left hemispheres were calculated by multiplying the adjusted thickness of the slices. In addition, the brain slice volumes of right and left hemispheres from focal cerebral ischemic mice were compared to assess ischemic injury using the imaging analysis.

RESULTArea measurement by computer-assisted imaging analysis was linear with different accurate areas (r=1.000). Slice volumes measured by imaging analysis correlated well with the accurate volumes measured by special gravity method, r=0.809 (n=45, P<0.001) in rats, and r=0.844 (n=74, P<0.001) in mice. The brain volumes in ischemic hemispheres were larger than in non-ischemic hemispheres in ischemic mice.

CONCLUSIONComputer-assisted imaging analysis can measure the brain slice volumes accurately and compare right and left hemisphere volumes quantitatively.

Animals ; Brain ; pathology ; Brain Ischemia ; pathology ; Image Processing, Computer-Assisted ; Male ; Mice ; Mice, Inbred ICR ; Rats ; Rats, Sprague-Dawley

AIMClinical studies stated that low molecular weight compounds (< 1.0 kd) extracted from the new born calf liver could effectively inhibit the proliferation of tumor cells. In this report, we observed inhibition effects and their regulative mechanisms of taurine, ornithine, carnosine on the proliferation of HL-60 cells.

METHODSThree active ingredients, i.e., taurine, ornithine and carnosine were separated by ion-exchange chromatographic column and identified from the low molecular weight filtrate of new born calf liver. MTT assay was used to test the survival rate of HL-60 cells and normal lymphocytes treated by the three ingredients. The various effects of the three compounds on HL-60 cells were respectively evaluated by agarose gel electrophoresis, ESR and immunohistochemical methods.

RESULTSThese compounds effectively inhibited the proliferation of HL-60 cells and induced apoptosis which was determined by apoptotic changes in morphology and nuclear DNA degradation. Whereas no inhibition effects on normal lymphocytes were observed. In addition, the results of ESR showed that the activity of oxygen radical within HL-60 cells treated with there compounds decreased to trace level. Furthermore, in the immunohistochemical experiments, we found that the level of p45/skp2 in HL-60 cells decreased while the level of p27/kip increased.

CONCLUSIONThe taurine, ornithine and carnosine compounds can selectively suppress tumor cells proliferation by regulating the level of cell cycle proteins.

Animals ; Animals, Newborn ; Apoptosis ; drug effects ; Carnosine ; pharmacology ; Cattle ; HL-60 Cells ; Humans ; Liver ; chemistry ; Ornithine ; pharmacology ; Taurine ; pharmacology

Adolescent ; Adult ; Dust ; analysis ; Female ; Health Status ; Humans ; Male ; Middle Aged ; Occupational Exposure ; analysis ; Pneumoconiosis ; prevention & control ; Sentinel Surveillance ; Surveys and Questionnaires ; Welding ; Young Adult

OBJECTIVETo investigate the toxicity of copper on MES23.5 dopaminergic cells and the probable mechanisms involved in this process.

METHODSMES23.5 dopaminergic cells were selected as our experimental model. [3-(4, 5-dimethylthiazol-2-yl)-2, 5 diphenyltetrazolium bromide] (MTT) assay was used to detect the influence of copper on the cell viability. The semi-quantitative reverse transcription polymerase chain reaction (RT-PCR), Western blotting and the high performance liquid chromatography-electrochemical detection (HPLC-ECD) have been used to detect the tyrosine hydroxlase (TH) mRNA and protein expression and the dopamine content in MES23.5 cells. The flow cytometry have been used to detect the changes of mitochondrial transmembrane potential.

RESULTS100 and 200 mumol/L copper had no effect on the MES23.5 cell viability, whereas 400 and 800 mumol/L of copper could decrease the cell viability (P < 0.01). Treating cells with 200 mumol/L copper for 24 h decreased the TH mRNA expression, the TH expression and the dopamine content compared with the control (P < 0.01, P < 0.01, P < 0.05, respectively). Besides, the mitochondrial transmembrane potential also decreased with the treatment of 200 mumol/L copper for 24 h (P < 0.01).

CONCLUSIONCopper could exert the toxic effects on MES23.5 dopaminergic cells and decrease the cell function. The dysfunction of mitochondria may be the mechanism of this toxicity effect.

Animals ; Cell Survival ; drug effects ; genetics ; Cells, Cultured ; Copper ; metabolism ; toxicity ; Dopamine ; biosynthesis ; Dose-Response Relationship, Drug ; Hybridomas ; Membrane Potential, Mitochondrial ; drug effects ; genetics ; Mice ; Mitochondria ; drug effects ; metabolism ; pathology ; Nerve Degeneration ; chemically induced ; metabolism ; physiopathology ; Neurons ; drug effects ; metabolism ; pathology ; Neurotoxins ; toxicity ; Oxidative Stress ; drug effects ; physiology ; Parkinson Disease ; etiology ; metabolism ; physiopathology ; RNA, Messenger ; drug effects ; metabolism ; Rats ; Tyrosine 3-Monooxygenase ; drug effects ; genetics ; metabolism