Abstract: Objective To establish an animal model of BALB/c mice infected with Burkholderia pseudomallei through the nose (inhalation route), provides a reliable animal model for the follow-up studies on the virulence of melioidosis and the pathogenesis of acute melioidosis. Methods　The experiment was carried out through infecting with Burkholderia pseudomallei through the nose (inhalation route). The pathophysiological response, visceral pathological damage and bacterial colonization of the mice infected with Burkholderia pseudomallei were observed by gross anatomy, histopathology and tissue homogenate count, and the biological characteristics of the mouse model of acute melioidosis were analyzed accordingly. Then we compared the physiological responses in BALB/c mice between the Burkholderia pseudomallei and non-pathogenic Burkholderia thailandensis. Results　In the model of acute nasal infection with Burkholderia thailandensis, most death happened between the 3rd to 5th day after infection, 3×105-3×106 CFU was the suitable dose for acute fatal melioidosis model of BALB/c mice, and the medium lethal dose was about 3×104-3×105 CFU. Both gross anatomy and tissue HE staining showed that abscesses or necrosis were found in the lung, spleen and liver, especially in the spleen and lung, which was positively correlated with the challenge dose. Viable bacteria was isolated from the blood, lung, spleen and liver of Burkholderia pseudomallei-infected mice, and the bacteria account colonization was related to tissue specificity. The concentration of live bacteria isolated from in the blood was the highest [Log2 value: (10.28±0.34) CFU/mL], and the organ with the maximum quantity of bacteria was the lung [Log2 value: (7.54±2.11) CFU/total organ]. It has been reported that the biological effects of Burkholderia pseudomallei and its homologous non-pathogenic Burkholderia thailandensis were similar at the cellular level, like multi-nuclear giant cell formation and active intracellular replication, while it is still unclarrified in the differences of virulence in mice. In this study, it was proved that Burkholderia thailandensis was not fatal to mice even at a high dose (8×107CFU), or detected from mice infected with it via nasal. Conclusion　We successfully established a reliable BALB/c mouse model (acute lethal model) of melioidosis via nasal infection, described its biological characteristics, and identified the different biological responses between Burkholderia pseudomallei and its homologous non-pathogenic Burkholderia thailandensis in mice.

Objective To investigate the clinical effect of lentinan and thymopentin injection combined with PTP chemotherapy in oral squamous cell carcinoma patients, and to observe the effect of lentinan on serum tumor specific growth factor ( TSGF) , IgG, IgA and IgM and quality of life in patients with oral squamous cell carcinoma .Methods The clinical data of 80 cases of oral squamous cell carcinoma ( postoperative lymph node metastasis) confirmed by operation and pathology were analyzed retrospectively, and the diagnosis and treatment were analyzed.The control group were treated with PTP regimen, 80 ~120 mg/m2 of cisplatin, 50 ~80 mg/m2 of teniposide on the first day, the second to fourth day, once daily +pingyangmycin 4~6 mg/m2 , once daily, 3 to 12 days, 3 weeks for a course of treatment, a total of 6 courses.In the control group based on the use of thymopentin injection 20 mg and lentinan 2 mg+5% glucose solution 250 mL intravenous infusion, chemotherapy 2 d before the start, infusion 14 d, 3 weeks for a course of treatment, a total of 6 courses, for the observation group;two groups were 40 cases.The levels of serum TSGF, IgA, IgG, IgM and CD3 +, CD4 +, CD8 +, NK cells and TSGF in the two groups before and after treatment were measured.The quality of life of the patients was investigated by hospital-made questionnaires.The clinical efficacy and side effects were analyzed.Results TSGF levels in the two groups were significantly lower than that before treatment, and the TSGF level in the observation group was (39.1 ±4.9)U/mL, which was significantly lower than that in the control group (52.3 ±5.8) U/mL(P<0.05).After treatment, the TSGF level in observation group was lower than that in control group [(39.1 ±4.9) U/mL vs. (52.3 ±5.8) U/mL](P<0.05).The CD3 +, CD4 +, NK, IgG levels in observation group were higher than those in control group and the CD8 +level in observation group was lower than that in control group (P<0.05).The scores of physical symptoms, sleep quality, mental state and social affection levels in observation group were higher than those in control group(P<0.05).The total efficacy in observation group was lower than that in control group (77.5% vs.45.0%, P<0.05).The adverse reaction rate was 20.0% in the observation group, which was significantly lower than that in the control group (P<0.05).Conclusion The use of lentinan and thymopentin injection combined with PTP chemotherapy in the treatment of oral squamous cell carcinoma has higher clinical efficacy and less adverse reactions.

Multimedia computing technology teaching is a new kind teaching method. It conquers many shortcomings such as poor video, poor expression in traditional teaching of microbiology. But it can also strangle the improvising creation of different teachers in teaching and then lead it to be typical " computer teaching" . So we should obey the teaching discipline in making multimedia computing technology courseware. It can lead many sections part from the class teaching if just emphasize the full use of video and audio. We should make the purpose obviously, and make it as easy as possible in teaching, and we should also explore the fixed regularity to make the teaching and studying integrated perfectly.

OBJECTIVETo establish an in vitro model of mouse cochlear basilar membrane impairment using cisplatin, and observe the protective effect of methionine on the hair cells.

METHODSThe cochlear basilar membrane samples of thirty two Kunming mice were harvested on the 2nd day after birth and randomly divided into four groups. Each group had 16 samples. Overnight preincubation the cochlear organ followed by appropriate treatment respectively as follows: the serum-free culture medium, the serum-free culture medium with methionine and cisplatin, the cisplatinum-containing serum-free culture medium, and the methionine-containing serum-free culture medium. The protective effect of methionine for injury of cochlea hair cells induced by cisplatin was observed by myosin-VI immunofluorescence, light microscopy, laser confocal scanning microscope and hair cells counting.

RESULTSThe outer hair cells (OHC) and inner hair cells (IHC) of control group and methionine group were not damaged. The outer and inner hair cells of cisplatin group were damaged in various degree, and had remarkable difference compared with control group and methionine group (P < 0.05). The outer hair cells and inner hair cells of cisplatin + methionine group were damaged less than the cisplatin group with remarkable difference (t(IHC) = 3.929, t(OHC) = 8.582, P < 0.05).

CONCLUSIONSCisplatinum could damage the cochlear hair cells of the basal membrane in Kunming mice. Methionine might protect against cisplatin's damage on the cochlear hair cells.

Animals ; Cisplatin ; adverse effects ; pharmacology ; Hair Cells, Auditory ; drug effects ; In Vitro Techniques ; Methionine ; pharmacology ; Mice ; Mice, Inbred Strains

OBJECTIVETo establish a method integrating multi-targets for determining critical process parameters of the manufacturing process of traditional Chinese medicine.

METHODThe ethanol precipitation process of Danhong injection was taken as an application case of the method established. Fractional factorial design of experiments were conducted. Mathematical models relating seven process parameters to ten targets in the ethanol precipitation process were established. Then the sums of the absolute values of the regression coefficients in the models were used to evaluate the criticality of process parameters.

RESULTWater content in the concentrate, ethanol concentration and ethanol consumption were identified as the critical process parameters.

CONCLUSIONThe method established can integrate multi-targets effectively for the evaluation of critical process parameters.

Chemical Precipitation ; Chemistry, Pharmaceutical ; methods ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Salvia miltiorrhiza ; chemistry

In this work, a feedforward control strategy basing on the concept of quality by design was established for the manufacturing process of traditional Chinese medicine to reduce the impact of the quality variation of raw materials on drug. In the research, the ethanol precipitation process of Danhong injection was taken as an application case of the method established. Box-Behnken design of experiments was conducted. Mathematical models relating the attributes of the concentrate, the process parameters and the quality of the supernatants produced were established. Then an optimization model for calculating the best process parameters basing on the attributes of the concentrate was built. The quality of the supernatants produced by ethanol precipitation with optimized and non-optimized process parameters were compared. The results showed that using the feedforward control strategy for process parameters optimization can control the quality of the supernatants effectively. The feedforward control strategy proposed can enhance the batch-to-batch consistency of the supernatants produced by ethanol precipitation.
Chemical Precipitation ; Chemistry, Pharmaceutical ; methods ; standards ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Quality Improvement ; Salvia miltiorrhiza ; chemistry

Extraction process trajectory of Danhong injection by near-infrared (NIR) spectroscopy in conjunction with multivariate data analysis techniques was developed for in-line monitoring of extraction process. To capture the variation of batch process, the use of score, Hotelling T2 and DModX control charts was investigated for real-time monitoring of extraction process. Various abnormal behaviors of the test batches were detected in time by comparing the extraction process trajectory. It was concluded that the process trajectory for in-line quality control based on NIR spectroscopy was a feasible technology tool of the total process quality control during traditional Chinese medicine (TCM) manufacturing process.
Chemistry, Pharmaceutical ; methods ; standards ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Online Systems ; Quality Control ; Spectroscopy, Near-Infrared ; methods

OBJECTIVETo establish a rapid quantitative analysis method for the quality control of Danhong injection extraction using near-infrared (NIR) spectroscopy.

METHODOnline collecting the NIR spectra during the mixed extraction process of Salvia miltiorrhiza and Carthamus tinctorius, partial least squares regression (PLSR) models were developed for the quality indicators rosmarinic acid (RA), salvia acid B (SaB), lithospermic acid (LA), hydroxysafflor yellow A (HSYA) and solid content (SSC), with HPLC and weight-loss method as reference methods.

RESULTThe correlation coefficients of the cross validation for RA, SaB, LA, HSYA and SSC were 0.909 3, 0.915 2, 0.901 9, 0.747 7 and 0.931 4, respectively. And the root mean square errors of cross validation (RMSECV) were 0.012 1, 0.251, 0.017 7, 0.038 1 g x L(-1) and 0.359%, respectively.

CONCLUSIONIn this study, NIR spectroscopy was successfully applied to achieve the real-time determination of the contents of RA, SaB, LA and SSC, while the performance of the HSYA calibration model needed to be improved.

Carthamus tinctorius ; chemistry ; Chemistry, Pharmaceutical ; methods ; standards ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; analysis ; isolation & purification ; Online Systems ; Salvia miltiorrhiza ; chemistry ; Spectroscopy, Near-Infrared ; methods

BACKGROUNDTo investigate the dynamics of amyloid fiber formation of yeast (Saccharomyces cerevisiae) prion protein Sup35NM under the native condition to provide materials and clues for the elucidation of amyloid fiber formation.

METHODSThe Sup35NM gene was cloned and expressed in E. coli. The recombinant Sup35NM protein was purified under denaturing conditions through Nickel-Sepharose chromatography. Aliquots were removed at designated time points for transmission electron microscopy (TEM), circular dichroism (CD) spectra, protease K resistance assay, as well as thioflavin T (ThT) binding assay.

RESULTSThe Sup35NM expressed and purified under denaturing conditions. The morphological alteration of the Sup35NM in PBS (pH7.4) during the protein aggregation and amyloid fiber formation was visualized by TEM. The CD assay showed that the course of amyloid fiber formation underwent a conformational shift from alpha-helix to beta-sheet. The fibers had higher capacity of resistance to protease K digestion compared to the monomers. ThT fluorescence assay displayed a rapid growth phase before reaching a final equilibrium phase during the fiber formation, and the higher concentration of Sup35NM could greatly accelerate the fiber formation in vitro.

CONCLUSIONYeast prion protein Sup35NM forms amyloid readily under native conditions in vitro. The dynamics of Sup35NM amyloid formation may provide supporting evidences for the nucleating polymerization models of amyloid fiber formation.

Amyloid beta-Peptides ; genetics ; metabolism ; ultrastructure ; Electrophoresis, Polyacrylamide Gel ; Endopeptidase K ; metabolism ; Kinetics ; Microscopy, Electron ; Peptide Termination Factors ; Prions ; genetics ; metabolism ; ultrastructure ; Protein Binding ; Recombinant Proteins ; metabolism ; ultrastructure ; Saccharomyces cerevisiae ; genetics ; metabolism ; Saccharomyces cerevisiae Proteins ; genetics ; metabolism ; ultrastructure ; Thiazoles ; metabolism

Objective To determine factors affecting postoperative survival of elderly patients with hepatocellular carcinoma (HCC). Methods A retrospective analysis of consecutive 54 elderly patients undergoing hepatectomy for hepatocellular carcinoma from Jan 1995 to Dec 2002 was performed. The data were analyzed by Kaplan-Meier method and Cox regression. Results Univariate analysis and Cox regression showed Child Pugh grading, vessel invasion, satellite nodule formation, Edmondson Steiner grading, intrahepatic recurrence and distant metastasis all related to postoperative overall survival or disease-free survival of the elderly with hepatocellular carcinoma (all P