Abstract Objective:FSM-2117 and FS-5416 are two bivalent hybrid strains of S.flexneri and S.sonnei, constructed by this laboratory-The FS-5416 expresses four invassive plasmid antigens(IpaA、IpaB、IpaC and IpaD) and has a good contact heamolysis activity(CHA+ ), butFSM-2117 hasn' t. To observe the immunogenicity of Shigelk vaccines through three different mucosal administration rout, the changes of ASCin the spleen and Peyer's patch(PP) , mesenteric lymph nodes(MLN) lymphocytes are detected.Methods:BALB/c mice were divided intothree groups, 20 mice per group. Mice were immunized respectively with the Ipa+ or Ipa- vaccines(4 x 10~7 CFU) three times with an intervalof two weeks by intranasal、intragastric or intraintestinal administration routs. The spleen , PP , MLN lymphocytes were isolated of seventh dayat random after immunization. An BA-EIISASPOT was done to account the numbers of ASC.Results:The numbers of SIgA and SIgG ASC ofspleen , PP , MLN lymphocytes of intranasal and intraintestinal group were significantly increased . Significant difference were only observed inthe number of spleen lymphocytes SIgA and SIgG ASC of intranasal group between Ipa+ and Ipa- . Conclusion:Two bivalent Shigelk vaccinescan induce spleen and GALT immunity reaction by nasal or small intestinal mucosal in a low dosage compared with intragastric rout (about 1/20). Ipa can significantly increased the number of spleen lymphocytes SIgA and SIgG ASC of intranasal group.

Objective To construct the lentivirus carrying human ?-catenin-EGFP(enhanced green fluorescent protein)and observe its expression in human follicle stem cells.Methods The ?-catenin gene sequence was amplified by RT-PCR from extraction of total RNA of human vascular endothelial cells.TA cloning technique was utilized to acquire gene subcloned pUCm-T-?-catenin.After transformation reaction,candidate clone was further analyzed by PCR and gene sequencing.Then the plasmid was transfected into FT293 cells.After identification by Western blotting,the plasmid was transfected into FT293 cells again for packaging.Infection titer was monitored by green EGFP expression.The expression of ?-catenin-lentivirus in human follicle stem cells were observed under inverted fluorescence microscope.Results The ?-catenin gene was cloned into the lentivirus successfully.The high expression of green fluorescence protein in FT293 cell line was found under fluorescent microscope.Viral titer checked by real-time PCR was about 2.0?108 TU/mL.When the multiplicity of infection(MOI)was 10,the infection efficiency of ?-catenin-lentivirus in human follicle stem cells was nearly 80% after infection 48 h around.After 3 weeks of continuous observation,we found the infection efficiency still keeping in the range of 80%-90%.Conclusion The lentivirus expression vector for ?-catenin was successfully constructed.It can steadily infect human follicle stem cells and the infection efficiency is considerable high.

Objective To investigate the clinical characteristics and prognosis of patients with different molecular subtypes of breast cancer.Methods A cohort of 716 breast cancer patients which had clear immunohistochemical detection were investiged.Their molecular subtypes were categorized as Luminal A,Luminal B,HER-2 over-expressing and basal-like subtypes,based on detection of ER,PR,HER-2 expression,and the clinical data including characteristics,relapse,prognosis and prognostic factors of the patients with different subtypes of breast cancer were analyzed retrospectively.Results There were no significant differences among different molecular subtypes at the age,menopausal status,production times,clinical stage,and radiation therapy(P ＞0.05).There were significant differences among different molecular subtypes at axillary lymph node starus (x2 =17.208,P =0.001),turner size (x2 =20.528,P =0.000) and operation method (x2 =24.242,P =0.000) and chemotherapy regimens (x2 =10.711,P =0.013).Univariate and multivariate analyses showed that clinical stage (x2 =17.005,P =0.002),axillary lymph node status (x2 =11.267,P =0.000) and molecular typing(x2 =125.634,P =0.000) were independent prognostic factors affecting long-term survisal rate.Conclusion Breast cancer patients in different subtypes have different long-term survival rate.The patients in basal-like subtype have the worst long-term survival rate.Molecular subtypes may provide important information to predict the prognosis of breast cancer.

Culturing objective and curriculum system for rural order directed free medical students were proposed scientifically and detail measure in designing general course module , basic course module, clinical course module, preventive care course module and professional practice module were put forward based on the analysis of guiding ideology of training rural order directed free medical educational talents.

Objective To explore the effect of applying functional movement screen (FMS) in the sport injury risk assessment of Chinese rugby athletes.Methods Rrugby athletes of Chinese national and provincial teams were selected and their data were collected using the standard FMS test.Their non-impact injury of the lower limbs and trunk were tracked and recorded.FMS diagnostic value and diagnostic cut-off value were evaluated using the receiver operating characteristic curve (ROC) and odds ratio (OR).Results The area under curve of all,male and female rugby athletes was 0.780 (P=0.000),0.877 (P=0.001) and 0.7130 (P=0.013) respectively,with significant differences from AUC=0.5.FMS score optimal cut-off point of all,male and female rugby athletes corresponding to the maxi mum Youden index was 13.5,15.5 and 13.5 respectively.Among all,male and female rugby athletes,the injury rates of the positive group (with FMS score less than the corresponding optimal cutoff point) were significantly higher than that of the negative group (with FMS total score greater than a corresponding cut-off point)(P＜0.01),and OR value of the positive group was 25.85 (95％CI:3.34～200.23),25 (95％CI:2.36～264.80) and 14.22 (95％CI:1.76～114.92) respectively.Conclusion In China,the average FMS score of rugby athletes had a strong correlation with non-contact sport injury,which might become an assessment index of non-contact sport injury risks.There is a significant difference in FMS score optimal cut-off points between the male and female rugby athletes,with that of the female being 13.5 points and the male being 15.5 points.

Through pot experiments, the disease index and control efficiency of TS67 cell, the fermentation liquid of TS67 and the supernatant of TS67 separately act on Fusarium oxysporum and Bipolaris maydis was detected. Experiment results analysis with SPSS statistical analysis software indicated all treatments of TS67 could inhibit both of soybean root rot disease and corn southern leaf blight (P

To construct an immortalized rat astrocyte strain genetically modified by rat preprogalanin gene (IAST/GAL) and detect its galanin (GAL) expression and secretion, a cDNA fragment of rat GAL in plasmid of pBS KS(+)-GAL was inserted into eukaryotic expression vector pcDNA3.1 (+) by DNA recombinant technology, then the restriction enzyme digestion and DNA sequencing were carried out to evaluate the recombinant. The pcDNA3.1 (+)-GAL and pcDNA3.1 (+) construct were transfected into immortalized rat astrocyte strain (IAST) by lipofectamine and the population of cells which stably integrated the construct was selected with 600 microg/mL G418. Individual clones were screened and expanded into clonal cell strains. Detection of Neo gene was used to validate the success of the transfection. Immunocytochemical staining, RT-PCR and radioimmunoassay were used to detect the expression and secretion level of GAL. The recombinant had been successfully constructed by restriction enzyme digestion and DNA sequencing. Detection of Neo gene showed that the pcDNA3.1 (+)-GAL and pcDNA3.1 (+) have been successfully transfected into IAST. After selection by using G418, IAST/GAL and IAST/Neo cell strains were obtained. IAST/GAL, IAST/Neo and IAST were immunostained positively for GAL, but the GAL average optical density of IAST/GAL was significantly higher than that of IAST/Neo and IAST (P< 0.01). The level of GAL mRNA expression and the supernatant concentration of GAL in cultured IAST/GAL were significantly higher than those of IAST and IAST/Neo (P<0.01), but no significant differences were found between the IAST and IAST/Neo (P>0.05). It was concluded that IAST/GAL strain was constructed successfully and it might provide a basis for the further study of pain therapy.
Astrocytes/cytology ; Astrocytes/*metabolism ; Cell Line, Transformed ; Cells, Cultured ; Galanin/*biosynthesis ; Galanin/genetics ; Genetic Vectors ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics ; Recombinant Proteins/biosynthesis ; Recombinant Proteins/genetics ; Transfection

Objective To investigate the expressions and correlations of Kiss-1 and matrix metalloproteinases-9 (MMP9) proteins in the colorectal cancer (CRC) tissues of patients with synchronous colorectal liver metastasis (SCRLM),and the association with the clinicopathologic factors and prognosis of patients.Methods The retrospective case-control study was adopted.The clinicopathological data of 96 patients with SCRLM and 69 patients with CRC and no metastasis who were admitted to the Eastern Hepatobiliary Surgery Hospital of the Second Military Medical University from January 2000 to May 2013 were collected.The 96 CRC tissues and 50 adjacent normal tissues (distance from resection margin ≥ 5 cm) were collected from 96 patients with SCRLM,and 69 CRC tissues were collected from 69 patients with CRC and no metastasis.Expressions of Kiss-1 and MMP9 protein were detected by immunohistochemistry (IHC).The follow-up of outpatient examination and telephone interview was performed to detect survival of patients till August 2014.Comparison of count data and correlation between expressions of Kiss-1 or MMP9 protein and clinicopathological factors were analyzed by the chi-square test and Fisher exact probability.Survival curve was drawn using the Kaplan-Meier method,and survival analysis was done using the Log-rank test.Correlation analysis was done by the Pearson correlation.Results Expression of Kiss-1 protein was located in the cytoplasm of tissue cells.The positive expression rates of Kiss-1 protein in CRC tissues of patients with SCRLM and with CRC and no metastasis and in adjacent normal tissues were 24.0％ (23/96),43.5％ (30/69) and 52.0％ (26/50),respectively,with a significant difference among the 3 tissues (x2 =14.307,P ＜ 0.05) and no significant difference between CRC tissues of patients with CRC and no metastasis and patients with SCRLM (x2 =0.845,P ＞ 0.05).The positive expression rate of Kiss-1 protein in CRC tissues of patients with SCRLM was significantly different from that in CRC tissues of patients with CRC and no metastasis and in adjacent normal tissues (x2 =0.702,11.594,P ＜ 0.05).Expression of MMP9 protein was located in the cytoplasm of tissue cells.The positive expression rates of MMP9 protein in CRC tissues of patients with SCRLM and with CRC and no metastasis and in adjacent normal tissues were 67.7 ％ (65/96),62.3 ％ (43/69) and 36.0％ (18/50),respectively,with a significant difference among the 3 tissues (x2=14.203,P ＜0.05) and no significant difference between CRC tissues of patients with CRC and no metastasis and patients with SCRLM (x2=8.038,P ＞ 0.05).The positive expression rate of MMP9 protein in CRC tissues of patients with SCRLM was significantly different from that in CRC tissues of patients with CRC and no metastasis and in adjacent normal tissues (x2 =13.475,13.475,P ＜ 0.05).The positive expression rates of Kiss-1 protein in CRC tissues of patients with SCRLM were 66.7％,21.9％ and 17.6％ in the high-,mederate-and low-differentiated tumor,50.0％,28.6％ and 17.5％ in the muscular layer of tumor invasion,outside of serosa and serosal layer,44.0％ and 16.9％ in patients with and without lymph node metastasis,respectively,showing significant differences among the tumor differentiation degree,depth of tumor invasion and lymph node metastasis (x2=6.546,6.172,7.453,P ＜0.05).The positive expression rates of MMP9 protein in CRC tissues of patients with SCRLM were 25.0％,66.7％ and 76.2％ in the muscular layer of tumor invasion,outside of serosa and serosal layer,44.0％ and 76.1％ in patients with and without lymph node metastasis,respectively,showing significant differences between the depth of tumor invasion and lymph node metastasis (x2 =12.094,8.690,P ＜ 0.05).All the 96 patients with SCRLM were followed up for a median time of 68 months (range,12-176 months).The median overall survival time,median tumor-free survival time,5-year cumulative survival rate and 5-year tumor-free survival rate were 31 months,26 months,69.6％ and 26.1％ in patients with SCRLM and positive expression of Kiss-1 protein and 26 months,19 months,24.7％ and 12.3％ in patients with SCRLM and negative expression of Kiss-1 protein,respectively,showing significant differences between the overall survival and tumor-free survival (x2=16.578,14.436,P ＜ 0.05).The median overall survival time,median tumor-free survival time,5-year cumulative survival rate and 5-year tumor-free survival rate were 31 months,19 months,24.6％ and 12.3％ in patients with SCRLM and positive expression of MMP9 protein and 28 months,16 months,58.1％ and 22.6％ in patients with SCRLM and negative expression of MMP9 protein,respectively,showing significant differences between the overall survival and tumor-free survival (x2=14.073,8.532,P ＜0.05).Of 96 patients with SCRLM,there were 23 patients with positive expression of Kiss-1 protein (10 with positive expression of MMP9 protein and 13 with negative expression of MMP9 protein) and 73 with negative expression of Kiss-1 protein (55 with positive expression of MMP9 protein and 18 with negative expression of MMP9 protein),with a negative correlation between expressions of Kiss-1 protein and MMP9 protein (r =-0.291,P ＜ 0.05).Conclusions The reduced expression of Kiss-1 protein and elevated expression of MMP9 protein are closely associated with invasion and metastasis of CRC and prognosis of patients.A combination detection of Kiss-1 and MMP9 proteins is expected to become a marker for predicting the prognosis of patients with SCRLM based on the negative correlation between them.

Objective To assess the contribution of the portable chest computed radiography (CR) in evaluation of monitoring devices of the patients in the intensive care unit (ICU). Methods One hundred and sixty-two cases with 387 chest radiographs in the ICU were analysed retrospectively. The location of the catheters of monitoring devices and complications were observed.Results The malposition of the catheters was detected in 47 cases(16.9%),including the endotracheal (ET) tubes too deep at the position, the central venous catheters placed into the internal jugular veins,and the position of the thoracic drain tubes to be deep not enough causing the drain to fall.The complications after operation of monitoring devices were not common,including pneumothorax caused by ventilatory assistance,atelectasis and pneumonia caused by malposition of the ET tubes,totally in 11 cases.10 cases with cardiopulmonary abnormalities were discovered accidentally in all 162 cases(6.2%) when evaluation of monitoring devices.Conclusion Bedside chest CR not only can show the catheter position and the complications of the monitoring devices ,but also the cardiopulmonary abnormalities of patients in the ICU.

Objective To investigate the role of human estrogen receptor-related receptor(ERR) ?,a submember of orphan receptors,in the tumorigenesis of endometrial cancer.Methods Plasmid of pSG-ERR? was transfected into endometrial cancer cell lines HEC-1A,HEC-1B,and Ishikawa.Real-time quantitative RT-PCR and western blot were used to analyze the mRNA and protein expression of ERR? in endometrial cancer cell.Flow cytometry was used to analyze the cellular growth.Results Expressions of the ERR? were significantly increased in the endometrial cancer cells transfected with pSG-ERR? plasmid; expression of the ERR? mRNA in HEC-1A cell was 9644.4 copies/ng,HEC-1B:9835.3 copies/ng,and Ishikawa:8008.6 copies/ng(P