Objective To assess the drug sensitivity of pancreatic cancer cells based on real-time cell analysis and provide a reference for individualized diagnosis and treatment of pancreatic cancer.Methods Three human pancreatic cancer cells lines SW1900, Capan-2 and PANC-1 were selected and treated with gemcitabine hydrochloride and tegafur gimeracil oteracil potassium capsules, respectively.After 24 hours of culture, the cells were treated with the two drugs in gradient concentration.The cell growth curves before and after the drug administration was monitored using a real-time cells analyzer and the growth inhibition rates (IC50) of the drugs of the pancreatic cancer cells were calculated.At the same time, the cells in the cell culture plate were treated with the drug, and acridine orange/ethidium bromide (AO/EB) staining and laser scanning confocal microscopy were performed to observe the changes of cells after the drug administration.Results 72 hours after the drug administration, IC50 values for the three cell lines were different.The IC50 values of gemcitabine hydrochloride for SW1900, Capan-2 and PANC-1 cells were 1.69 μmol/L, 10.05 μmol/L and 12.74 μmol/L, respectively.The IC50 values of tegafur capsule for SW1900, Capan-2 and PANC-1 cells were 180.29 μmol/L, 765.70 μmol/L and 95.57μmol/L, respectively.AO/EB staining confirmed the reliability of IC50.Conclusions SW1900 and Capan-2 cells can be used as the control for gemcitabine hydrochloride and tegafur gimeracil oteracil potassium capsules to establish cell models for drug screening in vitro, which provides a reference for the application of the technology in anticancer drugs screening.

Objective To develop a model that could roundly show the phenotypes of human alzheimer disease (AD), the triple-transgenic rat model harboring APP(Swe), PS1dE9, and TAU transgenes was established in view of the advantage of rat as an important animal model on the research of nerve system .Methods APPswe/PS1dE9/TAU triple transgenic rat AD rats were generated on a SD background by co-injecting rat pronuclei with two human genes driven by the mouse prion promoter:‘Swedish’ mutant human APP (APPsw) and exon 9 mutant human presenilin-1 (PS1dE9) and human microtubule-associated protein tau gene under the control of PDGF promoter .Transgene integration was confirmed by genotyping and expression levels were evaluated by western blot ( WB ) of brain homogenates .The pathological changes were detected by human Abeta, TAU and Phospho-PHF-TAU immunohistochemistry staining (IHC).The behavioral and cognitive changes were evaluated by Morris water maze .Results One transgenic rat lines with high human APP ( Swe ) , PS1dE9, and TAU transgenic expression was selected from three transgenic founders .Compared with the wild type rat , the transgenic rat showed significant learning and memory impairments in the Morris water maze at 6 months of age .The triple transgenic rat manifested hyperphosphorylated tau and obvious aggregation of amyloid -β( Aβ) in the brain cortex and hippocampus.Conclusion APPswe/PS1dE9/TAU triple transgenic rat AD model was established .The triple transgenic AD rat fills a critical need for a next-generation animal model to enable basic and translational AD research .

Objective To develop a more convenient and stable method for assessment of drug sensitivity of prostate cancer based on real-time cell analysis system as a reference for clinical treatment.Methods Human prostate cancer VCaP, DU145, PC-3, PC-3M-2B4 and PC-3M-IE8 cells were chosen to detect the sensitivity to three drugs, docetaxel, cabazitaxel and abiraterone acetate.Serial dilutions of the three drugs were used to treat the cell culture for 24 hours.The drug-induced effects on the cell lines after an incubation of 24 hours were recorded by the real-time cell analysis system to determine the half maximal inhibitory concentration (IC50).Results Docetaxel showd IC50 of 8.81 nmol/L, 11.61 nmol/L, 1.78 nmol/L, 1.44 nmol/L, 8.69 nmol/L for VCaP, DU145, PC-3, PC-3M-2B4, PC-3M-IE8 cells, respectively.Cabazitaxel showed IC50 of 3.73 nmol/L, 3.96 nmol/L, 10.41 nmol/L, 5.43 nmol/L, and 7.37 nmol/L, respectively, for the five cell lines.Abiraterone acetate showed IC50 of 8.34 μmol/L, 8.60 μmol/L, 24.20 μmol/L, 8.59 μmol/L, and 13.21 μmol/L for the five cell lines.Conclusions PC-3M-2B4 and DU145, VCaP and PC-3 cells can be used as control for docetaxel, cabazitaxel and abiraterone acetate to establish cell models for the drug screening in vitro and to provide reference for clinical applications.

Objective To obtain more physiological data of Leptin knockout SD rats available for the user , the long-term observation of fasting blood glucose and pathological phenotypes were performed .Methods The protein expression levels in liver tissues were determined by western blot .Body weight of Leptin knockout rats ( Leptin-/-) and littermate lean rats (Leptin+/+) were weighed up at 1,3,6,8 months of age.Fasting blood glucose of Leptin +/+rats and Leptin-/-rats at 1,3,6,8 months of age were measured using One Touch? brand blood glucose monitoring systems.Pathological changes of pancreas and livers of Leptin -/-rats were observed by the method of HE staining and Immunohistochemistry (IHC).Results Short null Lepin proteins were expressed in liver tissues from Leptin -/-rats. Leptin-/-rats become heavier than Leptin +/+rats since they were one month old .The body weight of Leptin -/-rats at 8 months of age was twice as heavy as Leptin +/+rats, female Leptin-/-rats weighing 884g, and male Leptin-/-rats weighing 1200g.Overt hyperglycemia was observed during the first month after birth .Compared with Leptin+/+female rats,the fasting blood glucose of Leptin -/-female rats was increased by 40%-26%from 1 to 6 months old. After that, blood glucose values decreased and eventually become nearly normal at 8 months of age.Pathological examination indicated that Leptin -/-rats at 8 months of age had a fatty liver , more pancreas islets with lager volume and more beta cells with increased insulin secretion .Conclusion Leptin-/-rat were characterized by obesity , fatty liver, islet cell hyperplasia and early hyperglycemia .

Objective To investigate the functional role of 25 microRNAs in breast cancer ,and to find new tumor suppressor microRNAs that may serve as specific targets of new gene therapies . Methods Twenty-five microRNAs expression vectors were constructed and stably transfected into mouse mammary tumor cells 4To7 by Lipofectamine2000. Cells were selected with G418 and sorted by Flow cytometry.The cells in logarithmic phase were collected and 2 ×105 cells/mouse was inoculated into BALB/c mice via tail vein .Lungs were harvested 14 days after tumor cell inoculation , and the number of metastasis foci was counted .Results Mice inoculated with mir-449a-expressing 4To7 cells via tail vein developed reduced lung metastases compared with mice inoculated with negative control cells .Mice inoculated with mir-1935-expressing 4To7 cells via tail vein developed increased lung metastases compared with mice inoculated with negative control cells .Other twenty-three microRNAs neither promoted nor inhibited lung metastases of breast cancer .Conclusions Two of twenty-five microRNA were identified to be associated with breast cancer metastasis .MiR-449a may play a tumour suppressor role in the regulation of migration and metastasis in breast cancer .miR-1935 transgenic over-expression promoted tumor growth and metastasis .

OBJECTIVE：To provide reference for the identification of Chebulae Fructus and Chebulae Fructus Immaturus . METHODS：UPLC method was adopted. The determination was performed on Waters Cortecs T 3 C18 column with mobile phase consisted of acetonitrile- 0.2％ phosphoric acid solution （gradient elution ）at the flow rate of 0.35 mL/min. The column temperature was 30 ℃，and the detection wavelength was set at 270 nm. The sample size was 1 μL. Using gallic acid as reference，UPLC fingerprints of 17 batches of Chebulae Fructus and 14 batches of Chebulae Fructus Immaturus were established and their similarity was evaluated by TCM Chromatographic Fingerprint Similarity Evaluation System （2012 edition）. By comparing substance control ， UV absorption spectrum and related literaturs ，common peaks were identified. PCA and PLS-DA were performed by using SPSS 20.0 and SIMCA 14.1 software. The contents of main difference components in Chebulae Fructus and Chebulae Fructus Immaturus were determined by above UPLC method and compared. RESULTS ：There were 8 common peaks in UPLC fingerprint of Chebulae Fructus and Chebulae Fructus Immaturus ，i.e. chebulic acid （peak 1），gallic acid （peak 2），punicalagin A （peak 3），punicalagin B （peak 4），corilagin（peak 6），chebulagic acid （peak 7）and chebulinic acid （peak 8）. The similarities of 17 batches of Chebulae Fructus were from 0.92 to 0.99，while 14 batches of Chebulae Fructus Immaturus were all above 0.99. The similarity of control fingerprint between Chebulae Fructus and Chebulae Fructus Immaturus was 0.909. PCA demonstrated the differences between Chebulae Fructus and Chebulae Fructus Immaturus . The results of PLS-DA were consistent with those of PCA ，and the variable importance in projection （VIP）values of peak 5，4，7，3 and 2 were above 1 in the PLS-DA model. In 31 batches of samples ，the contents of gallic acid （peak 2），punicalagin A（peak 3），punicalagin B （peak 4）and chebulagic acid （peak 7）were 2.63-10.31， 5.37-44.63，8.02-60.77，44.07-162.98 mg/g；RSDs were 40.14％， 47.91％ ，53.97％ ，36.22％（n＝31）. There was statistical significance in the differences of the mentioned 4 components between Chebulae Fructus and Chebulae Fructus Immaturus 719412818@qq.com （P＜0.05）. CONCLUSIONS ：There are significant differences between Chebulae Fructus and Chebulae Fructus Immaturus gallic acid ，punicalagin A ，punicalagin B and chebulagic acid are the main difference components for identification.