The study aims to establish a method for simultaneous determination of repaglinide and pravastatin sodium in rat plasma by LC-MS/MS and to study its pharmacokinetic interactions. Eighteen male SD rats were divided into repaglinide group, pravastatin sodium group and co-administration group. Blood samples were collected at different times after oral administration. Repaglinide and pravastatin sodium in rat plasma were separated by Agilent HC-C18 with the mobile phase consisting of methanol-0.1% formic acid (80 : 20). Detection and quantification were performed by using ESI-MS. The detector was operated in selected Reaction-monitoring mode at m/z 453.3-->230.1 for repaglinide, m/z 447.2-->327.4 for pravastatin sodium and m/z 285.1-->192.9 for diazepam as the internal standard. The calibration curve obtained was linear (R2>0.99) over the concentration range of 9.77-10,000 ng.mL-1 for repaglinide and 4.88-625 ng.mL-1 for pravastatin sodium. Compared with the single administration group, Cmax and AUC0-6h of repaglinide increased significantly (P<0.05) and tmax of pravastatin sodium prolonged (P<0.05) in co-administration group. The method is found to be simple, sensitive and accurate for determining the concentration of repaglinide and pravastatin sodium in rat plasma. There exists pharmacokinetic interactions in the co-administration of repaglinide and pravastatin sodium.
Administration, Oral ; Animals ; Carbamates ; administration & dosage ; blood ; pharmacokinetics ; Chromatography, High Pressure Liquid ; Drug Interactions ; Male ; Piperidines ; administration & dosage ; blood ; pharmacokinetics ; Pravastatin ; administration & dosage ; blood ; pharmacokinetics ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Reproducibility of Results ; Sensitivity and Specificity ; Spectrometry, Mass, Electrospray Ionization ; Tandem Mass Spectrometry

This study aims to investigate the change of plasma concentration of digoxin (DIG) in rats with ovariectomy. Twelve female SD rats were randomly assigned into ovariectomized group and sham group (n = 6). All rats plasma was collected after a single dose of 2 mg x kg(-1) DIG administrated orally, serum DIG concentration was determined by LC-MS/MS. The level of P-gp in the intestinal was analyzed by Western blotting. Pharmacokinetic calculations were performed on each individual using DAS 2.0 practical pharmacokinetic software. Compared with the sham group, C(max) of ovariectomized group decreased significantly (P < 0.01). There was no significant difference of AUC(0-t), and the level of P-gp was elevated in ovariectomized group. It was found that C(max) of DIG was significantly reduced after ovariectomy, and the change was associated with the decreased level of estrogen, which contributes to the increased level of P-gp.
ATP-Binding Cassette, Sub-Family B, Member 1 ; metabolism ; Animals ; Blotting, Western ; Chromatography, Liquid ; Digoxin ; blood ; pharmacokinetics ; Disease Models, Animal ; Estrogens ; blood ; Female ; Ovariectomy ; Rats ; Rats, Sprague-Dawley ; Tandem Mass Spectrometry

OBJECTIVEThe purpose of this study was to investigate the clinical significance of THY1 protein expression in epithelial ovarian cancer.

METHODSImmunohistochemistry (IHC) and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) staining were used to detect the protein expression of THY1, Ki67 and cell apoptosis in 76 epithelial ovarian cancers by tissue microarray. The correlation between THY1 expression and patients' clinical features was analyzed.

RESULTSOf the 76 epithelial ovarian cancer samples, 64 were informative for IHC and TUNEL assays and 42 (65.6%) among them showed down-regulated/loss expression of THY1 protein. A significant positive correlation of THY1 protein expression with clinical stage and distant metastasis was observed in this ovarian cancer cohort (P < 0.05). The more advanced the tumor stage, the more frequency of loss expression of THY1 protein. In addition, the mean positive rate of Ki67 staining in tumors with down-regulated/loss expression of THY1 was 33.7% +/- 3.5%, significantly higher than that in the tumors with normal expression of THY1 (17.3% +/- 6.1%, P = 0.0027). However, no significant correlation was observed between THY1 protein expression and tumor cell apoptosis as well as patients' survival in this series (P > 0.05).

CONCLUSIONDown-regulated/loss expression of THY1 protein in epithelial ovarian cancer is significantly correlated with cancer cell proliferation and metastasis in the epithelial ovarian cancer, and it may be used as one of the new molecular biomarkers to predict the disease progression in patients.

Adult ; Aged ; Apoptosis ; Cystadenocarcinoma, Mucinous ; metabolism ; pathology ; Cystadenocarcinoma, Serous ; metabolism ; pathology ; Down-Regulation ; Female ; Follow-Up Studies ; Gene Expression Regulation, Neoplastic ; Humans ; Ki-67 Antigen ; metabolism ; Middle Aged ; Neoplasm Metastasis ; Neoplasm Staging ; Ovarian Neoplasms ; metabolism ; pathology ; Survival Rate ; Thy-1 Antigens ; metabolism

Objective To establish a rapid,specific and sensitive TaqMan real-time fluorescence quantitative PCR assay for detection of murine polyomavirus ( MPyV) .Methods The specific primers and TaqMan probe were designed based on genome sequence of MPyV.The primers amplified a 69 bp fragment.After optimizing the reaction system and reaction condition, the standard curve was plotted by detecting recombinant plasmid standards.The specificity, sensitivity and reproducibility of this method were evaluated.In addition, samples of lungs, spleens and feces obtained from experimentally infected mice and 86 clinical samples were used to validate the efficacy of this real-time PCR assay.Results The specificity assay showed that this assay could specifically detect MPyV and the sensitivity for MPyV was about 100 copies/well.The coefficients of variation ( CV) of both intra-assay and inter-assay were less than 1.13%.All of the samples from experimentally infected mice were positive for MPyV and 3 out of 86 clinical samples were positive by this TaqMan-PCR detection with a positive rate of 3.5%.Conclusions The real-time fluorescence quantitative TaqMan-PCR assay established in this study has high specificity, sensitivity and stability.It can be used for clinical diagnosis, routine detection and epidemiological investigation of murine polyomavirus infections.

Objective: To improve the N biomarker in the amyloid/taueurodegeneration system by radiomics and study its value for predicting cognitive progression in individuals with mild cognitive impairment (MCI). Materials and Methods: A group of 147 healthy controls (HCs) (72 male; mean age ± standard deviation, 73.7 ± 6.3 years), 197 patients with MCI (114 male; 72.2 ± 7.1 years), and 128 patients with Alzheimer’s disease (AD) (74 male; 73.7 ± 8.4 years) were included. Optimal A, T, and N biomarkers for discriminating HC and AD were selected using receiver operating characteristic (ROC) curve analysis. A radiomics model containing comprehensive information of the whole cerebral cortex and deep nuclei was established to create a new N biomarker. Cerebrospinal fluid (CSF) biomarkers were evaluated to determine the optimal A or T biomarkers. All MCI patients were followed up until AD conversion or for at least 60 months. The predictive value of A, T, and the radiomics-based N biomarker for cognitive progression of MCI to AD were analyzed using Kaplan-Meier estimates and the log-rank test. Results: The radiomics-based N biomarker showed an ROC curve area of 0.998 for discriminating between AD and HC. CSF Aβ42 and p-tau proteins were identified as the optimal A and T biomarkers, respectively. For MCI patients on the Alzheimer’s continuum, isolated A+ was an indicator of cognitive stability, while abnormalities of T and N, separately or simultaneously, indicated a high risk of progression. For MCI patients with suspected non-Alzheimer’s disease pathophysiology, isolated T+ indicated cognitive stability, while the appearance of the radiomics-based N+ indicated a high risk of progression to AD. Conclusion We proposed a new radiomics-based improved N biomarker that could help identify patients with MCI who are at a higher risk for cognitive progression. In addition, we clarified the value of a single A/T/N biomarker for predicting the cognitive progression of MCI.

OBJECTIVETo investigate the cough-relieving, analgesic and antibiotic effects of durian shell extract (DSE) in relieving cough and its analgesic and antibiotic effects.

METHODSThe effect of DSE in relieving cough was assessed in mice challenged with ammonia and SO(2) to induce coughing. The analgesic and antibiotic effects of DSE in mice were evaluated by hot plate test and twisting reaction induced by acetic acid, and by minimal inhibitory concentration (MIC) and disc-agar diffusion tests, respectively.

RESULTSCompared with the control group, the mice treated with 300 and 900 mg/kg DSE showed significantly prolonged latency with decreased number of coughing induced by ammonia and SO(2), and the effect was dose-dependent. DSE markedly prolonged the latency and decreased the twisting number of the mice induced by acetic acid without affecting the pain threshold in hot plate test. DSE produced no significant inhibitory effects against Staphylococcus aureus, Staphylococcus epidermidis, or E. coli, and showed a week inhibition against Bacillus aeruginosus.

CONCLUSIONDSE shows obvious effect in relieving cough and produces better analgesic effect against chemical factor-induced pain than against physical agent-induced pain sensation. DSE has a moderate inhibitory effect against Bacillus aeruginosus.

Analgesics ; pharmacology ; Animals ; Anti-Bacterial Agents ; pharmacology ; Antitussive Agents ; pharmacology ; Bombacaceae ; chemistry ; Male ; Mice ; Plant Extracts ; pharmacology ; Random Allocation

OBJECTIVEITS2 of DNA barcoding was used to study genetic polymorphism of Platycodon grandiflorum.

METHODTotal genomic DNA was isolated from P. grandiflorum. PCR was used to amplified the region of internal transcribed spacer 2 (ITS2), and PCR products were sequenced. The sequences of ITS2 were analyzed and compared by Clustal. The intraspecies genetic distance was calculated based on Kimura 2-parameter model by using MEGA 5.05. The ITS2 sequence of Codonopsis pilosula was used as the outreach value for plants of the genus, and the phylogenic tree used constructed by Neighbor-Joining (NJ) method.

RESULTThe K2-P's genetic distance of all samples were ranged from 0 to 0.930. The K2-P's genetic distance of samples at the same area were ranged from 0 to 0.178. The K2-P's genetic distance of samples at different areas were ranged from 0.735 to 0.930. The analytical result showed that the degree of genetic variation were heavy in intraspecies of P. grandiflorum and significantly correlated with geographical location.

CONCLUSIONThe DNA barcoding of ITS2 can applied to study the intraspecific genetic diversity, it provides a reference for further development of DNA barcoding technology applications.

China ; DNA Barcoding, Taxonomic ; DNA, Plant ; genetics ; DNA, Ribosomal Spacer ; genetics ; Molecular Sequence Data ; Phylogeny ; Platycodon ; classification ; genetics ; Polymorphism, Genetic

This study is to explore the mechanism and effect of N, N'-di-(m-methylphenyl)-3, 6-dimethyl-1, 4-dihydro-1, 2, 4, 5-tetrazine-1, 4-dicarboamide (ZGDHu-1) on proliferation and apoptosis of A549 cells in vitro and on A549 xenograft tumor in nude mice. With different concentrations of ZGDHu-1 at different times were used to treat A549 cells in vitro. The proliferation was determined by living cell count, SRB assay and Brdu-ELISA. Cell apoptosis was determined by cell morphology, DNA agarose gel electrophoresis, DNA content, Annexin V/PI and Hoechst 33258 labeling method. The nude mice model of A549 xenograft tumor was established by subcutaneous inoculation. The suppression activity of ZGDHu-1 by intraperitoneal injection on xenograft mice model was detected. The expressions of bcl-2, bax and p53 gene and protein were analyzed by RT-PCR and flow cytometry. ZGDHu-1 can inhibit A549 cell proliferation viability within a certain range of treating time and does, and a majority of A549 cells were arrested in G2-M phase. The A549 cells apoptosis was confirmed by typical cell morphology, DNA fragment, Sub G1 phase, Hoechst 33258 and Annexin V/PI labeling method with a time and dose related manner. When the xenograft tumor mice model were treated with 10, 20 and 40 mg x kg(-1) ZGDHu-1 for 14 days, the tumor growth inhibition rate were 43.7%, 56.9% and 60.0%, respectively. The expression of bax, bax/bcl-2 and p53 gene and protein increased significantly and bcl-2 decreased slightly by the treatment of ZGDHu-1. ZGDHu-1 can significantly suppress the growth of A549 xenograft tumor in vivo and inhibited proliferation by inducing tumor cell apoptosis in vitro. The mechanism may associate with its up-regulation of bax and p53 during the apoptosis process.
Animals ; Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Dose-Response Relationship, Drug ; Female ; Flow Cytometry ; Heterocyclic Compounds, 1-Ring ; pharmacology ; Humans ; Lung Neoplasms ; metabolism ; pathology ; prevention & control ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Proto-Oncogene Proteins c-bcl-2 ; biosynthesis ; genetics ; RNA, Messenger ; biosynthesis ; genetics ; Random Allocation ; Reverse Transcriptase Polymerase Chain Reaction ; Tumor Suppressor Protein p53 ; biosynthesis ; genetics ; Xenograft Model Antitumor Assays ; bcl-2-Associated X Protein ; biosynthesis ; genetics

Objective To investigate the effect of bile acids on altered synthesis and transport in the pathogenesis of colitis-related cholestasis and the underlying mechanisms in rats.Methods Adult Wistar male rats were randomly divided into normal group and model group,ulcerative colitis(UC) model was established by administration 2,4,6-trinitro-benzenesulfonicacid in intro-colonic,while rats in normal group were colonic administered with saline.UC model was evaluated by calculating disease activity index,histopathological score and myeloperoxidase activity.Serum level of alkaline phosphatase (ALP),glutamyl transpeptidase (GGT),alanine aminotransferase (ALT) and aspartate aminotransferase(AST) were also determined.Moreover,the level of hepatic total bile acids,deoxycholic acid and lithocholic acid were determined by HPLC/MS.The hepatic protein expression of bile acids synthesis enzyme cholesterol 7α-hydroxylase(Cyp7a1) as well as bile acids export transporter multidrug resistance-associated protein 2 (Mrp2) were all investigated by Western-bolt.Results Compared to the normal group,disease activity index,histopathological score and myeloperoxidase activity in model group were all significantly increased to (4.51 ± 1.49),(1.36 ± 0.69) point,and (0.40 ± 0.07) U · mg-1,respectively (all P ＜ 0.05),suggesting the successful preparation of UC rat model.Compared to the normal group,the content of total bile acids and deoxycholic acid were all markedly increased to (85.50 ± 18.60) μmol · L-1 and (1.50 ± 0.68) ng · g-1 in model group,and the serum level of ALP and GGT were all significantly increased to (259.43 ±58.58) U · L-1and (1.50 ±0.68) U · L-1 in model group (all P ＜ 0.05),indicating the occurrence of cholestasis in model group of rats.Compared to the normal group,the protein expression of Cyp7a1 in the liver of model group was significantly increased to 0.72 ± 0.07 in gray value,but the protein expression of Mrp2 was markedly decreased to 0.66 ± 0.04 in gray value in model group (all P ＜ 0.05).Conclusion Acute colitis can induce intrahepatic cholestasis in rats,increased bile acids synthesis and decreased bile acids excretion may contribute to colitis-related cholestasis via up-regulating Cyp7a1 expression and down-regulating Mrp2 expression in the liver of colitis rats.

ObjectiveTo establish a mixed teaching mode characterized by online teaching and flipped class,and explore its application effect.Methods From February 2016 to July 2016,a total of 180 students from 6 classes of grade 2015 in Hangzhou Medical College were selected as the research object by cluster sampling method. According to random number table,students were divide into the control group(3 classes,n=90),and the observation group(3 classes,n=90). For the comprehensive skills and clinical thinking courses,traditional teaching mode was applied in the control group,while observation group adopted the mixed teaching mode which was combined the massive open online courses(MOOCs)and flipped class together. Teaching participation,teaching effect,teaching methods were evaluated to assess the implementation effect of two teaching methods one term after the implementation of mixed teaching model.Results The total person-time of online visits was 9564. The number of discussions and fullfiled tasks were 372 and 38. The total final exam score of students in the observation group was(91.75±5.40),which was higher than that in the control group(84. 57±5.32)(t=-2.683,P<0.05). The scores of evaluation of 8 aspects in the observation group were higher than those in the control group,such as urging to prepare,finding information actively, developing self-study ability,improving study enthusiasm,improving the class participation enthusiasm,being impressive for the mistake,assessing more objectively and overall evaluation(P<0.05).Conclusions Mixed teaching model could promote the autonomous learning and class participation of nursing students. It also can make the teaching appraisal more objective and fair. However,how to make the students love this mode and how to set up the examination ratio scientifically should be studied further.