This article introduces the independent experiment and social investigation activities in the course of medication analy- sis set up for strengthening students' comprehensive abilities.These activities create a good study atmosphere for enhancing stu- dents' ability to do research and their humanistic qualities.

Acetaminophen ; metabolism ; Animals ; Cells, Cultured ; Drug Interactions ; Flavonoids ; metabolism ; Hepatocytes ; metabolism ; Humans

To establish single- and double-transfected transgenic cells stably expressing hMATE1, hMATE1 cDNA was cloned by RT-PCR from human cryopreserved kidney tissue, and subcloned into pcDNA3.1(+) plasmid by virtue of both HindIII and Kpn I restriction enzyme sites. Subsequently, the recombined pcDNA3.1(+)- hMATE1 plasmid was transfected into MDCK, MDCK-hOCT1 or MDCK-hOCT2 cells using Lipofectamine 2000 Reagent. After a 14-day-cultivation with hygromycin B at the concentration of 400 µg · mL(-1), all clones were screened with DAPI and MPP+ as substrates to identify the best candidate. The mRNA content of hMATE1, the cellular accumulation of metformin with or without cimetidine as inhibitor, or transportation of cimetidine was further valuated. The results showed that all of the three cell models over expressed hMATE1 mRNA. The cellular accumulation of metformin in MDCK-hMATE1 was 17.6 folds of the control cell, which was significantly inhibited by 100 µmol · L(-1) cimetidine. The transcellular transport parameter net efflux ratios of cimetidine across MDCK-hOCT1/hMATE1 and MDCK-hOCT2/hMATE1 monolayer were 17.5 and 3.65, respectively. In conclusion, cell models with good hMATE1 function have been established successfully, which can be applied to study the drug transport or drug-drug interaction involving hMATE1 alone or together with hOCT1/2 in vitro.
Animals ; Biological Transport ; Cimetidine ; pharmacology ; DNA, Complementary ; Dogs ; Drug Interactions ; Humans ; Madin Darby Canine Kidney Cells ; Metformin ; pharmacology ; Organic Cation Transport Proteins ; genetics ; metabolism ; Transfection

OBJECTIVETo observe the content variation of luteolin and luteolin-7-beta-D-glucoside in Chrysanthemum morifolium Ramat. (CMR) from different collection time.

METHODSRP-HPLC was used to analyze these two components in CMR collected in 2001 and 2002.

RESULTThe content of luteolin was significantly lower than that of luteolin-7-beta-D-glucoside. Furthermore, the former showed no marked changes during collection, while the latter did not varied markedly in early collection but decreased significantly in later collection.

CONCLUSIONThe content of luteolin-7-beta-D-glucoside reflects the quality of Chrysanthemum morifolium Ramat. more viably than that of luteolin.

Chromatography, High Pressure Liquid ; Chrysanthemum ; chemistry ; Flavonoids ; analysis ; Glucosides ; analysis ; Luteolin

AIMTo search for flavonoids which possess stronger vasorelaxation action.

METHODSFour quercetin glycosides (1a - d) were synthesized from quercetin in three steps i. e. selective protection of quercetin, condensation with corresponding acetyiglycosyl bromide, and then removal of the protecting group; Six flavone compounds (2a - f) were prepared from phloroglucinol according to the conventional methods; The structures of synthetic compounds were confirmed by IR, 1H NMR, 13C NMR and MS. Vasorelaxation action of ten synthetic quercetin derivatives (or analogues) and four natural flavonoids compounds were examined on the isolated rat thoracic aorta rings; Comparative octanol-water partition coefficients (logP) were measured using a reversed-phase HPLC method.

RESULTSMost of the tested flavonoids showed concentration dependent relaxation effects against PE-induced contractions of rat aortic rings. These compounds had stronger action with the augment of logP values.

CONCLUSIONCompound 3-bromo-5 ,7-dihydroxyflavone (2d) was identified to have the most potent vasodilating action. These compounds exert vasodilating effects that are related to the logP values. A structure-activity relationship of flavonoids was suggested.

Animals ; Aorta, Thoracic ; drug effects ; Dose-Response Relationship, Drug ; Flavonoids ; chemical synthesis ; chemistry ; pharmacology ; Male ; Molecular Conformation ; Molecular Structure ; Quercetin ; administration & dosage ; analogs & derivatives ; chemical synthesis ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Structure-Activity Relationship ; Vasodilation ; drug effects ; Vasodilator Agents ; administration & dosage ; chemical synthesis ; pharmacology

OBJECTIVETo determine the possible difference in vasodialtation effect of quercetin and rutin.

METHODSThe isolated rat thoracic aorta was treated with phenylephrine (PE), and the effects of quercetin and rutin on the preconstricted aorta rings with or without endothelium were determined by organ bath technique. Nitric oxide synthase inhibitor L-N(G)-nitroarginine methyl-ester (L-NAME), guanylyl cyclase inhibitor methylene blue, cyclooxygenase inhibitor indomethacin were used to explore the mechanism.

RESULTSQuercetin (10-160 micromol/L) caused vasorelaxation of aorta rings preconstricted with PE in endothelium-intact and denuded aorta rings in a dose-dependent manner. Rutin(10-160 micromol/L) caused dose-dependent vasorelaxation in endothelium-intact rings preconstricted with phenylephrine, but not in denuded aorta rings. The maximal response (Rmax) values calculated from vasorelaxation curves of quercetin and rutin were (77.20+/-6.11)% and (44.28+/-7.48)%, respectively. There was no difference between median effective concentration (EC(50)) values of quercetin and rutin. Pretreatment with L-NAME (0.1 mmol/L) abolished the vasorelaxation by rutin,but did not influence the vasodilating effect of quercetin in endothelium-intact rings. Pretreatment with methylene blue (10 mmol/L) canceled the vasorelaxation both by quercetin and rutin. Pretreatment with indomethacin (10 micromol/L) attenuated the vasodilatation of quercetin, but did not affect the vascular effect of rutin.

CONCLUSIONThe vasodilatation effect of quercetin is more potent than rutin. The vasodilatation effect of quercetin might be mediated by guanylyl cyclase and cyclooxygenase-dependent pathway, while the vasodilatation by rutin might be via nitric oxide-guanylyl cyclase pathway.

Animals ; Aorta, Thoracic ; drug effects ; Dose-Response Relationship, Drug ; Guanylate Cyclase ; metabolism ; In Vitro Techniques ; Male ; Nitric Oxide ; metabolism ; Phenylephrine ; pharmacology ; Prostaglandin-Endoperoxide Synthases ; metabolism ; Quercetin ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Rutin ; pharmacology ; Vasodilator Agents ; pharmacology

OBJECTIVETo determine the effects of Varglaucocalyx on c-fos gene expression during global myocardial ischemia-reperfusion.

METHODForty Wistar rats were divided into 5 groups: group N as control; group CN as ischemia-reperfusion control and group XH, XM and XL treated with Varglaucocalyx 5%, 1%, 0.5% respectively prior to ischemia-reperfusion. The isolated rat hearts were perfused in condition of constant temperature and pressure, and then the left ventricular myocardiums were extracted for use. The expression of c-fos protein was detected by immunochemical method. The expression of c-fos protein were quantified by using computer image analysis system.

RESULTCompared with the values of group N, protein expressions relative area of c-fos gene (PERA) were increased significantly in group CN, XH, XM, XL(P < 0.01), but decreased significantly in group XH, XM, XL, compared with those of group CN (P < 0.05). The PERA of c-fos gene in group XM, XL were significantly lower than in group XH (P < 0.01), and the PERA of c-fos gene in group XM were lower than in group XL(P < 0.05).

CONCLUSIONVarglaucocalyx can effectively depress the expression of c-fos gene in myocardium which may account for its protection against myocardial ischemia-reperfusion injury, and the middle and the low concentrations of Varglaucocalyx are more effective than the high concentrations.

Animals ; Cardiotonic Agents ; pharmacology ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Female ; Gene Expression Regulation ; drug effects ; Genes, fos ; Isodon ; chemistry ; Male ; Myocardial Reperfusion Injury ; etiology ; metabolism ; Myocardium ; metabolism ; Plants, Medicinal ; chemistry ; Proto-Oncogene Proteins c-fos ; biosynthesis ; Random Allocation ; Rats ; Rats, Wistar

OBJECTIVETo gain stable genetic modification of neural stem cells (NSC) that constitutively secrete brain-derived neurotropic factor (BDNF) and to explore the biological role on the survival and neurite outgrowth of cultured dorsal root ganglion (DRG) neurons.

METHODSBDNF gene fragment from human brain cDNA libraries was obtained by using PCR. With molecular cloning technique, the recombinant stem cell viral vector with report gene was constructed, which is that MSCV-BDNF-IRES(2)-EGFP vector could encode Polycistronic mRNA. Viral particle was packaged by PT67 cell line and infected neural stem cells (mouse clone: C17.2). After selection with cloning cylinder, the expression of BDNF was assessed by immunohistochemistry enzyme linked immunosorbent assay (ELISA) and reverse transcriptase polymerase chain reaction (RT-PCR). The effects of stable gene-modified neural stem cells on embryonic mouse DRG neurons were evaluated in a dual-chambered cocultivation system at 3th, 10th day.

RESULTSRT-PCR analysis demonstrated expression of mRNA for human-BDNF. ELISA confirmed the presence of secreted BDNF 24 h after transfection and showed that the level of BDNF production by NSC-BDNF transfected C17.2 was at a rate of (14.6 +/- 0.8) ng x d(-1) x 10(-6) cells even after 3 months. With immunohistochemical analysis, compared with the control, the longer neurite outgrowth of cultured DRG cells and the more survival neurons were observed in NSC-BDNF transfected cells group.

CONCLUSIONSBDNF gene could be stably expressed in C17.2 cell line by MSCV, and the BDNF gene-modified NSC markedly enhances the survival and neurite outgrowth of cultured DRG neurons.

Animals ; Brain-Derived Neurotrophic Factor ; biosynthesis ; genetics ; Cell Culture Techniques ; Cell Survival ; Cells, Cultured ; Coculture Techniques ; Ganglia, Spinal ; cytology ; Humans ; Mice ; Neurons ; cytology ; metabolism ; Rats ; Rats, Sprague-Dawley ; Stem Cells ; cytology ; metabolism ; Transfection

New Chemical Entities (NCEs) development is a systematic long-term project that involves multiple disciplines. The translation research will help to build an advanced R&D system from the basic laboratory research, preclinical studies and clinical evaluation to clinical application of drug, for the purpose of shortening the R&D cycle and accelerate the launch of new drugs. In new drug R&D and its clinical application, drug disposition (absorption, distribution, metabolism, excretion, ADME) properties are important criteria for assessing drug-likeness of candidates. ADME evaluation of NCEs plays an important role in the translation research throughout innovative drug R&D process. Therefore, ADME evaluation at the early stage of drug design and development will be helpful to improve the success rate and reduce costs, and further access to safe, effective drugs.
Absorption ; Biological Transport ; Drug Design ; Drug Evaluation, Preclinical ; Pharmaceutical Preparations ; chemistry ; metabolism ; Pharmacokinetics ; Tissue Distribution ; Translational Medical Research

OBJECTIVETo investigate the inhibitive and inductive effect of (-)-tetrahydropalmatine (THP) and (+)-THP on main CYP450 isoforms in mouse liver microsomes.

METHODSThe in vitro inhibitory effect was evaluated by incubating (-)-THP or(+)-THP with the probe substrates of main phase I metabolic enzymes in mouse liver microsomes, and the remaining substrates were determined by HPLC or LC-MS/MS method. Mice were administered with (-)-THP or(+)-THP at dosage of 240 mg/kg or 60 mg/kg by gastric lavage for successive 7 days, then the cocktail-LC-MS method was applied to assess the activities of main CYP450 isoforms in mouse liver microsomes.

RESULTThe IC(50) values of both (-)-THP and (+)-THP on isoforms studied were higher than 100 μmol/L except that IC(50) value of (+)-THP on CYP2C was 43.89 μmol/L, indicating weak inhibition of (-)-THP and (+)-THP on CYP1A2, CYP2D22, CYP2E1 and CYP3A11 in vitro. Compared with the vehicle group, the activities of CYP2D22, CYP2E1 and CYP3A11 were not increased significantly in (-)-THP and (+)-THP treatment groups, while the activities of CYP1A2 in 60 mg/kg and 240 mg/kg (-)-THP groups were 68.7% and 73.0% higher, than that of the vehicle group (P < 0.05, P < 0.01, respectively), the activity of CYP2C37 in 240 mg/kg (-)-THP treatment group was 80.4%, higher than that of the vehicle group (P < 0.05).

CONCLUSIONThere is negligible or weak inhibition on main CYP450 in mouse liver microsomes by (-)-THP and (+)-THP in vitro. (+)-THP does not induce main CYP450 in mouse liver microsomes while (-)-THP weakly induces CYP1A2 and CYP2C37.

Animals ; Berberine Alkaloids ; pharmacology ; Cytochrome P-450 Enzyme System ; drug effects ; metabolism ; Male ; Mice ; Mice, Inbred ICR ; Microsomes, Liver ; drug effects ; enzymology