OBJECTIVETo investigate the apoptosis and proliferation effect of matrine on human medulloblastoma cell line D341 in vitro and the effect of the expression of the related caspase 3 and caspase 9 proteins.

METHODSThe D341 cells were cultivated successfully in vitro. Then the cells were divided into 5 groups according to the concentration of matrine (0.5 mg/mI group, 1.0 mg/ml group, 1.5 mg/ml group, 2.0 mg/ml group and the control group was 0 mg/ml). All the experiments were repeated three times. The cell morphologic and structure change was observed with the optical microscope and the transmission electron microscope. The proliferation of D341 cell was analyzed using Cell Counting Kit-8 assay. Apoptosis was detected by Annexin V-FITC/PI double staining. The expression of Caspase3 and Caspase9 was detected by Western blot.

RESULTSWith the effect of matrine, the proliferation inhibition rate gradually increased with drug concentrations increasing, and there was a significant difference (P < 0.01). The inhibitory effect of matrine on cell proliferation was different with the different treatment time, there was a significant difference between the 24 h to 72 h groups (P < 0.01). The apoptotic rate increased with matrine concentrations increasing. There were significant differences between the group of 0.5 mg/mI or 1.0 mg/mI to the group of 1.5 mg/mI or 2.0 mg/mI (P < 0.05). The apoptotic rate increased with the prolonged treatment time. There were significant differences between the group of 24 h or 48 h to the group of 72 h ( P < 0.05). With the increase of matrine concentration, the expression of Caspase 3 and Caspase 9 increased (P < 0.01).

CONCLUSIONMatrine induces the apoptosis, and inhibits the proliferation of human medulloblastoma D341 cells in vitro by up-regulation of the expression level of Caspase3, Caspase9.

Alkaloids ; pharmacology ; Apoptosis ; Caspase 3 ; metabolism ; Caspase 9 ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; Cerebellar Neoplasms ; metabolism ; pathology ; Humans ; Medulloblastoma ; metabolism ; pathology ; Quinolizines ; pharmacology ; Up-Regulation

Objective To explore the change of plasma orexin A concentration and the correlation between plasma orexin A concentration and energy intake in obese children.Methods Fasting plasma orexin A concentrations,boaly mass index(BMI) and energy intake were measured in 48 obese children(obese group) and 48 healthy children(healthy control group),and these indexes were compared,the correlation between plasma orexin A concentration and BMI,energy intake were analyzed.Results 1.The plasma orexin A concentration in obese group was significantly lower than that in healthy control group(F=5.632 P=0.008).2.In obsess group,there were negative correlation between plasma orexin A concentration and BMI(r=-0.478 P=0.012),positive correlation between plasma orexin A concentration and total energy intake(r=0.503 P=0.007),fat intake(r=0.659 P=0.006) and protein intake(r=0.381 P=0.026),and there was negative correlation between plasma orexin A concentration and carbohydrate(r=-0.316 P=0.022).3.In healthy control group,there were negative correlation between plasma orexin A concentration and BMI(r=-0.491 P=0.018),positive correlation between plasma orexin A concentration and total energy intake(r=0.512 P=0.009),fat intake(r=0.406 P=0.005),protein intake(r=0.313 P=0.020),and carbohydrate(r=0.432 P=0.025).Conclusions Orexin A may be involved in regulation of energy metabolism in obese children,and the interaction between plasma orexin A and energy intake might be different in different nutritional status in children.

A method of LC-QTOF/MS combining with chemical synthesis has been used to determine the structures of three novel bile acids from bear bile powder. Reference substances of tauroursodeoxycholic acid and taurochenodeoxycholic acid were oxidized by pyridinium chlorochromate. The products were analyzed by LC-QTOF/MS. Total 4 products including 3 isomers were predicted and identified according to the PCC oxidation theory and LC-QTOF/MS results. Bear bile powder samples were dissolved by methanol and analyzed by LC-QTOF/MS. Three unknown peaks were found and identified as 2-[[(3beta, 5beta)-3-hydroxy-7, 24-dioxocholan-24-yl]amino]-ethanesulfonic acid, 2-[[(5beta)-3, 7, 24-trioxocholan-24-yl]amino]-ethanesulfonic acid and 2-[[(5beta, 7beta)-7-hydroxy-3, 24-dioxocholan-24-yl]amino]-ethanesulfonic acid, separately, by matching their results with that of oxidation products above.
Animals ; Bile ; chemistry ; Bile Acids and Salts ; analysis ; chemistry ; Chromatography, Liquid ; methods ; Isomerism ; Molecular Structure ; Oxidation-Reduction ; Powders ; chemistry ; Spectrometry, Mass, Electrospray Ionization ; methods ; Taurochenodeoxycholic Acid ; chemistry ; Ursidae

Based on the literature study,the thoughts and possible therapeutic mechanism in treating male infertility from the perspective of spleen and kidney by regulating intestinal flora were explored.Disturbance of intestinal flora is one of the important factors leading to the development of male infertility,and the spleen and kidney have certain similarities to intestinal flora in the physiological function and pathological changes.Moreover,tonifying the kidney and strengthening the spleen can regulate the intestinal flora by fostering the growth of beneficial bacteria,inhibiting the reproduction of pathogenic bacteria,and protecting the barrier of the intestinal mucosa.Therefore,the possible therapeutic mechanisms in treating male infertility with the prescriptions for tonifying the kidney and strengthening the spleen to regulate intestinal flora are as follows:inhibiting the expression of inflammatory factors to reduce the inflammatory reaction of testicular tissues;improving the antioxidant capacity to alleviate the damage of spermatozoa caused by oxidative stress,and improving the bad mood to alleviate the impact of psychological stress on the reproductive system.The exploration of the thoughts for treating male infertility from the perspective of spleen and kidney by regulating intestinal flora may provide a new entry point for modern Chinese medicine clinical treatment of male infertility.

OBJECTIVETo establish and assess the Caco-2 cell in vitro absorption model.

METHODSCaco-2 cells were cultured on the millipore filters fixed in Snapwell transport chamber. The cell morphology, transepithelial electrical resistance, mannitol efflux rate and alkaline phosphatase activities were monitored during culture.

RESULTSAfter 21 days of in vitro culture, formation of tight junction was observed between the cells. The transepithelial electrical resistance reach a relatively stable value of 620-/+47 Omega.cm(2), the mannitol efflux rate was lower than 0.3%.h(-1).cm(-2), and the alkaline phosphatase activity in the apical side was significantly higher than that in the basolateral side.

CONCLUSIONThe established Caco-2 cell model shows similar morphology to intestinal epithelial cells with formation of polarity, and can be used as an in vitro model for absorption studies.

Caco-2 Cells ; Cell Culture Techniques ; methods ; Epithelial Cells ; cytology ; metabolism ; Humans ; Intestinal Absorption ; Intestines ; cytology

OBJECTIVETo establish a method for real-time recording the oxygen consumption of mice under normobaric hypoxia.

METHODSThe experimental apparatus was made up of animal container, filling water control system, electronic balance, hose, a computer with weight recording software, etc. The working principle was that the oxygen consumed by animal was replaced by water filling which was controlled by the pneumatic and hydraulic actuator. The water was weighted by an electronic balance and the weight signal was recorded into excel file at the same time. The accuracy and precision of the apparatus were detected by a 10 ml syringe. The oxygen consumption characteristics of 6 acute repetitive hypoxia mice and 6 normal mice were observed.

RESULTSThe P value for the paired t test was 1 and the CV value was 4%. The survival time and total oxygen consumption of acute repetitive hypoxia mice were both significantly increased compared to normal mice (P < 0.05), which were (58.8 +/- 6.8) min and (46.0 +/- 8.7) min respectively for the survival time and (85.1 +/- 8.5) ml and (73.6 +/- 5.4) ml respectively for total oxygen consumption.

CONCLUSIONThe hypoxia tolerance of the acute repetitive hypoxia mice can significantly improved by taking more oxygen in the animal cabin. The accuracy and precision of the apparatus are high and it can be used for the determination of oxygen consumption in hypoxia research.

Animals ; Hypoxia ; physiopathology ; Mice ; Monitoring, Physiologic ; instrumentation ; Oxygen Consumption ; physiology

OBJECTIVETo determine the effects of Chinese herbal monomers such as baicalin, berberine, and matrine on the androgen receptor (AR) mRNA expression in SZ95 sebocytes in vitro and to explore the possible mechanism of using traditional Chinese medicines to treat acne.

METHODSSZ95 sebocytes were cultured and then treated with berberine, baicalin, matrine, and 13-cis-retinoic acid for 24 hours. Reverse transcription polymerase chain reaction was applied to detect the changes of AR.

RESULTAR mRNA was downregulated by 13-cis-retinoic acid of 1 x 10(-5) mol/L and 1 x 10(-6) mol/L, and by baicalin of 1 x 10(-4) mol/L (P < 0.05).

CONCLUSION13-cis-retinoic acid and baicalin may exert antiandrogenitic action by inhibiting AR mRNA expression in human sebocytes.

Androgen Antagonists ; pharmacology ; Cell Line ; Down-Regulation ; Drugs, Chinese Herbal ; pharmacology ; Flavonoids ; pharmacology ; Humans ; RNA, Messenger ; biosynthesis ; Receptors, Androgen ; biosynthesis ; genetics ; Skin ; cytology

The study aimed to explore the changes of the immunocyte quantities and cytotoxicity in bone marrow, and how many hematopoietic progenitor cells retained after the bone marrow cells were activated by different combinations of various cytokines. Bone marrow cells were divided into four groups and were cultured in vitro: (1) Control: no cytokines were added. (2) IL-2 group: bone marrow cells were activated by IL-2. (3) CD3-AK group: bone marrow cells were activated by IL-2 and CD3-McAb. (4) CIK group: IFN-gamma, IL-1, IL-2 and CD3-McAb were added. CFU-GM assay and CD3 phenotype detection were performed before and after activating culture in all groups. The changes of cell quantities during culture and cytotoxicity of cultured cells were tested. CD3 positive cells markedly increased in both CD3-AK and CIK groups. The cell numbers and cytotoxicity of CD3-AK and CIK groups were higher than those of control or IL-2 group obviously after culture (P < 0.05). CFU-GM were decreased in all groups after culture and there had no significant difference among four groups. The combination of IL-2 and CD3-McAb not only stimulates the proliferation of marrow immunocytes and increases their cytotoxicity but retains enough hematopoietic progenitor cells as well. This combination of cytokines can be used to purge autologous bone marrow in vitro.

Ligustrazine, one of the major effective components of the Chinese traditional medicinal herb Ligusticum Chuanxiong Hort, has been reported plenty of biological activities, such as protect cardiovascular and cerebrovascular, neuroprotection and anti-tumor, et al. Because of its remarkable effects, studies on structural modification of ligustrazine have attracted much attention. Ligustrazine synthetic derivatives reported in recent decades are mainly derived from four primary intermediates (TMP-COOH, TMP-OH, TMP-NH2, HO-TMP-OH). To explore the neuroprotection activitiy of ligustrazine intermediates, six ligustrazine intermediates (2, 5, 8, 11, 12, 13) were synthesized and their protective effects against CoCl2-induced neurotoxicity in differentiated PC12 cells were studied. The target compounds were prepared via different chemical methods, including oxidation, substitution, esterification and amidation without changing the structure nucleus of ligustrazine. Compared with TMP (EC50 = 56.03 micromol x L(-1)), four compounds (2, 5, 12 and 13) exhibited higher activity (EC50 < 50 micromol x L(-1)) respectively, of which, compound 2 displayed the highest protective effect against the damaged PC12 cells (EC50 = 32.86 micromol x L(-1)), but target compounds 8 and 11 appeared lower activity (EC50 > 70 micromol x L(-1)). By structure-activity relationships analysis, the introduction of carboxyl, amino to the side chain of ligustrazine and appropriately increase the proportion of ligustrazine may contribute to enhance its neuroprotective activity, which provides a reference for the design, synthesis and activity screening of relevant series of ligustrazine derivatives in the future.
Animals ; Cell Differentiation ; drug effects ; Chemistry Techniques, Synthetic ; Cobalt ; toxicity ; Drugs, Chinese Herbal ; chemistry ; Neuroprotective Agents ; chemical synthesis ; chemistry ; pharmacology ; Neurotoxins ; toxicity ; PC12 Cells ; Pyrazines ; chemical synthesis ; chemistry ; pharmacology ; Rats

A rapid method of Liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-Q-TOF-MS) combined with pyridinium chlorochromate (PCC) oxidation has been developed to determine chemical structures of two novel isomers in bear bile powder. Derivatives of ursodeoxycholic acid (UDCA) and chenodeoxycholic acid (CDCA) were semi-synthesized by PCC oxidation, then were analyzed by LC-Q-TOF-MS. Separation was carried out on a reverse column with the mobile phase of acetonitrile-0.1% formic acid (45:55). The data of Q-TOF-MS was acquired by MS, MS/MS, positive and negative modes. Since UDCA and CDCA were stereochemical isomeric at an alcohol position, two oxidation products were same and have been confirmed by LC-Q-TOF-MS. Other two products were also determined based on the PCC oxidation theory. Samples of bear bile powder were dissolved by methanol and measured by LC-Q-TOF-MS. Two unknown peaks were found and identified by matching their retention times and accurate mass spectra ions with PCC oxidation productS. Finally, the structures of two new bile acids in bear bile powder were confirmed as 3alpha-hydroxy-7-oxo-5beta-cholanic acid, 7alpha-hydroxy-3-oxo-5beta-cholanic acid, respectively.
Animals ; Bile ; chemistry ; Chromatography, Liquid ; methods ; Isomerism ; Oxidation-Reduction ; Powders ; chemistry ; Pyridinium Compounds ; chemistry ; Tandem Mass Spectrometry ; methods ; Tissue Extracts ; chemistry ; Ursidae ; Ursodeoxycholic Acid ; chemistry