Aim To study the effect of halometasone in combination with scutellaria baicalensis georgi on the vitiligo mice induced by monobenzone. Methods 40% monobenzone cream was applied to induce vitiligo in C57BL/6 mice. Through the halometasone, halo-metasone and scutellaria baicalensis georgi combined with 40% monobenzone cream, the influence of halo-metasone and scutellaria baicalensis georgi on mice de-colorizing was studied. Hair decolorizing was observed with the naked eye, the skin decolorizing was observed by reflectance confocal microscopy ( RCM ) , and CD8 +T cell infiltration was tested with immunofluores-cence detection. The serum levels of interleukin-6(IL-6 ) and tumor necrosis factor-α( TNF-α) were deter-mined by enzyme linked immunosorbent assay ( ELISA) . Results Mice in model group showed de-pigmentation at both the monobenzone application part and non-application part. The halometasone group did not show significant therapeutic efficacy. In halometa-sone and scutellaria baicalensis georgi treatment group, there was less decolorization, the occurrence ratio, the scores of occurring time and size were lower compared with model group. There were fewer infiltrated lympho-cytes and CD8 +T cells. Halometasone and scutellaria baicalensis georgi group also showed that the serum levels of IL-6,TNF-α decreased. Conclusion Halo-metasone and scutellaria baicalensis georgi have thera-peutic effect on vitiligo mice induced by monobenzone.

Aim To observe the neuroprotective effect of sinomenine on hippocampal neurons from injury in-duced by oxygen glucose deprivation ( OGD ) and its underlying mechanism. Methods Hippocampal neu-rons were exposed to OGD for 4 h followed by 24 h re-oxygenation ( OGD-R) . Then cell viability was detec-ted by MTT. LDH release was detected by LDH kit. Cell apoptosis was detected by Hoechst stain. The ex-pression of Bax, Bcl-2 and caspase-3 were detected by Western blot. [ Ca2+] i of hippocampal neurons was detected by calcium imaging. Acid-sensing ion chan-nels ( ASICs ) current was detected by patch clamp technique. Results SN increased cell viability and reduced LDH release. SN also inhibited neuron apop-tosis and increased ratio of Bcl-2/Bax and reduced the expression of caspase-3 . OGD-induced increase of [ Ca2+] i was inhibited by SN. Furthermore, SN inhib-ited ASIC1 a current and also inhibited OGD induced increase of ASICs current in hippocampal neurons. Conclusion SN protects hippocampal neurons against OGD-R-induced injury. The inhibitory effect of SN on ASIC1 a and calcium overload was involved in the pro-tective effect of SN.

Aim To study the effects of ganoderma lu-cidum polysaccharides and metformin on myocardial fi-brosis of type 2 diabetic rats and its mechanism. Methods SD rats were fed with high fat diet for 4 weeks, and then were injected with streptozotocin (30mg·kg-1 ) to replicate type 2 diabetic model. The diabetic rats were randomized into normal control group,diabetes group, ganoderma lucidum polysaccha-rides group ( 600 mg · kg-1 ) , metformin group ( 600 mg·kg-1 ) , and combination group( ganoderma lucid-um polysaccharides 300 mg·kg-1 +metformin 300 mg ·kg-1 ) . After 12 weeks’ treatment,the levels of fast-ing serum glucose were determined and the extent of myocardial fibrosis was observed by Picro-sirius red staining. The contents of AGEs in serum were deter-mined by fluorescence spectrophotometer. The activities of CAT and GSH-Px in myocardium were detected. Im-munohistochemical method and Western blot were used to detect myocardial tissue AGEs and CTGF protein ex-pression. Results Combination group could repress patho-proceeding of myocardial fibrosis efficiently, im-prove the activity of CAT and GSH-Px in myocardium and lower the concentration of AGEs in serum, as well as reduce the expression of AGEs and CTGF in myo-cardium. Conclusions Ganoderma lucidum polysac-charides and metformin could prevent myocardial fibro-sis. The possible mechanism may be related to repress-ing oxidative stress of myocardium, lowering serum AGEs and down regulating AGEs and CTGF of myocar-dium.

Aim To determine the effects of high-fat meal and ABCB1 C3435 T polymorphism on the phar-macokinetics of nifedipine in the healthy Chinese sub-jects. Methods A total of 90 unrelated healthy Han subjects were divided into two groups:fasting group ( n=45 ) and high-fat meal group ( n=45 ) and then they received a single oral dose of 90 mg extended release tablet. Multiple blood samples were collected after 48 h, and the plasma concentrations of nifedipine were determined by high performance liquid chromatogra-phy- mass spectrometry ( LC-MS ) . PCR-restriction fragment length polymorphism ( RFLP ) analysis was performed to detect the C3435 T polymorphism in AB-CB1 gene. Results The numbers of individuals carry-ing C/C, C/T and T/T genotypes in fasting group were 13, 24 and 8, respectively. The mean area under the curve ( AUC0-∞) in subjects carrying T/T genotype distinctly increased by 46. 34% compared with subjects with C/C genotype, but there was no statistically sig-nificant difference (P=0. 066). In addition, pharma-cokinetic parameters including Tpeak, Cmax and AUC0-48 had statistically significant differences between fasting group and high-fat meal group ( all P<0. 05 ) . Con-clution High-fat meal can speed the absorption and increase the extent of nifedipine absorption; ABCB1 C3435 T polymorphism almost does not affect the phar-macokinetics of nifedipine.

Aim To study the role of TNF-α/NF-κB signaling in matrix metalloproteinase ( MMP)-9 expres-sion induced by lipopolysaccharide ( LPS ) in airway epithelial cells, and to investigate the effects of lenti-virus mediated RNAi targeting a disintegrin and metal-loproteinase 17 ( ADAM17 ) gene on MMP-9 expression induced by LPS. Methods The ADAM17 siRNA ex-pression vector was constructed, and packaged to re-combinant lentivirus in 293T cells. The HBE4-E6/E7 cells were pretreated for 30 min by NF-κB inhibitor ( PDTC) and a recombinant human TNFR p75-Fc fu-sion protein ( Etanercept) , or infected by the recombi-nant lentivirus for 72 h, and then stimulated for 24 h by LPS or TNF-α. The release of TNF-α was detected by ELISA. The mRNA and protein levels of MMP-9 were analyzed respectively by RT-PCR and Western blot. NF-κB activity was detected by electrophoretic mobility shift assay. Results LPS and TNF-α signifi-cantly increased MMP-9 mRNA and protein expressions and the activation of NF-κB in HBE4-E6/E7 cells ( P<0. 05 ) . Etanercept and PDTC significantly inhibited MMP-9 expression and the activation of NF-κB induced by LPS ( P<0. 05 ) . Lentivirus mediated RNAi targe-ting ADAM17 significantly decreased TNF-α produc-tion, inhibited MMP-9 mRNA and protein expressions and the activation of NF-κB induced by LPS in HBE4-E6/E7 cells ( P <0. 05 ) . Lentivirus mediated RNAi targeting ADAM17 did not inhibit MMP-9 mRNA and protein expressions and the activation of NF-κB in-duced by TNF-α ( P>0. 05 ) . And PDTC significantly inhibited MMP-9 mRNA and protein expressions and the activation of NF-κB induced by TNF-α ( P <0. 05 ) . Conclusions TNF-α/NF-κB signaling partic-ipates in the regulation of MMP-9 expression induced by LPS in airway epithelial cells, and lentivirus-media-ted RNAi targeting ADAM17 plays an important role in that signaling pathway upstream by regulating TNF-αrelease.

Aim To study the activation of Nrf2 in-duced by baicalein ( BAI ) , and its protection against carbon tetrachloride ( CCl4 ) , ethanol and acetamino-phen ( APAP )-induced hepatotoxicity. Methods A reporter gene assay was conducted in human normal liver L-02 cells to detect the activation of transcription factor Nrf2 induced by baicalein. APAP ( 10 mmol · L-1 ) , CCl4 (10 mmol·L-1 ) and Ethanol (100 mmol · L-1 ) were used to induce hepatotoxicity in L-02 cells. After the pre-incubation with Baicalein (0, 1, 10, 25, 50, 100 μmol·L-1 ) for 15 min, cells were administrated with or without those above hepatotoxins. 48 h later, cell viability was detected by 3-(4, 5-dim-ethylthiazol-2-yl ) 2 , 5-diphenyltetrazolium bromide (MTT) method. Results Baicalein (25, 50 μmol· L-1 ) induced the activation of Nrf2 ( P <0. 01 , P <0. 05) in the reporter gene assay. As compared with control, three hepatotoxins ( APAP, CCl4 , Ethanol ) all decreased cell viability ( P<0. 01 ) , and baicalein significantly reversed such decreases in a concentra-tion-dependent manner ( P<0. 01 ) . Conclusion Ba-icalein can induce the activation of transcription factor Nrf2 , which is probably one of the mechanisms con-tributing to the protection of baicalein against hepato-toxins (APAP, CCl4, Ethanol)-induced hepatotoxici-ty.

Aim To isolate cancer stem cells from human pan-creatic cancer cell line L3. 6pl and to identify their biological characteristics. Methods L3. 6pl cells were cultivated in com-mercial low adhesion plate with serum-free stem cell culture me-dium ( MEM/F12 1:1 ) supplemented with B27. The cancer stem cells reformed into floating spheres were isolated. The method of tumor sphere formation was used to isolate/enrich and characterize the cancer stem cells in pancreatic carcinoma cell line L3. 6pl. Cancer stem cell spheres were collected and sorted using magnetic cell sorting ( magnetic activated cell sorting, MACS) technology, with the cell surface markers of CD24 +CD44 + ESA+. Self-renewal and EMT-related oncogene expres-sion were measured with Western blot. Cancer stem cells differ-entiation potential and the expression of cancer stem cell related signs were checked with Immunofluorescence assay. To deter-mine tumorigenesis in vivo, Xenograft assay in NOD-SCID mice were performed respectively, then immunohistochemistry proto-oncogene c-Met and RON expression were also checked. West-ern blot was used to detect the changes of stemness relative tran-scriptional factors and epithelial markers expressed in spheres before and after differentiation. Drug resistance of pancreatic cancer stem cells to gemcitabine or paclitaxel was measured with MTT assay. Results CD24 + CD44 + ESA+ cells were signifi-cantly tumorigenic, and cultured in serum-free conditions to form spheroids, which had the characteristics of stem cells with self-renewal, EMT and drug-resistant capabilities, and had a posi-tive correlation with the c-Met, RON protein expression. Con-clusion Human pancreatic stem cells are successfully isolated, which provides a useful model for individualized therapy and e-valuation of the therapeutic efficacy for pancreatic cancer pa-tients.

Aim To develop a sensitive, rapid and ac-curate LC-MSn method for determination of midazolam/1′-hydroxymidazolam and their pharmacokinetic char-acteristics in rat brain. Methods SD rats received in-travenous injection of midazolam ( 5 mg · kg-1 ) via femoral vein, a probe drug of cytochrome P450 3A. The microdialysis ( MD ) samples in situ brain were collected every 8 mins at 2. 0μl·min-1 in 2. 4 hours. Analytes in brain dialysate were quantified by the pro-posed LC-MSn method. Gradient elution was performed on an Agilent Eclipse Plus-C18 column ( 2 . 1 × 50 mm, 3. 5 μm). The mobile phase consisted of 2 mmol ·L-1 ammonium acetate and acetonitrile. The analyte was detected using electrospray ionization ( ESI ) in multiple reaction monitoring ( MRM) modes. The reac-tion selected ions were 326 . 1/291 . 1 m/z for midazo-lam, 342. 1/324. 1 m/z for 1′-hydroxymidazolam and 285. 1/154. 0 m/z for diazepam as internal standard. Result The linear ranges of midazolam and 1′-hydroxymidazolam were 0 . 78~100 and 0 . 195 ~12 . 5μg·L-1 respectively. The lower limit of quantification was 0 . 2 μg · L-1 . The RSD of intra- and inter-batch precisions was less than 7 %. The RSD of accuracy was from -1 . 34 to -8 %. Conclusion This sensi-tive and rapid LC-MSn method is suitable for determi-nation of midazolam/1′-hydroxymidazolam in rat brain dialysate. MD combined with LC-MSn method may give assistance to deep and further studies of drug metabo-lism and CYP3A enzyme in brain.

Aim To develop a sensitive, specific and accurate method for the pharmacokinetic study of QO-58 ( a novel M channel opener ) in rats after intragas-tric ( ig) and intravenous ( iv) administration. Meth-ods QO-58 was administered at the doses of 25,50, 100 mg · kg-1 ( ig ) and at single dose of 100 mg · kg-1(iv), respectively. Blood samples were obtained at intervals after each administration. Plasma samples were deproteinized with acetonitrile after addition of in-ternal standard, and detected by RP-HPLC. The main parameters of pharmacokinetics were calculated by DAS2. 1. 1 software. Results The calibration curve in plasma was linear over the range of 0. 1 ~160 mg · L-1 in rat plasma, and the limit of detection ( LOD) was 0. 1 mg · L-1 . The intra-day and inter-day RSD was less than 20%. The recovery of QO-58 in rat plas-ma was 89. 56% ~101. 38%. The concentration-time curves of QO-58 in rat palsma were consistent with the two-compartment model after both oral and intravenous administration. The main pharmacokinetic parameters for QO-58 following oral administration with three doses (25, 50, 100 mg· kg-1 ) in rat were as follows:Cmax (mg·L-1):8.25,16.29,18.27;T12β(h): 8.24, 5. 01, 5. 92; AUC0-∞ ( g · min · L-1 ):261. 94, 189. 57,90. 65. Conclusion The developed method is simple and specific, and is suitable for preclinical pharmacokinetic studies of QO-58 .

Aim To study the transport of geniposide and geniposide in Zhizi Bopi Decoction in MDCK cell membrane model. Methods The safety concentration of geniposide and Zhizi Bopi Decoction in MDCK cells were determined by MTT assay. Then the MDCK cell membrane model was used to investigate the transport of drugs. Firstly, the effects of time, drug concentra-tion, P-gp inhibitor and EDTA on the absorption and transport of geniposide were studied systematically. Secondly, the differences were compared between the transport of the same concentration of geniposide as single compound and that in Zhizi Bopi Decoction in MDCK cell model. The drug concentration was deter-mined by high performance liquid chromatography ( HPLC) to calculate the apparent permeability coeffi-cient (Papp). Results Geniposide transport in MDCK cell monolayer was time and concentration dependent. P-gp inhibitors had no significant effect on its transport and the transport of geniposide was enhanced by ED-TA. The absorption Papp of different concentrations of geniposide in Zhizi Bopi Decoction were ( 8. 96 ± 0. 35 ) × 10 -7 cm · s-1 , ( 8. 95 ± 0. 38 ) × 10 -7 cm · s-1 and (9. 16 ± 0. 30) × 10 -7 cm·s-1, significantly higher than the absorption Papp of geniposide as single compound(5. 85 ± 0. 44) × 10 -7 cm·s-1, (6. 88 ± 0. 38) × 10 -7 cm·s-1 and (6. 31 ± 0. 19) × 10 -7 cm ·s-1 ( P<0. 05 ) . Conclusion The transport of ge-niposide in MDCK cell membrane model is passive transport and is not affected by P-gp. Geniposide may transport via the paracellular route. The Zhizi Bopi De-coction can increase the absorption of geniposide.