UNLABELLEDWith different ratios, Omnipague mixed with cod liver oil and Meglumine Diatrizoate mixed with cod liver oil are compared with the stardard marker under the scanning of CT and MR in order to find the better ratio and materials for the marker. The experimental results show that the performances of both Omnipague mixed with cod liver oil and Meglumine Diatrizoate mixed with cod liver oil are better than the stardard marker.

CONCLUSIONOmnipague mixed with cod liver oil and Meglumine Diatrizoate mixed with cod liver oil can be used in making marker for neuronavigation system.

Biomarkers ; Cod Liver Oil ; Diatrizoate Meglumine ; Humans ; Iohexol ; Magnetic Resonance Imaging ; methods ; Neuronavigation ; instrumentation ; methods ; Reference Values ; Tomography, X-Ray Computed ; methods

AIMTo prepare insulin powder for inhalation by spray-drying technology, determine the deposition of the insulin powder formulation in vitro and preliminarily investigate hypoglycemic response of the dry powder with/without absorption promoters.

METHODSThe depositions of the insulin powder for inhalation were determined by the China Pharmacopoeia 2000 version addenda XH and hypoglycemic effects were evaluated by testing serum glucose with glucose oxidase-peroxidase (GOD-PAP) method.

RESULTSThe depositions of the spray-dried insulin powder for inhalation were more than 40% under various humidity and their changes were not significant when air flow was no less than 18 L x min(-1). The coadministration of insulin with 8 mmol x L(-1)/dose sodium taurocholate [PA = 59.91%, Cnadir = (33 +/- 6) %] and 10 mmol x L(-1)/dose sodium deoxycholate [PA = 47.46% , Cnadir = (32 +/- 7)%] induced a significantly greater decline in blood glucose levels, while coadministration with 1% sodium caprylate, 1% sodium dodecyl sulfate, 250 microg/dose lecithin, 10 mmol x L(-1)/dose EDTA appeared to have no significant effect (P > 0.05).

CONCLUSIONInsulin powder for inhalation was relatively stable under various humidity conditions and different flow current. The use of 8 mmol x L(-1)/dose sodium taurocholate and 10 mmol x L(-1)/dose sodium deoxycholate could be able to potentially improve the bioavailability of insulin by pulmonary route.

Administration, Inhalation ; Animals ; Biological Availability ; Blood Glucose ; metabolism ; Deoxycholic Acid ; pharmacology ; Drug Synergism ; Female ; Humidity ; Hypoglycemic Agents ; administration & dosage ; pharmacology ; Inhalation ; Insulin ; administration & dosage ; pharmacology ; Male ; Powders ; Rats ; Rats, Sprague-Dawley ; Taurocholic Acid ; pharmacology

15-hydroxyeicosatetraenoic acid (15-HETE) plays an important role in hypoxia-induced pulmonary vasoconstriction. Release of nitric oxide (NO) is apparently decreased and activity of endothelial nitric oxide synthase (eNOS) is impaired in chronic hypoxia. However, little is known whether 15-HETE contributes to eNOS/NO pathway in the constriction induced by 15-HETE. We examined the response of rat pulmonary artery (PA) rings to 15-HETE, the production of NO, total eNOS expression and the phosphorylation of eNOS in bovine pulmonary artery endothelial cells (BPAECs) stimulated by 15-HETE. Rat PA rings were divided into three groups: endothelium intact group, endothelium denuded group, and nitro-L-arginine methyl ester (L-NAME, 0.1 mmol/L, an inhibitor of eNOS) group. Constrictions to 15-HETE were significantly enhanced in endothelium denuded group and L-NAME group (both P< 0.05 vs endothelium intact group, n= 9); BPAECs were incubated in different conditions to test nitrite production by Greiss method. Nitrite production was significantly reduced by 1 mumol/L 15-HETE (P<0.05), and increased by the lipoxygenase inhibitors, 10 mumol/L cinnamyl 3,4- dihydroxy-[alpha] -cyanocinnamate (CDC, P< 0.05) and 0.1 mmol/L nordihydroguiairetic acid (NDGA, P< 0.01 ); Western blot analysis of extracts from BPAECs incubated with 15-HETE in different time was carried out to test total eNOS expression, and the expression was changed unobviously. Immunoprecipitation (IP) and Western blot analysis of cell extracts from BPAECs treated with 2 mumol/L 15-HETE in different length of time were accomplished, using phospo-eNOS-threonine 495 (Thr495, an inhibitory site) antibody for IP, and eNOS or 15-lipoxygenase (15-LO) antibodies for Western blot. 15-HETE depressed eNOS activity by increasing the levels of phospho-eNOS-Thr 495. The data suggest that eNOS/NO pathway is involved in PA constrictions induced by 15-HETE and that 15-HETE depresses eNOS activity by phosphorylation in Thr495 site. The protein interaction between phospho-eNOS (Thr495) and 15-LO is discovered for the first time.
Animals ; Cattle ; Down-Regulation ; drug effects ; Endothelium, Vascular ; cytology ; drug effects ; enzymology ; Hydroxyeicosatetraenoic Acids ; pharmacology ; In Vitro Techniques ; Male ; Nitric Oxide Synthase Type III ; metabolism ; Pulmonary Artery ; cytology ; enzymology ; physiology ; Rats ; Rats, Wistar

AIMTo determine the protective effect of recombinant human interleukin-2 (rhIL-2) in inhalant form on experimental respiratory tract infection with Klebsiella pneumoniae in mice.

METHODSMice were infected with the method of nasal intubation drip. During infection, mice were given rhIL-2 by sc injection and the method of nasal intubation drip. There were normal group, vehicle group, model group, rhIL-2 groups and gentamicin group. In the end, the pathological changes in the lung were observed. The survival time and the mortality within a week of each group were recorded. The total protein content, the albumin content, the activity of alkaline phosphatase and the activity of lactic dehydrogenase of broncho-alveolar lavage fluid (BALF) were dertermined and compared.

RESULTSSymptoms of Klebsiella pneumoniae were remarkably relieved because of rhIL-2 administration. The total protein content, the albumin content, the activity of alkaline phosphatase and the activity of lactic dehydrogenase of BALF were less than those in the vehicle group and the model group.

CONCLUSIONInhalation of rhIL-2 can alleviate the pathological changes in the lung after infection. At the same dose, it could be seen that the effect of rhIL-2 in inhalant form was better than that of the injection.

Administration, Inhalation ; Animals ; Bronchoalveolar Lavage Fluid ; chemistry ; Female ; Interleukin-2 ; administration & dosage ; pharmacology ; Klebsiella Infections ; drug therapy ; metabolism ; pathology ; Klebsiella pneumoniae ; Lung ; pathology ; Mice ; Mice, Inbred ICR ; Recombinant Proteins ; administration & dosage ; pharmacology

Objective To examine the effect of niclosamide on thyroid endocrine disruption in larval zebrafish. Methods Zebrafish embryos (2 hours post-fertilization) were exposed to niclosamide at concentrations of 0, 5, 10, 20, 40 μg/L and 80 μg/L until 120 hours post-fertilization, and the body weight, hatching rate, malformation rate and survival rate of zebrafish embryos/larvae were measured. In addition, the triiodothyronine (T3) and thyroxin (T4) activities were determined in zebrafish, and the expression of tshβ and ttr genes that were associated with the regulation of thyroid hormones was quantified using a quantitative real-time PCR (qPCR) assay. Results Following exposure to niclosamide, there was no concentration-dependent hatching rate (F = 0.947, P = 0.924) or body weight of larval zebrafish (F = 1.042, P = 0.409); however, there were concentration-dependent survival rate (F = 9.309, P = 0.005) and malformation rate (F = 14.900, P = 0.001). As compared to controls, exposure to niclosamide at concentrations of 40 μg/L and 80 μg/L resulted in a significant reduction in the survival rate (both P values < 0.05), and a marked rise in the malformation rate of larval zebrafish (both P values < 0.05). In addition, the T4 activity increased (R² = 0.927, F = 6.858, P = 0.003) and T3 activity decreased (R² = 0.925, F = 8.212, P = 0.001) in larval zebrafish with the concentration of niclosamide. qPCR assay determined up-regulation of tshβ gene expression (R² = 0.840, F = 9.032, P = 0.002) and down-regulation of ttr gene expression (R² = 0.952, F = 9.130, P = 0.002). Conclusions Niclosamide exposure at environmental related concentrations may cause thyroid endocrine disruption of larval zebrafish.

Objective To investigate the effects of different dialysates on expression of protein kinase C-? (PKC?) and apoptosis of U937 cell line. Methods Different dialysates were added into culture fluid with U937 cell line at exponential phase of growth, and groups were divided: fluid A+fluid B group (dialysate A+dialysate B), fluid A+fluid B+rottlerin (PKC? specific inhibitor)group, fluid A+powder B group (dialysate A+powder B) and fluid A+powder B + rottlerin group. Besides, blank control group and normal control group were established. Cells were harvested 24 h and 48 h after treatment, morphological changes were observed by Hoechst33258 fluorescence staining, cell apoptosis was measured by Annexin-V-FITC/PI double staining, and expression of PKC? mRNA and protein was detected by RT-PCR and Western blotting, respectively. Results Cell apoptosis significantly increased in fluid A+powder B group, with typical morphology of apoptosis. After treatment for 24 h and 48 h, cell apoptosis rates in fluid A+powder B group were significantly higher than those at corresponding time points in blank control group, normal control group and fluid A+powder B+rottlerin group (P0.05). Conclusion Fluid A+powder B can significantly increase apoptosis of U937 cell line, the mechanism of which may be associated with the up-regulation of expression of PKC?. Compared with fluid A+powder B, fluid A+fluid B is superior in reducing apoptosis of peripheral blood monouclear cells.

Objective To explore the effect of platelet-rich plasma (PRP) on the generation of reactive oxygen species (ROS) and the phenotypes of photo-aging fibroblasts.Methods A photoaging cell model by repeating UVB irradiation was treated using appropriate concentration of PRP;Cell morphology and the rate of aging dying were observed under inverted microscope 24 hours later after establishment of the cell model;The expression of ROS between experimental and control group was detected using fluorescence microscope after single UVB irradiation.The relative intensity of fluorescence was analyzed using flow cytometry.Results PRP could ameliorate the large and sprawl appearance of photoaging fibroblasts obviously,reduce the generation of ROS as well as decrease the relative intensity of ROS.Conclusions PRP can decrease the level of intracellular oxidative stress caused by UVB irradiation,reduce the generation of ROS and ameliorate the senescence-like phenotypes of pho toaging fibroblasts.

Adenovirus vectors are one of the most promising gene transfer systems. They are of great value for gene therapy because these vectors achieve temporal high-level transgene expression and high gene transfer efficiency. To meet increasing needs of adenovirus vectors for gene therapy programs, parallel development of efficient, scalable and reproducible production processes is required. Perfusion cultivation of 293 cells is one of the most commonly used methods to produce adenovirus vectors and it is suitable for industrialized production specially. Experimental studies had been carried out to produce recombinant adenovirus containing the green fluorescent protein gene (Ad-GFP) by perfusion cultivation of HEK-293 N3S cells in a 5L stirring bioreactors. Perfusion rate was 1-2 volume/day. To infect the 293 N3S cells with Ad-GFP at the density of (2-4) x 10(6) cells/ ml. The time of collecting cells was 48 hours post infection. After three rounds of freeze/thaw and centrifugation, the crude viral lysates were stored at--80 degrees C until use. Then to get the Ad-GFP products by 2 x CsCl-gradient purification. The purity of the products was determined by the A260/A280 ratio and a high performance liquid chromatography (HPLC) assay. The infective titer was determined by a TCID50 assay. The culture term was 10-12 days. The infectious titer, the number of virus particle and the ratio of infectious titer to virus particle for the product were 1.0 x 10(11) IU/mL, 1.68 x 10(12) VP/mL and 6.0% IU/VP respectively. The A260/A280 ratio was 1.33, and the purity determined by HPLC was 99.2%. The cell specific productivity was around 1000 IU/cell. By perfusion cultivation of 293 N3S cells in a 5L stirring bioreactors, we established the production process for Ad-GFP, which paves a way to produce other recombinant adenovirus for gene therapy.
Adenoviridae ; genetics ; growth & development ; isolation & purification ; Bioreactors ; microbiology ; Cell Line ; Gene Transfer Techniques ; Genetic Vectors ; Green Fluorescent Proteins ; genetics ; Humans ; Kidney ; cytology ; virology ; Recombinant Proteins ; biosynthesis ; genetics ; Recombination, Genetic ; Virus Cultivation ; instrumentation ; methods

OBJECTIVETo investigate whether or not NADPH oxidase (NOX) participates in leptin-induced reactive oxygen species (ROS) production in hepatic stellate cells (HSC) and to explore the possible mechanism.

METHODSHSC-T6 cells (rat hepatic stellate cells line) were divided into nine groups: Group1: leptin (100 ng/ml) treated; Group2-6: leptin treated together with inhibitors that block different ROS-producing systems: diphenylene-iodonium (DPI) (20 micromol/L), Rotenone (20 micromol/L), Metyrapone (250 micromol/L), Allopurinol (100 micromol/L) and Indomethacin(100 micromol/L); Group7: leptin treated together with Janus kinase (JAK) inhibitor AG490 50 micromol/L; Group8: normal control group (treated DMEM with 0.1% DMSO); Group9: negative control group (untreated). Intracellular ROS levels were measured with dichlorodihydrofluorescein diacetate (DCFH-DA) dye assay by Fluorescence microscope and/or flow cytometry. NOX activity was analyzed by using spectrophotometer to calculate the absorbance of NADPH. The mRNA levels of Rac1 and p22Phox were evaluated by RT-PCR.

RESULTS(1) Leptin increased significantly the ROS production as compared to normal control group (92.91+/-4.19 vs.27.56+/-6.27, P<0.01) in HSC-T6 cells. Both the NADPH oxidase inhibitor DPI and AG490 (50 micromol/L) blocked the ROS production, inhibitors of other ROS producing systems had no significant effect on ROS production induced by lepin (P is more than 0.05). (2) Leptin treated HSC-T6 cells for 1 hour up-regulated the NOX activity significantly compared with that in normal control group [(1.90+/-0.22) pmol.min(-1).mg(-1) vs. (0.76+/-0.06) pmol.min(-1).mg(-1), P<0.05]. Furthermore, the NOX activity increased after being treated with leptin for 12 hours and 24 hours than being treated for 1 hour. Leptin-induced up-regulation of NOX activity was inhibited by pretreatment with DPI or AG490. (3) The RT-PCR results indicated that mRNA expressions of Rac1 and p22Phox in HSC-T6 cells with 12 hours of leptin stimulation increased significantly as compared with normal control group (0.41+/-0.13 vs 0.14+/-0.08, 0.45+/-0.12 vs 0.20+/-0.08, all P<0.05), while the DPI and AG490 had no effect on the mRNA expressions of Rac1 and p22Phox.

CONCLUSIONNOX is the main cellular source of the reactive oxygen species (ROS) generated by HSCs in response to leptin stimulation. The mechanism is probably that leptin can directly activate NOX through JAK signal transduction and hence induce the expression of NOX subunit to promote the activity of NOX which generates considerable ROS in HSC.

Animals ; Cells, Cultured ; Hepatic Stellate Cells ; drug effects ; metabolism ; Leptin ; pharmacology ; NADPH Oxidases ; genetics ; metabolism ; Rats ; Reactive Oxygen Species ; metabolism

OBJECTIVETo explore new methods to repair postburn contracture deformities in patients with extremely limited donor skin.

METHODSFive severely burned patients with extremely limited donor skin but severe deformities were enrolled in the study. The mature and the pliable scarred skin was utilized as the donor site for reconstruction of the postburn deformities. Split-thickness scarred skin was harvested for repair of postburn deformities after the scarred skin was expanded by expander, while thin razor-thin scarred skin with allo-acellular dermal matrix (ADM) was employed for the repair of postburn deformities when skin expansion was not feasible.

RESULTSAll the expanded scarred skin and composite skin grafts survived completely with good function and configuration. The long-term follow-up result was satisfactory, and the grafted skin was similar to that with split-thickness skin grafting.

CONCLUSIONIt is feasible to employ various split-thickness scarred skin for the reconstruction of postburn deformities. This technique is a new effective procedure for the reconstruction of postburn deformities, especially for those with extreme scarcity of donor site.

Adult ; Burns ; pathology ; surgery ; Cicatrix ; pathology ; Contracture ; surgery ; Female ; Humans ; Male ; Middle Aged ; Skin Transplantation ; methods