3.Preparation of cell penetrating peptide TAT and cleavable PEGco-modified liposomes loaded with paclitaxel and its in vitro apoptosis assay.
Han FU ; Guan-Lian HU ; Qin HE
Acta Pharmaceutica Sinica 2014;49(7):1054-1061
The preparation method, serum stability, efficiency of cellular uptake and apoptosis induction of the cell penetrating peptide TAT and cleavable PEG co-modified liposomes loaded with paclitaxel (C-TAT-Lipo) were investigated. The best preparation procedure was performed by orthogonal test based on single factor screening method. First, the paclitaxel (PTX)-loaded liposomes were prepared by filming-rehydration method, evaluated with entrapment efficiency and polydispersity index. The morphology of C-TAT-Lipo was characterized by transmission electron microscopy. Turbidity variations were monitored in the presence of fetal bovine serum (FBS) to evaluate the serum stability of the liposomes developed here. Next, the efficiency of cellular uptake of different Rho-PE-labeled liposomes on B16F1 cells in vitro was evaluated by confocal laser scanning microscopy (CLSM) and flow cytometry. The quantitative analysis of apoptosis induced by different PTX-loaded liposomes was performed by Annexin V-FITC/PI double staining. The optimal formulation was as follows: Chol : lipid: 1 : 8 (molar ratio); drug : lipid: 1 : 40 (mass ratio); lipid concentration: 3 mmol x L(-1); temperature of hydration: 25 degrees C. The mean size and polydispersity index of C-TAT-Lipo were about (97.97 +/- 3.68) nm and 0.196 +/- 0.037, the zeta potential was (-0.89 +/- 0.45) mV, the entrapment efficiency of paclitaxel was (90.16 +/- 1.53)%. The particle sizes did not exhibit significant variations in 50% FBS over 24 h at 37 degrees C. The efficiency of cellular uptake of the C-TAT-Lipo increased 1.40 fold following the cleavage of PEG. Apoptosis analysis showed 59.3% increase of the apoptosis and necrosis profile of C-TAT-Lipo after the detachment of PEG shells, which was markedly higher than that of N-TAT-LP with or without glutathione and SL, respectively. The results indicate that the C-TAT-Lipo is successfully prepared by filming-rehydration method and shows significant antitumor activities.
Animals
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Annexin A5
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Apoptosis
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Cell Line, Tumor
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Cell-Penetrating Peptides
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pharmacology
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Fluorescein-5-isothiocyanate
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analogs & derivatives
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Liposomes
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chemistry
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Melanoma, Experimental
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Mice
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Microscopy, Confocal
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Paclitaxel
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pharmacology
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Particle Size
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Polyethylene Glycols
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chemistry
4.Effect of ferulic acid on cholesterol efflux in macrophage foam cell formation and potential mechanism.
China Journal of Chinese Materia Medica 2015;40(3):533-537
The formation of macrophage-derived foam cells is a typical feature of atherosclerosis (AS). Reverse cholesterol efflux (RCT) is one of important factors for the formation of macrophage foam cells. In this study, macrophage form cells were induced by oxidized low density lipoprotein (ox-LDL) and then treated with different concentrations of ferulic acid, so as to observe the effect of ferulic acid on the intracellular lipid metabolism in the ox-LDL-induced macrophage foam cell formation, the cholesterol efflux and the mRNA expression and protein levels of ATP binding cassette transporter A1 (ABCA1) and ATP binding cassette transporter G1 (ABCG1) that mediate cholesterol efflux, and discuss the potential mechanism of ferulic acid in resisting AS. According to the findings, compared with the control group, the ox-LDL-treated group showed significant increase in intracellular lipid content, especially for the cholesterol content; whereas the intracellular lipid accumulation markedly decreased, after the treatment with ferulic acid. The data also demonstrated that the mRNA and protein expressions of ABCA1 and ABCG1 significantly increased after macrophage foam cells were treated with different concentrations of ferulic acid. In summary, ferulic acid may show the anti-atherosclerosis effect by increasing the surface ABCA1 and ABCG1 expressions of macrophage form cells and promoting cholesterol efflux.
ATP Binding Cassette Transporter 1
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analysis
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genetics
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ATP Binding Cassette Transporter, Sub-Family G, Member 1
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ATP-Binding Cassette Transporters
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analysis
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genetics
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Animals
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Cells, Cultured
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Cholesterol
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metabolism
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Coumaric Acids
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pharmacology
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Foam Cells
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drug effects
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metabolism
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Lipoproteins
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analysis
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genetics
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Mice
5.Awareness rate of child health management in parents of children aged 0 - 36 months
Xiaobing FU ; Jie LIAN ; Ping SHI
Chinese Journal of General Practitioners 2012;(7):519-521
A questionnaire survey on awareness of child health management was conducted in 238 family members ( 236 were parents) of children aged 0 - 36 months; among whom 92 of children aged 0 to 6 months,85 of children aged 7 to 12 months,33 of children aged 13 to 18 months,16 of children aged 19 to 24 months and 12 of children aged 24 to 36 months.52.1% of the surveyees had college education level.The awareness about medical examination in last half year in female subjects was higher than males ( P < 0.05).There were no differences in other health management knowledge.The results showed that the average times of medical examination was 2.8 ± 1.9 in last half year; the age for supplement of meat in food was 7.4 ± 2.4 mouths; supplement of meat,animal liver and eggs was more than 7 time a week.The subjects with college education or above paid more attention to the children's growth,risk for rickets and anemia than other peoples.Pure breast feeding accounted for only 37.0% (88/238).The study indicates that strengthening the knowledge and awareness is necessary to improve the children health management.
6.Experimental study on electroencephalographic characteristic in kainic acid induced temporal lobe epilepsy model established with stereotactic technique in rats
Ruobing QIAN ; Xianming FU ; Lian YU
Journal of Clinical Neurology 1992;0(01):-
Objective To explore the method for establish the temporal lobe epilepsy model in SD rats with stereotactic technique and to study the characteristic of electroencephalography (EEG) after epileptic seizure. Methods Kainic acid was injected into right hippocampus via stereotactic technique in 20 rats, and the behavior of rat was recorded by a video camera. The electrical activity changes of brain were recorded by EEG in bilateral hippocampus, amygdala and frontal lobe. Results According to Racine's scale, 6 rats were typeⅤ,10 rats were type Ⅳ,3 rats were type Ⅲ,1 rat was type Ⅱ. The spike wave was originated from hippocampus and spread to amygdala and frontal lobe. Pathologic changes showed that prolonged seizures induced pyramidal cell loss and death in hippocampus. Conclusions The method to establish kainic acid induced temporal lobe epilepsy model with stereotactic technique was credible, economical and useful. The spike wave originated from hippocampus was the basic feature of EEG of temporal lobe epilepsy model in rats.
7.Expression of Hypoxia Inducible Factor-1? in Vertebra Development of Mouse
Journal of Shanghai Jiaotong University(Medical Science) 2006;0(05):-
Objective To investigate the expression pattern of hypoxia inducible factor-1?(HIF-1?) in fetal vertebra development of the mouse. Methods The development of mouse fetal vertebra was observed dynamically,and the expression of HIF-1? mRNA at various stages was also detected by reverse transcription-polymerase chain reaction. Results The cartilaginous spine column began to form at E13.5.The primary ossification center was observed at E15.5,then the osteogenesis expanded and extended to both sides.HIF-1? mRNA began to express at E13.5,and more significantly at E14.5(P
8.The effect of different shoulder width on the compressive strength of full-contour zirconia crowns
Yan HAO ; Yunmin LIAN ; Qiang FU ; Shaoran DONG
Journal of Practical Stomatology 2017;33(4):451-454
Objective:To evaluate the effect of different shoulder width on the compressive strength of full-contour zirconia crowns.Methods:A computer aided design and computer aided manufacturing(CAD/CAM) system was used to prepare 24 full-contour zirconia crowns of the mandible first molar.1 mm occlusal thickness crowns with 4 shoulder width of knife-edge(group A),0.3 mm(group B),0.5 mm(group C) and 1.0 mm(group D) (n =6).The Co-Cr alloy PFM crowns made by the die of group C were used as the controls (n =6).All the full-contour zirconia crowns were bonded to the corresponding die using 3 M glass ionomer cement (GIC) and the compressive strength were measured by an universal testing machine.Based on the statistical analysis,crowns with the shoulder width of 0.5 mm were treated by the thermocycling test under cycling of 10000 times(group C1),20 000 times(group C2) and 30000 times (group C3) respectively(n =6).One-way ANOVA method was used for the statistical analysis.Results:The mean compressive strength(N) of group A,B,C,D and E was 3855.00 ± 305.47,2731.70 ± 261.80,3698.30 ± 276.87,3841.70 ± 544.88 and 2992.17 ± 168.41 respectively.The strength of group B and E were lower than that of group A,C and D(P < 0.05),B vs E P >0.05.The compressive strength(N) of group C1,C2 and C3 was 3220.00 ± 504.38 (group C1),3148.33 ± 425.60(C2) and 2992.17 ±168.41 (C3) respectively (P>0.05).Conclusion:For the full-contour zirconia crowns,both the knife-edge shoulder and the shoulder wider than 0.3 mm can meet the thermoclinical needs.The cycling times used in this study have no influences on the compressive strength.
9.The changes of pulmonary surfactant associated protein of sputum and oxygenation index in dysfunctional ventilatory weaning response patients
Yangjia DENG ; Qian WU ; Yuhong WEI ; Lei DU ; Lian FU
Chongqing Medicine 2014;(20):2572-2573,2576
Objective To explore the expression of SP-A in surface active substance of sputum pulmonary of dysfunctional venti-latory weaning response patients and to study changes of expression of SP-A and oxygenation index ,arterial partial pressure of oxy-gen .Methods The mechanical ventilatory patients were divide into dysfunctional ventilatory weaning response group (group A) and easily weaning group(group B) ,then collected sputum from ventilators at different times :on the first day ,two and seven days after using ventilators .Used ELISA to test level of SP-A and oxygenation index ,arterial partial pressure of oxygen in sputum of group A and group B .Results For group A ,SP-A content oxygenation index and arterial partial pressure of oxygen were significantly less than those for group B(P<0 .05);after two days ,for group A ,SP-A content in sputum was significantly less than that on the first day ,and SP-A content in sputum was significantly less than that on the second day (P<0 .05);two days later ,oxygenation index and arterial partial pressure of oxygen were significantly lmore than those on the first day (P<0 .05) and there was no statistical difference in oxygenation index and arterial partial pressure of oxygen between the seventh day and the second day (P>0 .05) .Con-clusion Dysfunctional ventilatory weaning response and lung injury related to ventilator can be diagnosed early through the test of SP-A content in sputum ,which provides a brand-new way and method of treatment in dysfunctional ventilatory weaning response and lung injury related to ventilator .
10.Expression of lentivirus-mediated neurotrophin-3 gene in Schwann cells
Jiandong YUAN ; Qiang FU ; Xiaofeng LIAN ; Tiesheng HOU ; Jie ZHAO
Chinese Journal of Tissue Engineering Research 2007;0(32):-
BACKGROUND:Neurotrophin-3 is found in the repair of spinal cord injury in the role of the strongest neurotrophic factor.It can effectively promote axonal regeneration through the glial scar tissue in repairing spinal cord injury.OBJECTIVE:To construct recombinant lentiviral vectors for gene delivery of homo sapiens neurotrophin-3(hNT3),and to investigate the expression of hNT3 gene in Schwann cells after transfection.DESIGN,TIME AND SETTING:Observational experiment was performed from June 2007 to March 2008 at the Central Laboratory of Changhai Hospital.MATERIALS:Bilateral sciatic nerves were harvested from 3-day-old Sprague Dawley rats for culture and identification of Schwann cells.Three-plasmid lentivirus systems:pGC-E1-EGFP,pHelper 1.0 and pHelper 2.0 were gained from Shanghai Chemical Technology Co.,Ltd.METHODS:pGC-E1-hNT3-EGFP plasmid was constructed by double restriction enzyme digestion and ligation,and then the plasmid was transformed into E.coli DH5?.Purified pGC-E1-hNT3-EGFP plasmids from the positive clones was confirmed by PCR and sequencing.293T cells were cotransfected with lentiviral vector pGC-E1-hNT3-EGFP,pHelper 1.0 and pHelper 2.0 by Lipofectamine 2000 to produce lentivirus.2 mL recombinant virus complete culture solution was added according to multiplicity of infection=1,4,8,10,12.The titer of virus was tested according to the expression level of enhanced green fluorescent protein.The control groups were Schwann cells and Schwann cells transfected by no-loaded lentivirus.MAIN OUTCOME MEASURES:The lentiviruses were transduced to Schwann cells,and the transfection efficiency was examined by flow cytometry,the overexpression of hNT3 was determined by Real-time PCR and Western blotting.RESULTS:The exogenous gene sequence of the recombinant hNT3 was completely in accordance with that of its open reading frame in GeneBank.The titer of concentrated virus was 5?107 TU/L.After recombinant LV-hNT3 infection,Schwann cells gave off strikingly bright green fluorescence,and the transfection efficiency amounted to 85%(multiplicity of infection=10).Real-time RCR test showed that the hNT3 mRNA were highly expressed in hNT3-Schwann cells,while did not express in the control group.Western blotting showed the hNT3 expression in Schwann cells.CONCLUSION:Construction of hNT3 gene lentiviral vector can transfect Schwann cells leading to an efficient overexpression of hNT3.