1.The Effects of Parvoviral Nonstructural Proteins Gene Transfection on the Expression of Cytokines in Gastric Cancer Cell
Xizhong SHEN ; Guanhua YU ; Shaoji JIANG
Chinese Journal of Cancer Biotherapy 1995;0(03):-
Human gastric cancer cells (MKN-45) were transfected with pULB3238 (a plasmid carrying MVMp NS gene). After the NS gene expressed, some of the NS-transfectants were died. The other cells were survived. In comparison with those of control MKN-45, the survived NS- transfected MKN-45 cells had the following characterizations: (a). nucleus/cytoplasm ratio reduced; (b). generation time prolonged;(c). adhesion ability between cells increased;(d). cloning efficiency decreased;(e). formation of tumors in nude mice suppressed. To investigate the possible mechanisms of above mentioned change of NS-transfectants, the effect of NS on the expression of some cytokine genes were explored. RNA dot blot indicated that the NS might enhance the expression of IL-l?(2.7 times), IL-1/?(4.6 times) IL- 6 nuclear factor (2.5 times) and had no effect on the expression of IL-6. The results suggested that the antineoplastic activity of parvoviruses may be medicated by NS interfere with the expression of cytokines in the host cells.
2.Research progress of animal models of systemic lupus erythematosus
Yu YAN ; Lianhua FANG ; Guanhua DU
Acta Laboratorium Animalis Scientia Sinica 2015;(4):428-433
Systemic lupus erythematosus ( SLE) is a chronic multisystem relapsing-remitting autoimmune disease, which affects human health seriously.There are numerous animal models that have long been employed in an effort to un-derstand the mechanism and treatment of SLE.Animal models of SLE were reviewed and compared in this paper, to provide references for the researchers to choose appropriate models for studying specific pathogenic mechanism and diagnostic crite-ria, searching for targeted treatment interventions and developing potential therapeutic drugs.
3.High-throughput screening of human soluble epoxide hydrolase inhibitors.
Shoubao WANG ; Jing GUO ; Xiaoming YU ; Guanhua DU
Acta Pharmaceutica Sinica 2010;45(11):1367-72
To screen potential human soluble epoxide hydrolase (hsEH) inhibitors, a high-throughput screening model in 384-well microplate with total volume of 50 microL was established. Recombinant hsEH was cloned and expressed in E. coli. and its specific substrate PHOME was synthesized. The HTS model was based on fluorescence analysis with enhanced sensitivity and specificity (Z' = 0.65). A total of 47 360 samples (including 25 040 compounds and 22 320 natural products) were screened, of which 950 samples with inhibition greater than 80% were selected for further rescreening. Finally, two compounds with high inhibitory activity were identified, whose IC50 value were 8.56 and 4.31 micromol x L(-1), separately. The results indicated that the method was stable, sensitive, reproducible and also suitable for high-throughput screening.
4.Establishment of a novel high throughput screening assay for identifying small molecular antagonists of human interleukin-6 receptor
Yu YAN ; Yangyang HE ; Chang ZHANG ; Xiaobin PANG ; Peng DU ; Zhiwei SUN ; Shuang WANG ; Guanhua DU
Military Medical Sciences 2014;(12):921-926
Objective To establish a high throughput screening assay for identifying human small molecular antagonists targeted IL-6R.Methods The full length gene of the human IL-6R extracellular region was amplified by PCR and cloned into a eukaryotic expression vector to construct recombination expression plasmid pABHis -IL6R that was then transfected transiently into HEK293T cells to prepare recombination protein IL-6R.Western blotting assay and receptor-ligand binding experiment were used to analyze the bioactivity of IL-6R.A new screening method based on ELISA was established using the function of IL-6R binding to its ligand and the characteristics of Fc fragment binding to IgG-HRP.Then Z′-factor was calculated and a known antagonist ab 47215 was used to assess the stability and reliability of the new assay .Results Recombination plasmid pABHis-IL6R was constructed and soluble IL-6R was prepared.IL-6R reported herein could be recognized by an anti-IL-6R antibody and specifically bind to its ligand in a dose response manner .A Z′-factor of 0.53 was obtained that could serve high throughput screening assay .Ab47215 , as a known specific antagonist , was able to block rhIL-6 from binding to the receptor in a dose-dependent manner in the new screening assay , the IC50 of which was (0.55 ± 0.11)μg/ml.Conclusion An innovative and easy screening assay for identifying human IL-6R antagonists is established , which might help discover potent and specific antagonists .
5.Vasorelaxant effect of Rho kinase inhibitor DL0805-0 on isolated rat aortic rings and its underlying mechanisms
Yu YAN ; Subo WANG ; Tianyi YUAN ; Xiaozhen JIAO ; Ping XIE ; Lianhua FANG ; Guanhua DU
Chinese Pharmacological Bulletin 2014;(4):473-477
Aim To investigate the in vitro vasorelax-ant effect of DL0805-0, a Rho kinase inhibitor, on iso-lated rat thoracic aorta and explore its underlying mechanism. Methods Tension was measured to eval-uate the vasorelaxant effect of DL0805-0 on rat endo-thelium-intact and endothelium-denuded thoracic aorta rings. Rho kinase inhibitor fasudil, nitric oxide syn-thase inhibitor Nω-nitro-L-arginine methyl ester ( L-NAME), guanylate cyclase inhibitor methylene blue, cyclooxygenase inhibitor indomethacin, calcium-activa-ted potassium channel blocker tetraethyl ammonium ( TEA ) , ATP-sensitive potassium channel blocker glibenclamide and voltage-dependent potassium chan-nel blocker 4-aminopyridine ( 4-AP ) were used to il-lustrate the mechanisms of vasorelaxant effect of DL0805-0 . Results DL0805-0 exerted vasorelaxation in a dose-dependent manner in KCl (60 mmol·L-1 ) or NE ( 0. 1 μmol · L-1 ) -induced contraction. DL0805-0-induced vasorelaxation was significantly re-duced by L-NAME. However, methylene blue and in-domethacin did not significantly affect vasorelaxation of DL0805-0. In endothelium-denuded rings, TEA re-markably attenuated the vasorelaxant effect of DL0805-0 , while glibenclamide and 4-AP did not affect vasore laxation of DL0805-0 significantly. DL0805-0 also re-duced NE-induced transient contraction and inhibited contraction induced by increasing extracellular calci-um. Conclusion These results suggest that DL0805-0 induces vasorelaxation through an endothelium-depend-ent pathway. The opening of calcium-activated K+channels and blocking of Ca2+ channels in vascular smooth muscle cells may be one of the mechanisms of DL0805-0-induced vasorelaxation.
6.Absorption dynamic characteristics of clopidogrel bisulfate polymorphs in rat.
Xiaoyan YU ; Qianxi CHEN ; Xiaoyu BAI ; Shuo TIAN ; Jialin SUN ; Yang Lü ; Guanhua DU
Acta Pharmaceutica Sinica 2011;46(10):1268-72
Four crystalline forms of clopidogrel bisulfate were characterized by analytical techniques. Aiming to research the absorption characteristics of clopidogrel bisulfate polymorphs after taken orally by rat, and to estimate the influence of crystal form to pharmacodynamic action, four crystalline forms of clopidogrel bisulfate were administered intragastrically to rats, and high performance liquid chromatography (HPLC) was used to measure the contents of clopidogrel bisulfate and its metabolite in rat plasma. The metabolite of clopidogrel bisulfate was detected in rat plasma. There were significant deviations among four crystalline forms in the areas under curve of the metabolite of clopidogrel bisulfate. We concluded that the different crystal forms of clopidogrel bisulfate showed different pharmacokinetic characteristics, which might affect pharmacodynamic action.
7.Effects of paclitaxel loaded-drug micelles on cell proliferation and apoptosis of human lung cancer A549 cells.
Lin WANG ; Ruishuang YU ; Wenliang YANG ; Shujuan LUAN ; Benkai QIN ; Xiaobin PANG ; Guanhua DU
Acta Pharmaceutica Sinica 2015;50(10):1240-5
This study was conducted to investigate the paclitaxel loaded by hydrazone bonds in poly(ethylene glycol)-poly(caprolactone) micelles (mPEG-PCL-PTX) on proliferation and apoptosis of human lung cancer A549 cells and its possible mechanisms of anti-tumor activity. The cell proliferation was measured with MTT assay. Flow cytometry were used to analyze the cell cycle. The cell apoptosis was analyzed using Hoechst/P staining. The expression levels of apoptotic genes expression in the mitochondrial apoptosis pathway were detected by RT-PCR and Western blotting, respectively. The mPEG-PCL-PTX could inhibit the proliferation of A549 cells and promote the apoptosis. The Bax, caspase-3 protein expression were increased while Bcl-2 protein expression was decreased in A549 cells. Results showed that the polymer containing hydrazone bond is non-toxic in vitro, the mPEG-PCL-PTX micelles can inhibit the proliferation and induce the apoptosis of A549 cells. Key words: paclitaxel; micelle; A549 cell; proliferation; cell cycle; apoptosis
8.System and intranasal immunization of group B streptococcal C5a peptide
Guanhua XUE ; Shentao LI ; Yonghong YANG ; Lihua YU ; Aihua WANG ; Guan WANG ; Liqin YUE ; Guorong YIN ; Xuzhuang SHEN
Chinese Journal of Microbiology and Immunology 2008;28(12):1137-1142
Objective To study the system of streptococcal C5a peptidase (ScpB) and the specif-ic antibody levels in serum, lung, vagina and recta after subcutaneous and intranasal immunization with dif-ferent doses of C5a peptide. Methods Recombinant protein C5a peptide was expressed in E. coli strain BL21 and purified by affinity chromatography. The expressed product was identified by SDS-PAGE and pep-tide mass fingerprinting (PMF). BALB/c mice were subcutaneously and intranasally injected with different doses of ScpB. Antibody titer was tested by ELISA. Opsonophagocytosis assay was used to test the function of antibody. Results ScpB protein was successfully expressed and purified. The probability based mouse score of ScpB was 175 by PMF analysis. ELISA data showed that both subcutaneous and intranasal immtmi-zation could elicit significantly higher levels of IgG in immunized mice serum than that of control group (P <0.01), 30 μg group waa better than 5 μg and 10 μg group. Intranasal immunization could elicit higher lev-els of IgA in lung, vagina and rectum (P <0.001) while system immunization could not. Opsenophagocyto-sis tests indicated that anti-serum of ScpB had opsenophagocytic activity than that of control (P < 0. 05).Conclusion The results demonstrated that intranasal immunization with ScpB could induce significantly higher levels of lgG and IgA, and its anti-serum had better opsenic activity.
9.Blood compatibility of two novel polyurethane coating materials.
Guanhua YU ; Jian JI ; Dongan WANG ; Linxian FENG ; Jiacong SHEN
Journal of Biomedical Engineering 2004;21(2):184-187
Amphiphilic coupling-polymer of stearyl poly (ethylene oxide)-co-4, 4'-methylendiphenyl diisocyanate-co-stearyl poly(ethylene oxide), MSPEO, was specially designed as surface-modifying additives. The blends of MSPEO in both polyether urethane (PEU) and chitosan(Chi), as the coating materials for intravascular device were investigated. Two kinds of static clotting time tests, plasma recalcification time (PRT) and prothrombin time(PT), as well as the static platelet adhesion experiment were carried out. And the dynamic anti-coagulation experiment was performed with a closed-loop tubular system under a blood shear rate of 1,500 s-1. The results demonstrate that both blend coatings can improve the anti-coagulation of polyurethane greatly and will not lead to hemolysis, and that more platelets adhere to the surface modified by Chi-MSPEO blend coating as compared with those adhere to the surface modified by PEU-MSPEO blend coating. The surface modified by Chi-MSPEO has longer PRT, whereas the surface modified by PEU-MSPEO has longer PT.
Blood Coagulation Tests
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Chitin
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analogs & derivatives
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chemistry
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Chitosan
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Coated Materials, Biocompatible
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chemistry
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Ethylene Oxide
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chemistry
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Humans
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Materials Testing
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Platelet Adhesiveness
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Polymers
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chemistry
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Polyurethanes
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chemistry
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Prothrombin Time
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Surface Properties
10.Effects of sevoflurane pretreatment on myocardial injury in patients undergoing cardiac valve replacement with cardiopulmonary bypass
Chengjie GAO ; Jingui YU ; Huixia WANG ; Bo LI ; Xiaoming Lü ; Lufeng XU ; Guimao CAO ; Jishun NING ; Guanhua JIANG ; Aijun NIU ; Hailong DONG
Chinese Journal of Anesthesiology 2012;32(3):278-281
Objective To investigate the effects of sevoflurane pretreatment on the myocardial injury in patients undergoing cardiac valve replacement with cardiopulmonary bypass (CPB).Methods Twenty NYHA class Ⅱ or Ⅲ patients,aged < 60 yr,undergoing cardiac valve replacement with CPB,were randomly divided into 2 groups (n =10 each):sevoflurane group (group S) and control group (group C).The patients were premeditated with intramuscular morphine and scopolamine.Anesthesia was induced with iv injection of midazolam 0.05-0.08 mg/kg,fentanyl 10-15 μg/kg and pipecuronium 0.08-0.10 mg/kg.Anesthesia was maintained with intermittent iv boluses of midazolam,fentanyl and pipecuronium and in addition sevoflurane was inhaled before aortic clamping and the end-tidal concentration was rapidly increased to 1.0% and maintained at the level for 5 min in group S.Blood samples were taken from the central vein before skin incision (T1),immediately after aortic clamping (T2 ),at 0 and 30 min after aortic unclamping (T3-4),and at 2,6,12 and 24 h after operation (T5-8) for determination of the concentration of serum cardiac troponin Ⅰ (cTnI) and activities of creatine kinase (CK) and creatine kinase isoenzyme-MB (CK-MB).Myocardial specimens were taken from right auricle before aortic clamping and at 10 min after aortic unclamping for electron microscopic examination.Results The concentration of serum cTnI and activities of CK and CK-MB were significantly increased at T4-8 in both groups ( P < 0.05).The serum cTnI concentration at T4-8 and the activities of CK and CK-MB at T8 were significantly lower in group S than in group C ( P <0.05).Different degrees of mitochondrial swelling were observed after aortic unclamping in both groups,but the changes were milder in group S than in group C.Conclusion Sevoflurane pretreatment can attenuate the myocardial injury in patients undergoing cardiac valve replacement with CPB.