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OBJECTIVE@#To establish a micellar electrokinetic capillary chromatography-based method for identification and quantitative detection of interleukin-12 (IL-12) and analysis of its unfolding process.@*METHODS@#An uncoated fused-silica capillary (inner diameter 50 μm) with a total length of 48.5 cm (40 cm to the detector) was used for the experiment. The factors influencing the separation efficiency of IL-12 were analyzed, and a standard curve of IL-12 concentration was established. The mixture of IL-12 and anti-IL-12 antibody was incubated in a water bath at 38 ℃ for 40 min, and capillary electrophoresis was then performed under the same conditions. The results were compared with those of IL-12 and anti-IL-12 antibody to identify IL-12. IL-12 and dithiothreitol (DTT) were incubated at 60 ℃ in water bath for different lengths of times, and the unfolding process of IL-12 was analyzed based on electrophoresis results of IL-12 in different states.@*RESULTS@#A micellar capillary electrophoresis on-line sweep method was established with 80 mmol/L borate (pH=9.3) containing 30 mmol/L sodium dodecyl sulfate (SDS) as the buffer solution. This system showed a good linear relationship between the peak area and the mass concentration of IL-12 with a linear correlation coefficient of 0.9991 within the linear range of 2 to 120 ng/L. As the incubation time of IL-12 and DTT prolonged, the disulfide bond of IL-12 gradually opened and resulted in distinct changes in the protein peak.@*CONCLUSIONS@#This capillary electrophoresis-based method is simple and sensitive for IL-2 analysis and allows rapid detection of changes in IL-12 content in the setting of tumors and analysis of the possible causes.
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Objective To compare the EEG complexity between rats under awaking and differ-ent depth of anesthesia via analyzing sample entropy and fractal dimension.Methods Sixteen SD rats were intraperitoneally injected with urethane twice,first with 500 mg/kg and second with 800 mg/kg one hour later.The scalp EEG was collected in stage of awaking (W),light anesthesia (LA)and heavy anesthesia (HA).The sample entropy (SampEn)and fractal dimension (FD)were computed by MATLAB.The characteristic values were denoised by linear dynamic system method during the whole process.Results The value of SampEn and FD gradually dropped from awaking to heavy anes-thesia.The SampEn and FD in W was significantly higher than the value in LA or in HA (P <0.05). The SampEn and FD in HA was significantly lower than that in LA (P < 0.05 ).Conclusion The SampEn and FD of EEG could be used to monitor the depth of anesthesia.
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Objective To investigate the mechanism of platelet inhibitor from Agkistrodon halys venom (AHV-PI) on platelet aggregation. Methods Protein kinase Akt phosphorylation levels in platelet were measured by Western blot. XS-1000I blood cell counter was used for platelet count. The plasma content of 5'-NT and platelet membrane GPIb were determined by Enzyme-Linked Immunosorbnent Assay (ELISA). The effect of AHV-PI on binding rate between the fluorescence labeled monoclonal antibody CD61 (FITC-CD61) and platelet membrae glycoprotein Ⅱb/Ⅲa (GPⅡb/Ⅲa) was observed by flow cytometry (FCM). Results AHV-PI can reduce the level of Akt-phosphorylation level and the number of platelet. AHV-PI can increase the content of 5'-NT in plasma, reduce the expression of platelet GPIb. Flow cytometry displayed AHV-PI can not affect the rate of combination between platelet GPⅡb/Ⅲa and FITC-CD61. Conclusion The mechanism of inhibition of platelet aggregation may be inhibit protein kinase Akt phosphorylation to block the signal transduction pathway of Akt. Limit cell grouth and reduce platelet number, also it may be related to its 5'-NT activity, it can degradate ADP to prevent the formation of platelet thrombus.
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AIM: To investigate the effects of protein C activator (PCA) from Agkistrondon acutus venom ( AAV) on the tension of thoracic aorta rings isolated from the rats with sepsis.METHODS:The model of sepsis was es-tablished by intraperitoneal injection of lipopolysaccharide ( LPS) .SD rats were randomly divided to 6 groups ( n=6 ):sham group, LPS group, PCA intervention group (LPS+PCA, PCA at doses of 0.1 mg/kg, 0.3 mg/kg and 0.6 mg/kg) and LPS+polymyxin B (at dose of 0.2 mg/kg) group.Using perfusion experiment in vitro, the tension of the aortic rings was measured by biological signal analytical system.RESULTS:The values of MABP, HR, LVDP and ±dp/dtmax were significantly lower in LPS group than those in sham group and LPS+PCA groups.Compared with sham group, the relaxa-tion response to acetylcholine ( ACh) and the contractile response of aorta rings induced by phenylephrine ( Phe) were sig-nificantly decreased in LPS group, which were increased significantly in PCA intervention group ( especially at dose of 0.6 mg/kg) compared with LPS group.The dose-response curve of aorta contraction with denuded endothelium induced by Phe shifted down significantly in LPS group compared with sham group, and no significant difference between LPS group and PCA intervention group was observed.Also no statistical difference was found in non-endothelium dependent relaxation of aortic rings induced by sodium nitroprusside among the groups.Pretreatment of N-nitro-L-arginine methl ester and methyl-ene blue increased the contraction amplitude of aortic rings induced by Phe.CONCLUSION:PCA from AAV effectively reverses the hypoergia of the vessels in rats with sepsis through protecting vascular endothelium, the mechanism of which may be mediated by inhibiting NO-GC-cGMP signal transduction pathway.
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Cultivating the talents with scientific research and innovation has been the emphasis of medical education in 21st century.We make a deep exploration and practice on how to cultivate the innovative ability of undergraduates in functional experimental teaching.The article points out that it is an effective way to convert educational sense,to update educational mode,to strengthen the scientific research practice,and to enhance the innovative experiment.
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AIM To evaluate the changes of serum and brain tissue endoxin in model of bilateral cerebral hemisphere ischemic reperfusion injury, and effect of anti digoxin antiserum (an antagonist of endoxin). METHODS The bilateral cerebral hemisphere ischemic model was prepared by ligating three vascular by Kameyama's manner. SD rats were randomly divided into 7 groups and each group had 8 rats. Sham group, ischemic reperfusion group, negative control group, nimodipine group, low concentration anti digoxin antiserum group, middle concentration anti digoxin antiserum group, high concentration anti digoxin antiserum group. The blood was collected at the end of reperfusion, meanwhile rats were killed, and the bilateral cerebral hemisphere were took out and used to prepare encephlon homogenate and made into samples of light microscope. RESULTS Compared with sham group, the serum CK content increased; Brain tissue SOD activity reduced and MDA content increased importantly in ischemia reperfusion group; The levels of serum and brain tissue endoxin in ischemia reperfusion group were significantly higher, while ATPase activity in brain tissue decreased; Mitochondrial Ca 2+ content in brain tissue increased significantly and Mg 2+ content decreased significantly. In brain tissue,there was some inflammatory change and local necrosis;The rank order and structure of cell wasn't clear;The morphology of pyramidal cell was abnormal. Compared with ischemic reperfusion group, Anti digoxin antiserum reduced serum CK content; It antagonized lowering of SOD activity and increase of MDA content in brain tissue; It remarkably reduced the level of brain tissue endoxin; It reduced abnormal ion content of brain tissue mitochondrion induced by cerebral ischemic reperfusion injury; The high and middle concentration anti digoxin antiserum had a significant effect on raising brain tissue ATPase activity. It reduced neuron denaturation. CONCLUSION Cerebral ischemic reperfusion can increase the level of brain tissue and serum endoxin and higher endoxin can promote brain injury. Endoxin is a major factor involved in cerebral ischemic reperfusion injury. Anti digoxin antiserum can reduce brain tissue injury and had a protective and treatment effect on cerebral ischemic reperfusion injury by antagonizing the effect of endoxin.
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AIM: To investigate the protective effects and its mechanism of Ginkoglide A on stress ulceration in fats. METHODS: Wistar rats were randomly assigned to control, and Ginkgolides pretreatment group. The stress ulceration was produced by water immersion restraint, and gastric myoelectric activity during during was recorded by implanting electrodes in the stomach antrum smooth muscle. The malondialdehyde (MDA) concentration and superox- ide dismutase (SOD) activity in plasma, and gastric mucusal lesions were examined after the rat was killed. RE- SULT: Compared with the control group, previous intraperitoneal administratin of Ginkgolide A (5 - 20 mg/kg) not only had a highly significant decrease on stress - induced myoelectric activity disorder (P
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AIM:To explore the effects of the new drug of sulfonylurea(1-{4-[2-(3-ethyl-4-methyl-2-oxo-3-pyrroline-1-carboxamido)ethyl]-phenylsulfonyl}-3-(1,4-tetramethylene)-urea,BGW) on the glucose uptake and the activation of Akt/PKB in SMMC7721 cells.METHODS:Cultured SMMC7721 cells were divided into control group,glibenclamide group,insulin group,BGW group and BGW+insulin group.Scintillation was used to detect the glucose uptake in SMM7721 cells.The activation of Akt/PKB was tested by Western blotting.RESULTS:Compared to control cells,gibenclamide,insulin,BGW and BGW+insulin significantly increased the glucose uptake(P