AIM: To investigate the effect of hyperbaric oxygen (HBO) on gelatinase,nitric oxide synthase and the permeability of brain blood barrier(BBB) in ischemia-reperfusion(I/R) mice. METHODS: Using cerebral I/R models, during the reperfusion period, 0.25 MPa (ATA) HBO were applied 5 times. matrix metalloproteinase(MMP)-2,9, nitric oxide synthase(NOS) and evans blue (EB) in brain were measured. RESULTS: ①HBO had significanty effect on MMP-9, but had little effect on MMP-2. ② HBO decreased the activity of NOS.③ The maxium amount of EB in IR group was at 4 hours after reperfusion and gradually decreased at 11 h, 23 h,48 h, 72 h. CONCLUSION: HBO may decrease the activities of MMP-9,NOS and the permeability of BBB in cerebral ischemia-reperfusion mice.

Objective:To obtain the high-efficiency expression of the biological active rabies virus nucleoprotein in the prokaryotic expression system.Methods:This experiment uses codon optimization technology to re-encode the nucleoprotein gene of rabies virus CTN-1 strain, artificially synthesize the full-length gene and clone it into pET-43.1a prokaryotic expression vector, induced expression in BL21 (DE3) strain of Escherichia coli( E. coli), and used Western blot to detect its reactogenicity. Results:The results showed that after induction, SDS-PAGE electrophoresis analysis showed that an obvious expression band appeared at a molecular weight of 50×10 3, which was consistent with the expected protein band size. Among them, the E. coli concentration A600 is about 0.5, and the expression yield is the highest (about 32.3%) when induced at 37℃ for 5 h. Nucleoprotein expression product is mainly inclusion body when it is expressed in large quantities. After purification by Ni 2+ chelating chromatography, the purity of the target protein can reach over 95%. The purified product was identified by Western blot and positively reacted with the sera of mice immunized with rabies vaccine, indicating that the prokaryotic expression of the CTN-1 strain nucleoprotein has biological activity. Conclusions:This experiment successfully established a high-efficiency expression method for the nucleoprotein of the CTN-1 strain in the prokaryotic expression system, and obtained high-purity target protein, which provides a basis for further clinical diagnosis and preparation of new vaccines.

Objective:To analyze the delay in the publication of medical postgraduates' research papers and provide them with a guidance for the management of academic articles.Methods:A total of 293 articles were chosen from the Journal of Medical Postgraduates published from 2005 to 2006,of which,229 were original articles and 45 were reviews,including 184 funded and 109 non-funded research papers,and their delay in publication in different periods was statistically analyzed by SPSS13.0 software.Results:The total delay in the publication of the articles was(234.7 ? 116.9) d,including(57.0?36.8) d in manuscript examination,(109.4 ? 96.4) d in editing and(61.8 ? 52.4) d in suspension.The original articles were less delayed than the reviews,(224.2 ? 7.4) d vs(272.8 ? 19.3) d,P

Objective To explore the effect of hyperbaric oxygenation (HBO) on generation of free radicals and the permeability of blood-brain barrier(BBB) during cerebral ischemia-reperfusion(CIR). Methods Three hundred and twenty Kunming mice were randomly divided into four groups: a sham operation group, a HBO group, a CIR group, and a HBO + CIR group, with 80 mice in each group. Conscious mice were used to establish the CIR model, and 0.25MPa (ATA) HBO were applied 5 times after operations. The activities of SOD, CAT, GSH-PX and the concentration of MDA, EB in the cerebral hippocampal tissues in each group were measured with colorimetry. The cerebral hippocampal tissues were harvested and processed, then observed and compared with transmission electron microscopic observation. Results Compared with the sham operation group, the activities of antioxidation enzymes (GSH-PX,SOD, CAT) in CIR group decreased significantly (P

Objective To investigate the effects of ketamine combined with moderate hypothermia on brain ischemia-reperfusion (I/R) injury in a rat model of asphyxial cardiac arrest. Methods Fifty healthy Wistar rats of both sexes aged 4.0-4.5 months, weighing 410-510 g were randomly allocated into 5 groups (n = 10each): group Ⅰ sham operation (group S), group Ⅱ asphyxial cardiac arrest (group ACA), group Ⅲ ketamine (group K), group Ⅳ moderate hypothermia (group MH) and group Ⅴ K + MH. The animals were anesthetized with intraperitoneal (IP) phenobarbital 20 mg/100 g, tracheostomized and mechanically ventilated (RR 60 bpm,FiO2 50%), PaCO2 was maintained at 35-45 mm Hg. Cardiac arrest was induced by clamping tracheal tube until ECG activity disappeared and MAP ＜ 15 mm Hg. Resuscitated was started 5 min later. MAP ＞ 60 mm Hg and HR ＞ 250 bpm were considered to be signs of successful resuscitation. Dead animals and animals in which resuscitation time was longer than 5 min were excluded from the study. In group K ketamine 100 mg/kg was administered IP at 5 min before asphyxia. In group MH hypothermia was started as soon as asphyxia was started and body temperature was maintained at 30-35 ℃. After successful resuscitation, the animals were sacrificed. Their brains were removed for determination of brain water content and p-caspase-3 expression in hippocampus. Results Brain I/Rsignificantly increased brain water content and p-caspase-3 expression in group ACA. MH alone significantly attenuated 1/R-induced brain edema and decreased p-caspase-3 expression, while ketamine alone only significantly decreased p-caspase-3 expression but did not decrease I/R-induced brain edema. MH + K decreased p-caspase-3expression further but did not reduce brain edema further as compared with MH alone. Conclusion Ketamine combined with moderate hypothermia provides better protection against brain I/R injury.

We constructed prokaryotic recombinant expression vector of P61 gene from Nocardia brasiliensis,expressing P61 protein with biological activity in E.coli,and lay a foundation for further studies related to P61.P61 gene was synthesized and cloned into an expression vector pET-30a(+).The recombinant vector was transformed into Escherichia coli BL21 and induced with IPTG.The production was analyzed with Western blot and the catalase activity of P61 was tested with Catalase Assay Kit.The protein of P61was successfully expressed in E.coli with solubility and high catalase activity,and could be identified by anti-N.brasiliensis sera from mice.The prokaryotic expression plasmid of protein P61 was constructed successfully and can be expressed efficiently in E.coli BL21 cells with higher catalase.

In view of problems such as non-innovative,useless and non-standard topic in topic-chosen stage of postgraduate of biomedical engineer,the countermeasures are discussed in aspects of aim,standpoint and methods of topic-chosen to improve its scientificity,feasibility and utility,thus providing a good beginning for postgraduates to perform scientific research and apply the result to practice.[Chinese Medical Equipment Journal,2008,29(3):105-106]

Objective:The recombinant shuttle peptide-single chain antibody with neutralizing activity is expected to be obtained by combining the shuttle peptide sequence with rabies single chain antibody and optimizing the expression conditions in E. coli.Methods:In this study, the SO57 single-stranded antibody sequence was recoded and the shuttle peptide of Arg 12 was fused at the N-terminal to construct pET22 (b)-rSO57 expression vector, which was then expressed in E. coli. The growth density OD 600, induction time, induction temperature and inducer concentration were optimized to obtain a higher expression effect. The recombinant protein was purified by immobilized metal chelating chromatography, and the relative affinity of the recombinant single-chain variable fragment (scFv) was determined. The neutralization effect of the recombinant scFv was verified by rSO57/CVS-11 virus strain mixed with infected cells and rapid fluorescent focus inhibition test (RFFIT). Results:The recombinant rSO57 was successfully constructed and induced in BL21 (de3). The optimal expression condition was that when the cell density of OD 600 was 0.8, IPTG of 0.4 mmol/ml was used for induction. After induction at 16 ℃ for 24 hours, rSO57 was expressed in soluble form, accounting for 19.8% of the soluble protein in the cell. After chromatography purification, the recombinant rSO57 with a purity of 84% was obtained. ELISA showed that the relative affinity coefficient Ka of rSO57 was 4.3×10 5. When rSO57 was mixed with CVS-11 strain, the infection rate of cells increased with the increase of dilution, indicating the neutralization activity of recombinant antibodies. Using the RFFIT, 50 μg rSO57 was equivalent to 2.17 IU of rabies neutralizing serum. Conclusions:In this study, the recombinant scFv fusion with shuttle peptide was expressed, which could neutralize rabies virus, in order to prepare specific and targeted antiviral drug carriers for rabies treatment.

Total aralosides of Aralia elata (Miq) Seem (TASAES) from Chinese traditional herb Longya Aralia chinensis L was found to improve cardiac function. The present study was to determine the protective effects of TASAES on diabetic cardiomyopathy, and the possible mechanisms. Therefore, a single dose of streptozotocin was used to induce diabetes in Wister rats. Diabetic rats were immediately treated with low, medium and high doses of TASAES at 4.9, 9.8 mg/kg and 19.6 mg/kg body weight by gavage, respectively, for eight weeks. Cardiac function was evaluated by in situ hemodynamic measurements, and patch clamp for the L-type Ca2+ channel current (ICa2+-L) and transient outward K+ channel current (Ito). Histopathological changes were observed under light and electron microscope. The expression of pro-fibrotic factor, connective tissue growth factor (CTGF) was monitored using immunohistochemistry staining. Compared with diabetic group, medium and high doses, but not low dose, of TASAES showed a significant protection against diabetes-induced cardiac dysfunction, shown by increased absolute value of left ventricular systolic pressure (LVSP) and maximum rates of pressure development (+/-dp/dt(max)), and enhanced amplitude of ICa2+-L (P < 0.05). Histological staining indicated a significant inhibition of diabetes-caused pathological changes and up-regulation of CTGF expression (P < 0.05). The results suggest that TASAES prevents diabetes-induced cardiac dysfunction and pathological damage through up-regulating ICa2+-L in cardiac cells and decreasing CTGF expression.
Animals ; Aralia/*chemistry ; Calcium Channels, L-Type/physiology ; Cardiomyopathies/*drug therapy/etiology/physiopathology ; Connective Tissue Growth Factor/metabolism ; Diabetes Mellitus, Experimental/*complications ; Drugs, Chinese Herbal/*chemistry ; Heart/drug effects/physiopathology ; Hemodynamics ; Male ; Myocardium/pathology ; *Oleanolic Acid/analogs & derivatives/therapeutic use ; Patch-Clamp Techniques ; Potassium Channels/physiology ; Rats ; Rats, Wistar ; Saponins/*therapeutic use ; Treatment Outcome

Objective:To investigate the in vitro viability of rabies virus in tissues and body fluid samples. Methods:The viability of rabies virus in tissues and suspensions was analyzed by virus titer determination method, direct immunofluorescence, RT-PCR and laboratory techniques for virus isolation.Results:With the increase of temperature, the viability of rabies virus in brain tissues and suspensions decreased gradually. Rabies virus lost infectivity after 30 min at 56℃, but remained viable in tissues for 7 d at 37℃. The virus showed no viability after 1 h at pH9.6. The rabies virus in suspensions could be completely inactivated after the stimulation with ethanol at a final concentration above 30%, sodium hypochlorite above 500 mg/L or benzalkonium bromide above 100 mg/L for 3 min. It was found that 80% acetone had the strongest inactivation effect on rabies virus in tissues, and no virus could be isolated after soaking for 4 h.Conclusions:Rabies virus was not tolerant to high temperature and relatively stable in the environment with pH6.8-7.4. Common disinfectants could kill the virus. This study provided detailed data about the viability of rabies virus in vitro, which would be conducive to the prevention and control of rabies.