OBJECTIVE To explore the clinical distribution and drug resistance of extended-spectrum ?-lactamases(ESBLs) producing Enterobacter cloacae.METHODS The clinical distribution and drug resistance of 73 strains of E.cloacae and ESBLs producing E.cloacae identified in our hospital from Jan 2002 to Jan 2004 were analyzed.RESULTS Among them,the highest detectable rate was in respiratory department and from phlegm specimen.No ESBLs producing E.cloacae and non-(ESBLs) were resistant to imipenem;no non-ESBLs were resistant to cefepime,but some ESBLs producing E.cloacae was resistant to it;all ESBLs producing E.cloacae and non-(ESBLs) were resistant to ceftriaxone and amoxicillin/clavulanic acid.CONCLUSIONS E.cloacae is one of the(commonest) pathogens in nosocomial infection,and ESBLs producing E.cloacae has a high detectable rate,its drug resistance has increased;the clinic should choose antimicrobial agents rationally according to drug sensitive tests in vitro.

AIM: To construct engineered E.coli strains which can express nattokinase with fibrinolysis activity using gene engineering technology. METHODS: The pro-nattokinase (pro-NK) gene was amplified by PCR and inserted into expression vector pET3c. The recombined plasmid pENK which expressed the fusion protein of pro-NK and 22 amino acid peptide was then transferred into lysogenic host strains BL21(DE3)pLysS - and BL21(DE3)pLysS +. Both SDS-PAGE and the fibrin plate assay were used to examine the expression and the activity of the target protein. RESULTS: SDS-PAGE assay showed the fused gene encoding 42 kD fusion protein was expressed in both expression strains pENK-(DE3)pLysS - and pENK-(DE3)pLysS +, and the fibrin plate assay indicated that the expression product had fibrinolysis activity. pENK-(DE3)pLysS - exhibited the basal expression of the target gene, while fusion protein was only induced by IPTG in pENK-(DE3)pLysS +. Basal expression of the fused toxic gene in pENK-(DE3)pLysS - led to bacteriolysis and hollow lawns. CONCLUSION: A pro-NK fusion protein with fibrinolysis activity is successfully expressed in E.coli , which lay a foundation for the exploitation of nattokinase.

To improve the expression level of non fusion hbFGF in E coli , the coding sequence of human bFGF gene, which had been cloned from primarily cultured human fibroblast, was mutated according to the principle of lowering the GC content and increasing the codon preference After being ligated into pET 3c and transformed into BL21(DE3), the recombinant induced by IPTG Expression level was up to 30% of the total bacterial protein The result indicated that optimizing of the TIR would promote the expression level of recombinant protein

Objective To investigate the correlation between serum cystatin C (CysC) levels and cerebral microbleeds (CMBs) in patients with acute ischemic stroke.Methods The patients with acute ischemic stroke were enrolled.Susceptibility weighted imaging was used to identify the presence of CMBs.Particle-enhanced turbidimetric immunoassay was used to detect the levels of serum CysC.Results A total of 485 patients with acute ischemic stroke were enrolled,including 151 (31.1％) with CMBs.The level of serum CysC of the CMB group was significantly higher than that of the non-CMB group (1.24± 0.13 mg/L vs.1.02± 0.11 rmg/L; t=4.261,P＜ 0.001).Multivariate logistic regression analysis showed that the increased serum CysC level was an independent risk factor for the presence of CMBs in patients with acute ischemic stroke (each increase in one standard deviation,odds ratio 4.063,95％ confidence interval 2.142-8.127; P ＜0.001).Multiple linear regression analysis showed that the number of CMBs in patients with acute ischemic stroke increased with the increasing serum CysC level after adjusting for other confounders (r2 =0.361,P =0.017).Conclusions In patients with acute ischemic stroke,the serum CysC levels are independently associated with the CMBs,and the number of CMBs increases with the serum CysC level.

Objective By comparing the change of expression of AQP1 in vitro lung preserved by the continuous infusion of a heart‐lung machine ,continuous pressure perfusion and single low temperature ,to explore the best method of vitro lung preserva‐tion .Methods Thirty Mongrel dogs were randomly divided into 3 groups ,and both lungs were completely resected under the condi‐tion of keeping mechanical ventilation .The vitro lungs were preserved by the way of the continuous infusion of a heart‐lung ma‐chine ,continuous pressure perfusion and single low temperature ,and collecting specimens according to the time point .HE staining was used to observe the morphological changes of vitro lung tissue .Immunohistochemistry and Western blot were used to detect the expression of AQP1 in vitro lung .Results HE staining found that as the time went by alveolar structure gradually collapsed ,in‐flammatory cells increased ,alveolar interval also gradually broadened and exudation could be seen in the alveolar cavity ;at the same time point ,organization structure of extracorporeal circulation group changed lighter than pressure perfusion group and low‐temper‐ature preservation group .In each experimental group ,the expression of AQP1 showed a trend of decline;at each time point ,the ex‐pression of AQP1 in extracorporeal circulation group was higher than pressure perfusion group ,and pressure infusion group was higher than that of low‐temperature preservation group .Conclusion The protective effect of the continuous infusion of a heart‐lung machine on vitro lung was better than continuous pressure perfusion and single low temperature .

Objective To investigate the effect of simvastatin on gene expression of L-type calcium channels(LCCs) of myocardial cells in BALB/c mice infected by coxsackievirus B3(CVB3) so as to study the therapeutic effect of simvastatin on viral myocarditis.Methods Sixty male BALB/c mice were divided into five groups randomly(n=12).Mice in viral control group and three groups of oral administration of simvastatin(10,30 and 90 mg?kg-1)were inoculated intrapritoneally with 0.2 mL of CVB3(Nancy strain).Mice in normal control group were inoculated intrapritoneally with 0.2 mL Eagle's solution.The heart samples of all the mice were obtained for hispathological study and detection of myocardial LCCs alpha1 subunits mRNA expression by semi-quantitative reverse transcription-polymerase chain reaction(RT-PCR).Results In viral control group,the mononuclear inflamematory infiltrate was focal or diffuse in myocardium of mice,severe hearts revealed a large area of myocardial necrosis.The degree of inflammatory cell infiltrate and area of necrosis were significantly less in simvastatin groups as compared with viral control group.The myocardial LCCs alpha1 subunits mRNA expression by semi-quantitative RT-PCR in normal control group was much lower than that in viral control group(0.06?0.01 vs 1.37?0.32,P

By the survey of topic selection of medical master's dissertation for candidates with the same educational level,the writers think that the decided topics should be in close relation to their respective subjects and the operational and technical conditions of their institutes and the training institutes should be taken into full consideration.The decided topics should enhance theoretical and practical values and will be achieved in the required time as well.

Objective To explore the methods and standards of catastrophic health expenditure for China,and improve the operability in defining those impoverished due to diseases.Methods 600 households with inpatients were randomly sampled from 12 villages of three townships in City M in Hubei province for household survey.Literature review and expert consultation methods were used to measure and determine the threshold of catastrophic health expenditure( CHE) of City M,with improvements made by grouping.Results In a uniform standard,the CHE occurrence of the income method and expenditure method is lower than the non-existence expenditure method and WHO method,verifying theoretically that the standard set for the former should be higher than the latter.In addition,the income method and expenditure method present differently in China.Standards set should vary with different measurement methods,maintaining approximate CEH expenditure for the same region.For the income method and expenditure method,the recommended benchmark reference is 25%,with an adjustable range of 35%~45%.Mutual verifications of these methods can further determine a rational CHE value.Conclusions The CHE methods support the definition of people impoverished due to diseases beyond their household payment capacity.Different thresholds however should be set for different methods,and the relative standards should be translated into absolute standards by grouping.

Objective: To establish the quality control standards for Shimingbao Granules.Methods:Radix Angelical Sinensis, Cortex Phellodendri and Pericarpium Citri Reticulatae in granules were identified by TLC. The content of paeoniflorin in Shimingbao Granules was determined by HPLC with external standard. Results:The average recovery was 97.54% and RSD was 1.7%( n =5). The RSD in a duplicate test was 1.7%( n =5). Conclusion: These methods are simple, accurate and specific and can be used for the quality control of Shimingbao Granules.

AIM: To get high biological activity of recombinant human bone morphogenetic protein -2 (rhBMP-2) expressed in Escherichia coli by the methods of refolding. METHODS: The rhBMP-2,expressed in Escherichia coli, was washed by Triton X- 100 and further purified by DEAE chromatography.The inclusion bodies were resolved in 8 mol/L urea, and were refolded and dimerized in the redox systems (reduced and oxidized glutathione). Finally, a one - step purification procedure based on the heparin affinity chromatography was implemented. The biological activity of purified rhBMP - 2 were tested by induction of the alkaline phosphatase activity in C2C12 cells. RESULTS: The rhBMP - 2 was expressed in Escherichia coli in a non - active aggregated form of inclusion bodies using a temperature - inducible expression system. The high - purified rhBMP - 2 was obtained in the form of inclusion bodies by several purification courses. The rhBMP - 2 was refolded and dimerized in the redox systems (reduced and oxidized glutathione) and a one - step purification procedure based on the heparin affinity chromatography was implemented to isolate the rhBMP - 2 dimers and monomers. The purified rhBMP - 2 dimers showed the higher biological activity than the commercial rhBMP - 2. CONCLUSIONS: The method achieved the refolding of rhBMP - 2 would be applied to the whole TGF - β superfamily because the BMP - 2 belongs to the superfamily. Meanwhile, the inexpensive,high yield rhBMP - 2 is suitable for clinic application.