According to bias in codon choice of Pichia pastoris, The phytase phyA gene from Aspergillus niger N25 was mutated without changing its amino acid sequence. The expression plasmid pPIC9k-phyAm was constructed and transformed into GS115 strain. Positive clones,of which the chromosomes were integrated with phyA gene,were identified by the phenotype and PCR. SDS-PAGE analysis suggust that the size of enzyme protein of the expression product was about 70.15kDa.Southern blotting analysis to the yeast transformants showed that phyA gene was intergrated into the chromosome genome. The phytase activity of PP-NP m-4-4 with codons optimized reached 136 000U/ml in malt wort culture medium after being induced with 36h, which was the 2.8 times of the original strain PP-NPm-8.

Objective To discuss the effect and feasibility of BiPAP in refractory asthma patients. Methods A total of 15 coses of refractory bronchial asthma admitted by our hospital during Feb 2003 to May 2006 were treated by BiPAP in addition to medical therapy。 The symptoms、signs and the arterial blood gases were observed and analyzed。Results Among 15 patients,2 gave up treatment,13 patients were successfully treated with the BiPAP therapy。There were significant improvement of PaCO2、PaO2、pH and respiratory rate ,heart rate after 2h BiPAP。Conclusions Ventilation in BiPAP is an effective method for refractory asthma ,which can save the patients life and decrease the complications.

A method was established for the simultaneous determination of the total fatty acid esters of chloropropanols in edible oils by gas chromatography-mass spectrometry combined with isotope dilution technology. The samples were hydrolyzed with sodium methylate-methanol, and then purified by diatomite cartridge. After being derivatized with heptafluorobutyrylimidazole ( HFBI ), the target analytes were determined by GC-MS with the deuteriumchloropropanols esters as the internal standards. An excellent linear correlation in the range of 0. 050-2. 000 mg / L was acquired for 3-monochloropropane-1,2-diol (3-MCPD) esters, 2-MCPD esters, dichloropropan-2-ol (1,3-DCP) esters and 2,3-dichloropropan-1-ol (2,3-DCP) esters, with all the correlation coefficients (r) higher than 0. 9995. The limits of detection (LODs) for 3-MCPD esters, 2-MCPD esters, 1,3-DCP esters and 2,3-DCP esters were 0. 015, 0. 015, 0. 030, and 0. 030 mg / kg, respectively, and the limits of quantitation (LOQ) were 0. 050, 0. 050, 0. 100, and 0. 100 mg / kg, respectively. The average spike recoveries of the four kinds of chloropropanols esters in blank extra virgin olive oil matrix were typically in a range of 87. 0% -110. 5% with the relative standard deviations (RSDs) less than 10. 1% . The detection rates of 3-MCPD esters, 2-MCPD esters, 1,3-DCP esters and 2,3-DCP esters in 74 edible oil samples were 94. 6% , 63. 5% , 5. 4% , and 0% , respectively. The contamination levels of 3-MCPD esters, 2-MCPD esters and 1,3-DCP esters were in the range of not detected (ND) to 10. 646 mg / kg, ND to 3. 617 mg / kg and ND to 0. 089 mg / kg, respectively. This method is accurate and rugged for the simultaneous determination of total fatty acid esters of chloropropanols in edible vegetable oils.

Objective To investigate the therapeutic effect of ambroxol in bleomycin-induced pulmonary fibrosis of rats.Methods Sixty Wistar rats were divided into three groups:The normal control group(group N),the model group treated with bleomycinA5(group B)and the group treated with ambroxol(group A).Pulmonary fibrosis was induced by intratracheal instillation of bleomycin.Then the rats received daily ambroxol 35mg/kg(group A),or normal saline(group B).Five rats in each group were killed on 3.7.14 and 28 days after intratracheal instillation.Histological changes of the lungs were evaluated by HE stain and Masson's trichrome stain.The levels of tumor necrosis factor-?(TNF-?)in lung homogenates were measured by radioimmunoassay.Results Pulmonary fibrosis of Group A was significantly improved as compared with that of Group B(P

Objective To discuss the effect and possibility of NIPPV in ILD combined with respiratory failure. Methods A total of 37 cases of ILD combined with respiratory failure admitted by our hospital during Feb 2004 to Oct 2007 were divided into treatment group (20 cases) and control group (17 cases). Routine pharmaceutical intervention was adopted in both groups while NIPPV was given in the patients of treatment group. The symptoms、signs and the arterial blood gases were observed and analyzed. Results Among 20 patients,3 gave up treatment , 17 patients were successfully treated with the NIPPV therapy. There were significant improvement of PaCO2、PaO2、pH and respiratory rate ,heart rate after 2h NIPPV. Conclusion NIPPV is an effective method for ILD combined with respiratory failure,which can save the patients life and decrease the complications.

Objective To investigate the relationship between the apoptosis,expressions of Bcl-2 and Bax protein in perihematomal brain regions of rats and neurologic dysfunctions after intracerebral hemorrhage (ICH). Methods Seventy Sprague-Dawley rats were divided randomly into two groups:an experimental group and a control group.A model of ICH was established by injection of 0.5 U bacterial collagenaseⅦinto the caudate nucleus in the rats.Neurological impairment was evaluated at 6 h,12 h,24 h,48 h,72 h,7 d and 14 d after 1CH,respectively, before the rats were sacrificed.TUNEL method was used to detect apoptosis,and SP method to detect expressions of Bcl-2 and Bax protein in the perihematomal brain tissues.Results Neurological impairment occurred in all the rats after ICH,and peaked at 48 h after ICH.The apoptosis and expressions of Bcl-2 and Bax protein were peaked at 48 h,6 h and 48 h after ICH,respectively.Conclusion The degree of the neurological impairment after ICH is parallel to that of the apoptosis.Apoptosis may play an important role in neurological impairment after ICH.

Objective:To prepare the first extracellular domain of human ? chemokine receptor 5(huCCR5)NH 2 terminal's specific antibody F(ab′) 2 and its detection method Methods:Use computer analyzed and located the least homologous domain of the extracellular first loops The gene fragment was amplified by PCR and cloned into the pGEX IN vector A recombinant GST fusion protein was constructed After comfirming the correctness of the inserted sequence,the transformation and expression of this fusion protein were performed in E coli The expression products of the fusion protein were purified and 2 New Zealand rabbits were immunized An anti huCCR5 NH 2 terminal antibody F(ab′) 2 was prepared by protein A Sepharose CL 4B affinity chromatography ,pepsin digestion and S 200 column seperation.Results:Reduced,unreduced SDS PAGE and ELISA block examination analysis demonstrated that this F(ab′) 2 had high specificity to combine with huCCR5 Conclusion:In this paper,not only introduce a simple and quick method to get a specific antibody F(ab′) 2 of certain functional domain but also a good idea and technique to study other high similar superfamily members

Ashi point (tonsil) in combination of the pricking and bleeding technique on Shaoshang (LU 11 )and Shangyang (LI 1) were used to treat 58 outpatients of acute tonsillitis. The results showed cure in 38 cases,remarkable effect in 17 cases and failure in 3 cases by one treatment, and the total effective rate in 95%.

Objective To evaluate the academic status of the researchers in pediatric department of Peking university first hospital, we conducted bibliometric analysis of research on the scientific publications. Methods The total number, first authors, and the citing and cited conditions of these papers published from 1999 to 2000 were analyzed by bibliometric methods. Results There were total 513 papers including 275 original articles published in 59 journals from 1999 to 2003.There were 238 papers written in Chinese, of which 188 papers (79 %) were from the core journals. In the 37 original articles written in English, 31 papers(83.8 %) were select-ed by science citation index(SCI).The average of impact factors (IF) of sourcing journals was 1.447(0.107 -7.717). The averages of IF per year ranged from 0. 768 to 2.206.The original articles selected by SCI were totally cited 38 times. Every article was cited 1.41 times in average.Of all 275 papers,each had an average of 9.8 citing reference, increasing from 8.1 in 1999 to 12.0 in 2003.About 80.7 % of those references were written in foreign languages.The Price's index of those papers was 43.2%.Conclusions The number of papers kept improving in the past 5 years, and had a significant improvement in 2003.The authors showes great talents to use English articles as their main information sources. But the utilization of the last published papers in this subject should be im-proved.

Objective To evaluate the clinical effectiveness in screening asymptomatic diabetic peripheral neuropathy(ADPN) by the Michigan neuropathy screening instrument (MNSI) and the Toronto clinical scoring system(TCSS).Methods MNSI,TCSS and neural electrophysiological test (NET) were conducted in 232 neurologically asymptomatic type 2 diabetes patients.By using the results of NET as the golden criteria for diagnosis of ADPN,we evaluated the effectiveness of the two different scoring system by the receiver operator characteristic curve.The sensitivity,specificity,positive and negative predictive values,accuracy,Youden indexes and kappa values on different diagnostic cut-off points of MNSI and TCSS were analyzed.The correlation between the two different scoring system and the risk factors of diabetic peripheral neuropathy (DPN) were also analyzed.Results The area under the ROC curve of MNSI and TCSS were 0.792,0.704,respectively.The sensitivity,specificity,accuracy,Youden indexes and kappa values of MNSI over 2 and TCSS over 2 were 66.2％ vs 73.3％,90.4％ vs 63.7％,78.3％ vs 68.5％,0.566 vs 0.370,and 0.588 vs 0.345,respectively.MNSI was better than TCSS in the effectiveness of diagnosing ADPN and consistence with the result of NET.Moreover,MNSI was associated with the most related risk factors of DPN including age,glycosylated hemoglobin (HbA1c),HbA1c × disease duration,islet function and HDL-C.Conclusions MNSI could be used as a relatively simple and reliable method for clinical and epidemiological screening and assessment of ADPN.