Objective:To dectect the effect of granulysin and IL-12 genes'expression products on proliferation and apoptosis of melanoma B16 cell in vitro.Methods:Co-expression plasmid including granulysin peptide and murine interleukin 12(mIL-12)genes was transfected into melanoma B16 cell with Lipofectamin TM2000 and its expression products were detected by RT-PCR.Growth suppression was detected with MTT colorimetric assay,and cell apoptotic alterations were evaluated by Hoechst 33258 staining,AO/EB staining,and Annexin V-FITC flow cytometry(FCM).Results:GLS peptite and IL-12 genes could be expressed in B16 cells.Expression products inhibited the proliferation of melanoma cells under MTT observaton.Cells apoptosis with nuclear chromatin condensation,fragmentation and cell membrane change were observed under Hoechst 33258 staining and AO/EB staining.FCM analysis showed the apoptotic rates in test group was 21.02%,which was higher than that in control in control group(15.57%).Conclusion:Expression products of granulysin and mIL-12 genes can not only inhibit proliferation but also induce apoptosis of murine melanoma cell line B16 in vitro.

Objective To apply Gukang gold far-infrared magnetic therapeutic paste to the treatment of lateral epicondylitis.Methods 64 cases of lateral epicondylitis patients were randomly divided into treatment group and control group,each with 32 ones.The subjects from the treatment group were treated with Gukang gold far-infrared magnetic therapeutic paste,and the remained with ibuprofen sustained-release capsules.The results after one and two courses were analyzed statistically.Results After two-course treatment,the patient from the treatment group had an higher improvement rate than the others from the control group,with a P value less than 0.05.The treatment course of the treatment group was shorter than that of the control group,with a P value less than 0.05.Conclusion When compared with ibuprofen sustainedrelease capsule,Gukang gold far-infrared magnetic therapeutic paste is more effective for the treatment of lateral epicondylitis.

Objective To evaluate the results of patients operatively treated transverse plus posterior wall fractures of the acetabulum.Methods Review forty-five patients who had operated for transverse plus posterior wall fractures of the acetabulum 1 retrospectively with fracture displacement,from August 1993 to January 2005 in Department of Orthopaedics,Beijing Shunyi Hospital.The radiographs were graded according to the criteria described by Matta.The functional outcome was evaluated using a modification of the clinical grading system developed by Merle d'Aubigné and Postel.Result Forty-five patients were followed up 16 to 48 months with an average of 34 months.The radiographic result was excellent in seventeen patients,good in eighteen,fair in seven,and poor in four.The clinical outcome at the time of final follow- up was graded as excellent in fourteen patients,good in twenty- two,fair in eight,and poor in two.Conclusion Operative treatment for transverse plus posterior wall fractures of the acetabulum has a satisfying therapeutic effect.The appropriate operation time,reasonable operation approach,anatomic reduction and stable internal fixation is the key to obtain good results.

This article aims at exploring ways of integrating subject of pathobiology under the guidance of scientific outlook on development and improving and enriching pathobiology inte-gration and construction through curriculum systems recombination and integration,teaching con-tents optimization to reflect the latest research progress,network education platform establish-ment,comprehensive and exploratory laboratory course teaching system construction,new syl-labus and teaching material making up.

Background and purpose:To explore the possibility of genetic re-expression silenced by DNA aberrant hypermethylation which is a common epigenetic modification in carcinogenesis. 5-Aza-CdR, an inhibitor of DNA methylation, was used to determine the effects of expression of tumor suppressor gene E-cadherin in tumor cell lines. Methods:Methylation specific PCR(MSP) was utilized to examine methylation status of E-cad gene on breast carcinoma cell line MDA-MB-435 before and after the treatment with 5-Aza-CdR. Immunohistochemistry(IHC) was used to test the expression of E-cad protein. Semi-quantitative RT-PCR method was used to detect the changes of E-cad mRNA.Results:1).E-cad methylation was positive(116bp) and unmethylation was negative on MDA-MB-435 cell before the treatment with 5-Aza-CdR. After being treated with 5.0umol/L 5-Aza-CdR for 3 days, methylation turned negative and unmethylation positive bands(97bp) were detected. 2).The E-cad protein expression was not detected by immunohistochemistry on MDA-MB-435 cell before the treatment, while E-cad staining was positive on the cell membrane after the treatment. 3). The E-cad mRNA failed to be amplified in cells before the treatment. After incubation at variable concentrations of 0.5 ?mol/L, 1.0 ?mol/L, 2.0 ?mol/L and 5.0 ?mol/L 5-Aza-CdR for 3 days, respectively, E-cad mRNA expression was detected on the fourth day in a dose-dependent manner. Correlation between the mRNA expression level and the agent concentration was observed.Conclusions:The demethylation agent 5-Aza-CdR can reverse the aberrant E-cad methylation status in MDA-MB-435 and re-expressed E-cad mRNA and protein.

Objective To construct an eukaryotic co-expression plasmid pBM9 carrying human granulysin active peptide and murine IL-12 and determine its expression.Methods The primer pairs including granulysin leader peptide DNA sequence were designed to amplify granulysin from the plasmid pZM03 carrying granulysin gene by polymerase chain reaction(PCR).PCR product was directly cloned into an eukaryotic co-expression plasmid pBudCE4.1 to construct plasmid pBudCE4.1-S9K.pBudCE4.1-S9K plasmid was identified by DNA sequencing.Murine IL-12 gene was subcloned into pBudCE4.1-S9K to construct eukaryotic co-expression plasmid pBudCE4.1-S9K/mIL-12.Mycobacteria replicon Orim from plasmid pZM03 was subcloned into NotⅠsite of pBudCE4.1-S9K/mIL-12 to construct eukaryotic co-expression shuttle plasmid pBM9.pBM9 was tansfected into RAW264.7 cells.RT-PCR was used to detect the mRNA expressions of granulysin and IL-12.The protein expression of them were observed by immunocytochemical method and ELISA respectively.Results RT-PCR identified the expressions of granulysin and mIL-12 in transfected cells and culture supernatant.Immunocytochemical method and ELISA verified the protein expressions.Conclusion The recombinant shuttle plasmid pBM9 is successfully constructed and expressed in vitro,which laid a foundation of gene therapy for tumors with granulysin and mIL-12.

Pathogen biology is a main course of curricula in medical university.In developing quality courses,we optimized Pathogen biology course content,created the PBL teaching mode suitable for this course,set up network teaching platform,enhanced teachers' qualification,constructed comprehensive and exploratory experiment teaching system,wrote new teaching syllabus and other measures,made the Pathogen biology course construction more reasonable and perfect.

Objective; To clone, sequence and analyze the coding sequences of the Mr 15000 and Mr 9000 active segments of natural granulysin derived from human CTLs activated by allogenic antigen ,in order to establish the basis for further purifying and investigating the immune - impairing mechanism of granulysin. Methods; The coding sequences of the Mr 15000 and Mr 9000 granulysin gene were amplified from the total RNA of activated CTLs of healthy person after reverse transcription by Nested - PCR, inserted into pET32a ( + ) vectors and then transformed into E. Coli TOP10, respectively. The recons were identified by PCR, endonuclease digestion and sequencing. Results: The whole coding sequences of the Mr 15000 and Mr 9000 active segments were successfully cloned as expected. The accurate pET32a - Mr 15000 and pET32a - Mr 9000 recons were obtained through Colony - PCR, endonuclease digestion and sequencing. There lied polymorphism on the 119 th amino acid of the product encoded by GLS gene. Conclusion; The coding sequences of the Mr 15000 and Mr 9000 active segments of human granulysin can be obtained and cloned by the methods mentioned above and can be used for subsequent research.

To fully incarnate"specialization"peculiarities of microbiology teaching courses by means of optimizing microbiology teaching contents and curriculum systems,and making up a new syllabus suitable to different specialties will lay a very solid foundation for cultivating medical talents with specialty traits.

AIM: To study and establish the fingerprint of raw Herba Polygoni Orientalis by RP-HPLC. METHODS: The chromatographic conditions were as follow: an Inersil-ODS-3 column was used;the mobile phase was composed of acetontrile and 0.1% phosphoric acid with gradient elution;the flow was 1.0 mL?mim~(-1) and the UV absorbance detection was set at 300 nm. RESULTS: Under the selected chromatographic conditions. Similarity of 10 batches of good HPLC fingerprint of raw Herba Polygoni Orientalis were obtained. no less than 0.9.( CONCLUSION): Quality of raw Herba Polygoni Orientalis can be controlled effectively by HPLC-UV fingerprint.