Objective To explore the changes of human mitochondrial COXI,ND1 and ND6 genes expression induced by ionizing irradiation.Methods Changes of human COXI,ND1 and ND6 gene expression were detected by RT-PCR and Real-time PCR 8 h after the irradiation in human lymphoblastoid cell lines,which were exposed to 1-10 Gy 60Co γ-rays.And the dose-effect relationships between expression changes of the genes and the doses were analyzed.The changes of these three genes expression were also analyzed at different post-radiation time-points between 0.5 h and 72 h after irradiation of 5 Gy in order to explore the time-effect.Results The expression of three genes COXI,ND1 and ND6,showed either the dose-effect or the time-effect after irradiation.The gene expression levels of three genes up-regulated generally and the peak change time-point was 4 h after irradiation.Conclusion Ionizing radiation,msht induce the changes of mitochondrial gene expression,and the gene expression level is up-regulated.

Objective To investigate the effect of mixed application of lecture-based learning and flipped classroom and problem-based learning in microbiology teaching. Methods The five-year program students of Grade 2013 and Grade 2014 were randomly divided into two groups, and the experimental group (48 students in the teaching reform class) carried out the blended teaching, while the contrast group (48 students of parallel class) carried on the traditional teaching. After the lecture, the teaching effect of the hybrid teaching method and traditional teaching method was analyzed after the theoretical and the experi-mental operation test were adopted, and the four part questionnaire surveys including the integrated use of knowledge, active classroom atmosphere, innovation ability and teaching satisfaction were proceeded in each group. The data of each group was analyzed by t test analysis with SPSS 19.0 respectively. Results The theory test scores of experimental group was (90.16±3.14), which was higher than the control group (82.33± 4.21). The difference between them was very significant (P=0.000). Survey results showed that the integrated use of knowledge, active classroom atmosphere, innovation ability and teaching satisfaction were higher than traditional group. The difference was statistically considered significant (P<0.01). Conclusion Hybrid teaching method is very good for training and development of students' comprehensive quality and ability. It has important significance in improving the students' score.

Objective:To construct GM-CSF and EB virus LMP2A fusion gene recombinant adenovirus and identificate it , using recombinant adenovirus to do immunologic study.Methods:With RT-PCR and Western blot to detect GC 2A gene expression of the recombinant adenovirus , with ELISA to analysis antibodies of vAd-GC2A stimulation in murine body , lactate dehydrogenase assay to detect the mouse cellular immunity effect , thereby the recombinant adenovirus immune effect was initially analysis.T-test method was used to do preliminary analysis and evaluation of the immune effect of recombinant adenovirus .Results:GC2A fragment amplified by PCR, the size was 1 961 bp and it was the expected.The results showed, the recombinant adenovirus vAd-GC2A proteins can be recognized by sera antibody of GM-CSF ( LMP2A); the results of ELISA showed that vAd-GC2A can stimulate the generation of antibodies.EBV-positive tumor cell killing rate (%) by CTL induced recombinant adenovirus was (66.7 ±6.9)%, at the same time the adenovirus type 5 wild strains and PBS were (24.3 ±2.5)%and(32.4 ±3.1)%only(P≤0.05).Conclusion:Fusion gene has been successfully constructed , vAd-GC2A in mice have humoral and cellular immune function.The data suggest that the vAd-GC2A should play a potent role in preventing the positive EB virus tumor.

Objective To identify the expression of a fusion gene GCA formed from GM-CSF gene and LMP2A gene of Epstein-Barr virus (EBV) in a recombinant BCG (rBCG) and to study its immunoge-nicity.Methods The rBCG was constructed to express the fusion gene GCA and the expressed products were detected by Western blot assay .ELISA was performed to measure specific antibody titers in serum sam-ples from mice immunized with rBCG .Lactate dehydrogenase assay was used to analyze the cellular immuni-ty of mice.A mouse model of EBV-positive gastric carcinoma was established to evaluate the therapeutic effects of rBCG.Results The target proteins of GM-CSF and LMP2A were successfully expressed in rBCG . The specific antibodies were detected in rBCG immunized mice as indicated by ELISA .The maximum anti-body titer reached 1 ∶27 900 [(326.5±7.8) pg/ml] as injection with rBCG 5×108/mouse.The rBCG in-duced cytotoxicity of cytotoxic lymphocytes (CTLs) to EBV-positive gastric carcinoma cells (GT39) (with a killing rate of 89.6%±6.8%) was significantly higher than that of control group (P<0.05) The sizes of tumor in PBS control group [(1964.0±548.7) mm3] and BCG group [(1268.65±72.4) mm3] were big-ger than those in rBCG group [(168.64±78.80) mm3].Conclusion The rBCG expressing GM-CSF and LMP2A fusion gene was successfully constructed .The rBCG could induce humoral and cellular immune re-sponses in mice and inhibit the growth of tumor .

Objective To establish the analysis criteria of nucleoplasmic bridges (NPB) and a dose-response curve of NPB frequencies in human peripheral blood lymphocytes irradiated by 60Co γ-rays.Methods Human peripheral blood samples were collected from three healthy males,and were irradiated with 0,1,2,3,4,5 and 6 Gy of 60Co γ-rays.A cytokinesis-block micronucleus assay was carried out to analyze NPBs and micronuclei (MN) in binucleated cells.Results NPBs in binucleated cells at each dose level of γ-ray was conformed to the Poisson distribution.The dose-response curve of the γ-ray-induced NPB frequencies in human peripheral lymphocytes followed the linear-quadratic model y =(1.39 × 10-3)x2 + (4.94 × 10-3)x (R2 =0.981,P ＜ 0.05).Conclusions The dose-response curve of NPB frequencies in human peripheral blood lymphocytes induced by 0-6 Gy 60Co γ-rays was established.

Objective: To develop an HPLC method for the determination of rutin, hyperin and quercetin in Vicia sepium L. . Methods:The samples were separated on an Agilent ZORBAX Eclipse XDB-C18 column(250 mm × 4. 6 mm, 5 μm) with the mobile phase consisting of acetonitrile-1‰ phosphoric acid solution with gradient elution at the flow rate of 0. 8 ml·min-1 . The column tem-perature was 30℃, and the detection wavelength was set at 370 nm. Results: The linear range of the three components was 4. 090-130.940 μg ·ml-1(r=0.999 9), 4.600-147.200 μg ·ml-1(r=0.999 9) and 0.810-25.780 μg·ml-1(r=0.999 8), and the average recovery was 103. 45% (RSD=1. 25%), 98. 96% (RSD=1. 77%) and 102. 88% (RSD=0. 84%)(n=6), respectively. Conclusion:The method is stable, reproducible and simple, which can be used in the quality control of Vicia sepium L. .

BACKGROUND:Recombinant adeno-associated virus serotype 9 has a high affinity in myocardial tissue, and the expression of recombinant adeno-associated virus serotype 9-enhanced green fluorescent protein (rAAV9-eGFP) in the aorta of atherosclerosis mice is not clear. OBJECTIVE:To explore the optimal time point of rAAV9-eGFP expression in the aorta of atherosclerosis mice. METHODS:Atherosclerosis model was established with high-fat diet in 30 ApoE-/-mice for 16 weeks. Among them, 25 mice were injected with 5.0×1011 vg (virus genomes) rAAV9-eGFP through the tail vein, while the remaining 5 mice were injected with saline, serving as the control group. The virus-transfected mice were kil ed at 14, 21, 28, 35 and 60 days after transfection, and aortic tissue was harvested. The expression of enhanced green fluorescent protein was detected with laser scanning confocal microscope. Western blot assays were used to detect the expression of enhanced green fluorescent protein in aorta. The expression of enhanced green fluorescent protein in vivo was observed and the optimal expression time point was determined. RESULTS AND CONCLUSION:rAAV9-eGFP effectively transfected the aorta of atherosclerosis mice, enhanced green fluorescent protein was expressed in aortic tissue, and the expression intensity increased gradual y with the increasing transfection time. The highest expression level was found at 35 days after transfection and then maintained stable at 60 days. There were significant differences at different time points after transfection (P<0.001). These data indicate that rAAV9-eGFP can be effectively expressed in the aorta of atherosclerosis ApoE-/-mice and rAAV9-eGFP can be regarded as the optimal vector in the treatment of atherosclerosis.

Objective To investigate the dose-effect relationship of mRNA level of sensitive mitochondrial genes in human lymphoblastoid cells induced by ionizing radiation.Methods Seven human sensitive genes,including ND3,Cyt b,COX Ⅰ,COX Ⅱ,COX Ⅲ,ATPase6 and ATPase8 were chosen.Changes of mRNA level of these genes were detected by RT-PCR and Real-Time PCR at 24 h after irradiation in human lymphoblastoid cells,which were exposed to 0 - 15 Gy of 60 Co γ-rays.Results The expression of these 7 genes at mRNA level was up-regulated 24 h after irradiation in human lymphoblastoid cells.The level of gene expression of COX Ⅰ,which belongs to complex Ⅳ of mitochondrial respiratory chain,was most obvious,and the peak occurred after irradiation of 8 Gy,which was 13 times of the control group.A good dose-effect relationship was showed for COX Ⅲ gene expression at dose range of 3 -10 Gy as well as 3 - 15 Gy for other 3 genes including ND3,ATPase6 and ATPase8.Conclusions Gene expression levels of COX Ⅲ,ND3,ATPase6 and ATPase8 24h post-irradiation at certain irradiation dose range could be used for radiation damage biomarkers.

Objective To evaluate the accuracy of 99mTcO4- gastrointestinal imaging in the diagnosis of Meckel’s diverticulum in children. Methods The clinical data of 99mTcO4-imaging, surgery and pathological results of 95 children with gastrointestinal bleeding were retrospectively analyzed. Results Forty-four cases of 95 patients had positive ifnding of 99mTcO4-ectopic gastric mucous membrane imaging, and positive rate was 46.3%. In the positve cases 52.3%cases (23/44) were diagnosed of intestinal heterotopic gastric mucosa, 47.7%cases (21/44) were suspected of intestinal heterotopic gastric mucosa. The sex ratio (males to females) was 7.8:1. Thirty-seven positive cases underwent operation;among them, 35 cases were found to have Meckel’s diverticulum and 1 case had duplication of the digestive tract;Diverticulum lesion was not found in 1 case. The samples of 35 cases diagnosed surgically of Meckel’s diverticulum were examined by pathologic histology. All 35 cases were conifrmed as Meckel’s diverticulum and ectopic gastric mucosa epithelium, including 2 cases with ulcer formation, 1 case with small patches of pancreatic tissue, 1 case with ileocecal junction of suppurative enteritis and 1 case with chronic appendicitis. Conclusions 99mTcO4-radionuclide imaging technique is of high value for the non-invasive diagnosis of children’s Meckel’s diverticulum.

Objective To investigate the dose response of S100A4 gene expression in the irradiated lymphoblastoid cells AHH-1 at different time points post irradiation.Methods AHH-1 cells was exposed to different doses(0,1,3,5,8,10,15 and 18 Gy)of 60Co γ-rays,and its mRNA levels of S100A4 was detected by reverse transcription PCR and real-time PCR at 4,8,12,24,48 and 72 h after irradiation.Results Within the range of applied doses,the level of S100A4 gene expression was upregulated with a good dose-response (R2 =0.79-0.93,P ＜ 0.05) and had obvious difference at different time points (F =8.91,P ＜ 0.01).Conclusion S100A4 gene expression at transcriptional level could be detected easily and had optimum dose-responses at certain time points after irradiation,and hence is applicable as a dosimeter.