Grass carp reovirus (GCRV), a disaster agent to aquatic animals, belongs to Genus Aquareovirus of family Reoviridea. Sequence analysis revealed GCRV genome segment 8 (s8) was 1296 bp nucleotides in length encoding an inner capsid protein VP6 of about 43kDa. To obtain in vitro non-fusion expression of a GCRV VP6 protein containing a molecular of fluorescence reporter, the recombinant baculovirus, which contained the GCRVs8 and eGFP (enhanced green fluorescence protein)genes, was constructed by using the Bac-to-Bac insect expression system. In this study, the whole GCRVs8 and eGFP genes, amplified by PCR, were constructed into a pFastBacDual vector under polyhedron (PH) and p10 promoters, respectively. The constructed dual recombinant plasmid (pFbDGCRVs8/eGFP) was transformed into DH10Bac cells to obtain recombinant Bacmid (AcGCRVs8/eGFP) by transposition. Finally, the recombinant bacluovirus (vAcGCRVs8/eGFP) was obtained from transfected Sf9 insect cells. The green fluorescence that was expressed by transfected Sf9 cells was initially observed 3 days post transfection, and gradually enhanced and extended around 5days culture in P1(Passage1) stock. The stable high level expression of recombinant protein was observed in P2 and subsequent passage budding virus (BV) stock. Additionally, PCR amplification from P1 and amplified P2 BV stock further confirmed the validity of the dual-recombinant baculovirus. Our results provide a foundation for expression and assembly of the GCRV structural protein in vitro.

Objective To investigate the vaginal flora characteristics,clinical symptoms and signs of lactating women 6-8 weeks after delivery and the influence factors.Methods Two hundred and twenty-three puerpera 6-8 weeks after delivery who went to outpatient department of Peking University First Hospital for postpartum review (study group) and 200 normal non-pregnant women (control group) from April to May,2010 were enrolled into this study.All in study group were evaluated by a questionnaire regarding their symptoms,sexual behavior,breast-feeding situation and delivery mode.Vaginal secretions from upper 1/3 of vagina were collected and evaluated by vaginal microbial evaluation system.Bacterial vaginosis (BV) was diagnosed if Nugent score ≥ 7.Enumeration data between groups was compared by t test,and quantitative data by Chi-square test.Results The density of vaginal flora in study group was different from that of control group (x2 =96.302,P＜0.001) ; so did the flora diversity of two groups (x2 =69.871,P＜0.001).There was no difference between mean age of study group [(30.5 ± 3.9) years] and control group [(30.9 ± 2.4)years],P＞0.05.The cesarean section rate was 52.7％ (89/169),13 women (7.7％) had began their sexual behavior before interview,116 women (70.3％) took breast-feeding.Dominant bacteria in the vagina of lactating women were Gram-positive cocci (69/175,39.4％) and Gram-positive bacillus (54/175, 30.9％), which was different from control group (Gram-positive bacillus,180/189,95.2％),P＜0.001.The vaginal pH of study group was 5.0±0.5,which was higher than that (＜4.5) of control group.In study group,incidence of lower hydrogen peroxide level of vagina was 84.3 ％ (177/210) ; positive rate of sialidase was 6.7 ％ (14/210) ; positive rate of leukocyte esterase was 91.0％ (191/210).A large number of postpartum women had an abnormal Nugent score,most of them showed a state of intermediate BV (132/195,67.7％) or BV (8/195,4.1％),while a few patients had symptoms of BV (23.5％,33/140).Sexual behavior had no significant effect on postpartum vaginal flora.The proportion of abnormal vaginal flora in breast feeding mothers (74.1 ％,86/116) was significantly higher than that (57.1％,28/49) in artificial feeding mothers,P=0.034.Delivery mode also affected postpartum vaginal flora.Women underwent cesarean section were more susceptible to abnormal vaginal flora (77.5％,69/89) than those underwent vaginal delivery (57.5％,46/80),x2 =7.726,P=0.005.Conclusions Vaginal flora of postpartum women 6-8 weeks after delivery had significant difference from that of nonpregnant women.The incidence of asymptomatic abnormal vaginal flora was high.Breastfeeding and delivery mode are influence factors of abnormal vaginal flora.

Antineoplastic Agents ; pharmacology ; Cell Cycle ; Cell Line, Tumor ; drug effects ; Cell Proliferation ; Granulocyte Colony-Stimulating Factor ; pharmacology ; Humans ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; metabolism ; pathology

Objective To explore the clinical effects of atomization inhalation pulmicort respules in treatment with pediatric acute infectious laryngitis. Methods 104 cases with pediatric acute infectious laryngitis admitted in Xiaoshan Hospital of Zhejiang Province from January 2011 to July 2013 were radomly divided into experimental group(n=52)and control group(n=52). Patients in experimental group were given atomization inhalation pulmicort respules,and patients in control group were received dexamethasone by intravenous injection. The symptom improvement,disappearing times of symptoms and therapeutic effect in two groups were observed and compared. Results Compared with control group,the improvement scores were higher,and disappearing time of symptoms were shorter in experiment group(P<0.05). The therapeutic effect in experimental group was up to 100%, while control group was 90.74%,and the difference between them was statistically significant(P<0.05). Conclusion Compared with dexamethasone, atomization inhalation pulmicort respules has advantages in treatment with pediatric acute infectious laryngitis,with better efficacy,shorter treatment time and fewer side effects.

Objective To investigate the suppression effect of RNA interference(RNAi) on the expression of epidermal growth factor receptor(EGFR) of ovarian cancer SKOV3 cell line. Methods SKOV3 cells were transfected with synthetic EGFR sequence-specific dsRNA by lipofectamin 2000.The expression level of EGFR mRNA and protein were detected by RT-PCR and Western blotting,respectively.The proliferation of transfected SKOV3 cells was detected by WST-1 cell proliferation assay.The abilities of adhesion,migration and invasion were detected by cell adhesion assay,cell migration assay and Matrigel invasion assay,respectively. Results The suppression rates on EGFR mRNA and protein by sequence-specific dsRNA-EGFR were 73.65% and 58.94%,respectively.The cell proliferation was inhibited by 22.54%,35.76% and 31.07%,respectively at 24,48 and 72 h after transfection.The adhesion of sequence-specific dsRNA transfected SKOV3 cells for 30 min and 90 min were reduced by 12.11% and 18.66%,respectively.The migration and invasion of SKOV3 cells were decreased by 25.47% and 22.08%,respectively. Conclusion The down-regulation of EGFR expression of ovarian cancer by sequence-specific dsRNA can lead to the suppression of proliferation,adhesion,migration and invasion of ovarian cancer cells.

Objective To explore the feasibility and value of MSCT for the evaluation of hypophysoma before and after endonasal transphenoidal microsurgery.Methods 82 patients with hypophysoma intended to be treated by endonasal transphenoidal microsur-gery,were examined by 64-slice head CT angiography (CTA).All the images were transmitted to workstation and reconstructed by MIP、VR and MPR.The hypophysoma were typed according to the relationship between hypophysoma and its surrounding vessels. The shortest and longest diameter between internal carotid arteries of two sides,tumor length and width,the diameter between ex-ternal naris and the rear wall of sphenoid top and the length from external naris to the rear wall of tumor top were measured in VR images.Sphenoid division was observed in coronal plane image.Major vascular injury,bleeding,and residual tumor situation were observed on postoperative CT review.The surgery effect was evaluated by three associate professorwith SPSS 13.0.Results 39 ca-ses were type-A,30 were type-B,13 were type-C,tumor length ranged from 1.7 cm to 3.9 cm,width ranged from 1.3 cm to 2.5 cm,the diameter between external naris and the rear wall of sphenoid top and the length between external naris and the rear wall of tumor top were 6.9 cm~7.6 cm and 9 cm~12.2 cm.There are 1 5 cases of sphenoid without division,34 cases with one divi-sion,26 cases with two division and at least three division in 7 cases.The surgery effect was better in those which tumor size was relatively smaller compared to the diameter of the internal carotid artery.The evaluation of endoscopic endonasal transsphenoidal surgery with three types of pituitary adenomas had a significant difference (P <0.05).Conclusion MSCT can preferably reflect the spatial location relationship between pituitary adenimas and its surrounding structure,and it is effective evaluation methods for pre-operative and postoperative evaluation of transnasal microsurgical resection.

Objective To explore the application value of multi-slice CT angiography (MSCTA)in posterior approach of upper cervical vertebrae.Methods Fifty-one patients with suspected or conformed upper cervical disease were collected and performed cer-vical CTA examination.Twenty three patients underwent internal fixation surgery of posterior upper cervical and reviewed with CTA examination.Volume Render (VR),Maximum Intensity Projection (MIP)and Multi-planar Reformation (MPR)were used to show the courser and anatomical variation of vertebra artery,the shape of vertebrae,the relationship between the vertebra artery and adjacent structure,advantage artery.The distribution of screw placement,including perforation of the anterior wall,the inner wall, the outer wall,the upper wall,the interior wall and the inner wall of occipital were observed by VR,MIP and MPR after surgery, especially the relationship between the screw and vertebra artery,the screw and spinal cord.Comparing the correlation between the screw placement and postoperative symptoms.SPSS1 9.0 statistical packages were used for statistical analysis.Results VR,MIP and MPR can show the courser and nearby space relationship of cervical artery.The number of advantage artery was 25,equilibra-tion was 26,dysplasia variation was 13,course variation was 4.The number of tiny pedicle was 1 6,including 9 cases of ipsilateral advantage artery.The postoperative data analysis of 23 patients showed that:C1 -20,C2 -36,C3 - 1 5,C4 -2,occipital 40.The number of accurate screw placement was 83.Different type of perforation was 30,including 3 screws closing to vertebra artery,1 screw closing to spinal cord.All 23 patients were asymptomatic after surgery.Conclusion MSCTA can reflect the morphology of cervical vertebral,the spatial relationship between cervical vertebral and surrounding structure,which can provide an accurate refer-ence for clinical screw placement and improve the accuracy .The postoperative CTA can show screw placement accurately,which has great reference for the evaluation of clinical success rate of surgery and post-surgery treatment.

Objective Chiral recognition for the glimepiride and glimepiride-cis-isomer by applying three amino acid (D-Lysine,L-Glutamine and L-Tyrosine) as chiral probes based on electrospray ionization mass spectrometry (ESI-MS) was achieved.Methods glimepiride/glimepiride-cis-isomer solutions were mixed with three amino acid solutions.The complex was extracted by ESI-MS and then the fragmentation abundance was investigated applying collision induced dissociation (CID) by MS/MS,which is the basis of chiral recognition for glimepiride and glimepiride-cis-isomer.Results Chiral recognition effect was achieved with the recognition rate (R) 1.61,2.92 and 2.17 for D-Lysine,L-Glutamine and L-Tyrosine respectively.Conclusion 3 kinds ofchiral amino acids were used as probes to distinguish between stereoisomers,and rapid identification of glimepiride and glimepiride cis isomer by mass spectrometry come true for the first time.

Whether InfusaSleeve(IS) catheter can deliver antisense oligodeoxynucleotide (ODNs) following arterial denuation is unknown. We evaluate the feasibility of local endoluminal delivery of C-myc ODNs to the site of arterial denudation by using IS catheter and to determine the biological importance of these effects. IS catheter was introduced into right side of iliac artery of 21 rabbits after angioplasty of iliac artery. Animals were randomized to the control group (n＝6) receiving saline injection and the treated group receiving c-myc antisense (n＝15, 1 mg ODNs per vessel). In two weeks and 40 days following the operation, angiography was performed. Morphometric analyses were carried out in balloon-denuded iliac arteries. The expression of c-myc protein was detected by using a mouse monoclonal antibody to c-myc. Morphometric analyses carried out at 40 days after transcatheter c-myc antisense oligomer administration. The results showed that maximal neointimal area was reduced from 7.66±3.7(×105 μm2) in the control group (n＝6) to 4.04±1.02(×105 μm2) in the antisense treated group (n＝6, P＜0.05). These changes in vascular remodeling following denuding injury resulted in an increase in residual luman from 20～50% in the control group to 70～90% in the antisense-treated group. C-myc protein expression was virtually undetectable at baseline in locally ODNs-delivered arteries and detectable in control denuded arteries. The results show that: ①Single IS transcatheter administration allowed endoluminal delivery of ODNs to the site of arterial injury; ② c-myc antisense oligomer reduced the formation of neointime in denuded arteries, implying a therapeutic potential of this approach.

OBJECTIVEThis research explores the regulatory role of bone morphogenetic protein 2 (BMP2) in the expression of sclerostin in OCCM-30 cementoblast.

METHODSOCCM-30 cementoblasts were treated with 50 and 100 ng · mL⁻¹ BMP2 for 3, 5, and 7 days. SOST mRNA was detected by real-time quantitative polymerase chain reaction (RT-PCR). Western blot analysis was employed to detect the sclerostin levels in the nucleus. Five groups were prepared for the experiments: control, BMP2, BMP2+dorsomorphin, BMP2+SB202190, and BMP2+PD98059. OCCM-30 was pretreated with BMP2 for 3 and 5 days, and then the sclerostin and SOST mRNA levels were measured.

RESULTSRT-PCR and Western blot analyses showed that BMP2 upregulated the expression of SOST in a concentration-dependent manner. SOST expression increased with time (P < 0.05). Moreover, sclerostin levels of BMP2+dorsomorphin, BMP2+SB202190, and BMP2+PD98059 groups were lower than that of the BMP2 group, and the sclerostin level in BMP2+dorsomorphin group was lowest (P < 0.05).

CONCLUSIONThe upregulation of SOST by BMP2 in OCCM-30 is mainly mediated by the BMP2/Smad signal pathway.

Blotting, Western ; Bone Morphogenetic Protein 2 ; metabolism ; Bone Morphogenetic Proteins ; metabolism ; Dental Cementum ; metabolism ; Genetic Markers ; Signal Transduction ; Up-Regulation