In order to find the characteristics of two members of gene family of squaleneexpoxidase (SE) , a quantitative real time PCR method was developed to analyze the expression of Eleutherococcus senticosus SE1 and SE2 gene from different growth periods and in different organs. The result indicated that all the expression of SE2 more than SE1 in the whole growth period and organs of E. senticosus. And in the whole growth period, expression of SE1 showed a low-high-low characteristic. Both expression of SE2 and growth period showed the same trend. The lowest content of the expression was in the roots. SE1 expression have been improved more than SE2 when treated with MeJA. The expression of E. senticosus SE1 and saponins content had significantly positive correlation (P < 0.05) and the correlation coefficients was 0. 858, while the correlation was not significant for SE2. That indicated that SE1 played a key enzyme gene in the biosynthesis of triterpenoidsaponins
Eleutherococcus ; chemistry ; enzymology ; genetics ; growth & development ; Gene Expression Regulation, Plant ; Peroxidase ; genetics ; metabolism ; Plant Proteins ; genetics ; metabolism ; Saponins ; analysis ; metabolism ; Transcriptome

According to the sequence of P450 cDNA of Eleutherococcus senticosus, specific primers were designed. Frokaryotic ex pression vector pET30a-P450 was constructed and the prokaryotic expression conditions were optimized. Results showed that the BL21 after being transformed with the recombinant expression vector accumulated the high amount of recombinant protein. SDS-PAGE analysis showed that the recombinant protein was about 53 kDa. The recombinant accumulated the highest amount of recombinant protein af ter IPTG (1 mmol x L(-1)) at 27-37 degrees C for 24 h. Consequently P450 gene of E. senticosus could be expressed successfully by prokaryotic expression vector pET30a-P450. Induction temperature, IPTG concentration, medium type and amount of induction time could all influence the expression of target protein, but the impact strength was different.
Cloning, Molecular ; Cytochrome P-450 Enzyme System ; genetics ; metabolism ; Eleutherococcus ; enzymology ; genetics ; Escherichia coli ; genetics ; metabolism ; Gene Expression ; Genetic Vectors ; genetics ; metabolism ; Plant Proteins ; genetics ; metabolism ; Plasmids ; genetics ; metabolism ; Recombinant Proteins ; genetics ; metabolism

Adenocarcinoma, Papillary ; blood supply ; metabolism ; pathology ; Adult ; Aged ; Carcinoma, Pancreatic Ductal ; blood supply ; metabolism ; pathology ; Chemokine CXCL12 ; metabolism ; Female ; Gene Expression Regulation, Neoplastic ; Humans ; Lymph Nodes ; metabolism ; Lymphatic Metastasis ; Male ; Microvessels ; pathology ; Middle Aged ; Neoplasm Staging ; Neovascularization, Pathologic ; metabolism ; pathology ; Pancreas ; metabolism ; Pancreatic Neoplasms ; blood supply ; metabolism ; pathology ; Receptors, CXCR4 ; metabolism

BACKGROUND: The particular bystander effect of suicide gene can remarkably kill tumor cells. Meanwhile, it can be used together with radiotherapy as well as immune gene therapy, and overcome the defect of low gene transduction efficiency.Cytosine deaminase (CD) can generate a powerful bystander effect.OBJECTIVE: To observe the effect of a eukaryotic expression plasmid plRES2-AcGFP1-CD mediated by liposome transfected into bone marrow mesenchymal stem cells (BMSCs) and its gene expression.DESIGN, TIME AND SETTING: A cytologic experiment at genetic level was performed at Research and Development Center of Stem Cell and Tissue Engineering, Dalian University of Technology from May to December 2007.MATERIALS: A total of 6 C57BL mice of SPF degree and weighing 18-20 g were used for separation and culture of BMSCs.Escherichia coli DH5a was provided by Research and Development Center of Stem Cell and Tissue Engineering, Dalian University of Technology. Lipofectamine~(TM) 2000 was provided by Invitrogen, China.METHODS: The DNA plasmids were extracted from transformed Escherichia coli DH5a. plRES2-AcGFP1-CD plasmid was identified by BamHI/Xhol double digestion. The BMSCs derived from mouse femur and tibia were cultured and purified by adhesion method in vitro. Signal cell suspension prepared by BMSCs of the third generation was cultured by adding fluorescence-labeled CD44, CD45, CD90 and CD105 antibodies. BMSCs of the third generation were transfected by LipofectamineTM 2000 mediation.MAIN OUTCOME MEASURES: Identification of recombinant plasmids; the expressions of surface markers on BMSCs were detected by flow cytometry; the expressions of cytosine deaminase gene were observed at 36 and 48 hours after transfection under fluorescent inverted microscope.RESULTS: After agarose gel electrophoresis, a band appeared at 1.0-1.5 kb of the digested products of plRES2-AcGFP1-CD plasmids, and the band was accorded with the length of cytosine deaminase gene in the length. Flow cytometry demonstrated that the cells were negative for CD45 but positive for CD44, CD90 and CD105. Fluorescent inverted phase contrast microscope suggested that at 36 hours after plRES2-AcGFP1-CD gene transfection an expression of green fluorescent protein was found in the BMSCs; additionally, at 48 hours after transfection, the expression of green fluorescent protein remained and the intensity was remarkably increased.CONCLUSION: The liposome-mediated plRES2-AcGFP1-CD gene successfully expressed in BMSCs, and the expression peaked at 48 hours after transfection.

Porcine circovirus type 2 (PCV2) can cause immunosuppression on herds. PCV2, as an essential pathogen of PCV2-systemic disease (PCV2-SD), has caused considerable economic losses in pig industry worldwide. Here we review and address the evolution, viral protein and immunolesion of PCV2 and preventive techniques of PCV2-SD.
Animals ; Circoviridae Infections ; veterinary ; Circovirus ; genetics ; Phylogeny ; Swine ; Swine Diseases ; virology ; Viral Proteins ; genetics

Background Fibrin glue has been utilized to adhere the graft during the pterygium excision with conjunctival autografts.Several relevant clinical randomized controlled trial(RCT) and retrospective studies have been published abroad,but the samples for its effectiveness and safety issue of fibrin glue and suture are still underinvestigation.Objective Current study was to quantificationally assess the efficacy and safety of fibrin glue versus sutures in the application of pterygium excision with conjunctival autografts.Methods Based on established search strategy,a computerized literature search was conducted to identify all citations concerning the RCT for effectiveness and safety evaluation of fibrin glue and suture for the graft fixation during the pterygium excision with conjunctival autografts from MEDLINE ( from 2000 to October,2010 ),EMbase ( from 2000 to October,2010 ),Cochrane Central Register of Controlled Trials ( Issue 4,2010 ),CBMdisc ( 2000 to October 2010 ),CNKI ( 2000 to October 2010 ),and the relevant conference proceedings and references searched by hand was performed as supplement.The included literature was scored with Jadad table.The Cochrane Collaboration' s RevMan 5.0 software was used for the test of heterogeneity or test for overall effect.The effective indexes,such as operative duration,recurrence rate and complication,were evaluated by Meta analysis.Results Six RCTs involving 401 eyes of 377 participants were identified.These literatures were published with English in 2004-2010 from China,New Zealand,Sweden,Israel,Turkey and Malasia and the Jadad scores were 4-5.The quantitatively analysis revealed that fibrin glue appeared to short the operative time compared with suturing method (MD =14.23 ;95％ CI:- 16.18- 12.29;P=0.00) and drop the rate of recurrence ( RR =0.49,95％ CI:0.26 -0.95 ; P =0.03 ).No significant differences were found in the rate of postoperative graft dehiscence or absence (RR =3.41,95 ％ CI:0.85-13.68;P =0.08 ).Conclusions Fibrin glue shows the good effectiveness and easy application during the pterygium excision with conjunctival autografts.Long-term follow-up of multi-central RCTs with a larger number of cases are still needed to support this conclusion.

Objective To analyze the characteristic and the clinical application of high -frequency stimulation ABR for the patients with unilateral sudden deafness .Methods 40 patients aged 17~45 with unilateral sudden deaf-ness(40 health ears ,subject group)and 20 normal volunteers (40 ears ,control group)were selected to receive high and low frequency stimulation ABRs and pure tone audiometry .Results There were no statistical differences be-tween subject group and control group in pure tone audiometry .The wave Ⅰlatency of high -frequency ABR was longer in the subject group than that in the control group(P<0 .001) .The abnormal rate of Ⅰ - Ⅴ interval between high and low frequency stimulation ABRs was 62 .5% (25/40) in the subject group ,and 5% (2/40) in the control group .There were statistical differences between the two groups (χ2 =29 .574 ,P<0 .001) .Conclusion The waveⅠlatency of high-frequency ABR was longer and the abnormal rate of Ⅰ - Ⅴ interval markedly increased in the health ear of patients with unilateral sudden deafness .This may suggest that the contralateral ear may have an ab-normal blood supply of the inner ear when single side sudden deafness happened .High frequency stimulation ABR can be used to detect the potential changes of contralateral ear before the hearing loss occurred .

Objective To investigate the changes of serum bilirubin level in elderly patients with acute cerebral infarction and its significance.Methods 164 hospitalized elderly patients,who suffered from acute cerebral infarction within 1 week after onset,were divided into 2 groups according to age:group A aged over 60 years(n=85) and group B aged 40-60 years(n=79),and 66 healthy subjects aged over 40 years were collected as controls(group C).Serum bilirubin levels in all subjects were determined.The ratio of pulse pressure over mean arterial pressure(PP/MAP) in group A and B was calculated.Nerve function scores in the three groups were detected before and after 2 weeks of treatment.Meanwhile,the data of risk factors including blood glucose,blood pressure,blood lipids,smoking and drinking in group A and B were collected.Results Compared with group C,serum total,direct,indirect bilirubin levels were increased in group A and B(both P＜0.01),and the change was smaller in group A than in group B(P＜0.05).The nerve function scores was lower in group A than in group B before and after treatment [(35.2±12.6) vs.(44.3±7.9),(40.7±9.1) vs.(51.3± 4.1),t=5.58,9.73,both P＜0.01],but PP/MAP and the numbers of risk factors were higher in group A than in group B [(0.46±0.06) vs.(0.38±0.06),93.01 vs.71.20,both P＜0.01].There were no significant correlations of serum total,direct and indirect bilirubin levels with nerve function scores in group A or B(all P＞0.05).Conclusions Serum bilirubin level is increased in patients with acute cerebral infarction,but the endogenous antioxidant capacity is decreased because of aging,multiple risk factors and more serious atherosclerosis in elderly patients,and the increment of bilirubin level is relatively smaller in acute cerebral ischemia,leading to the reduced protective effect against stress.Serum bilirubin level may influence the prognosis in patients with acute cerebral infarction.

Objective: To clone Fd and light chain genes of monoclonal antibody HAb18 against human hepatoma and verify their accuracy and liability. Methods: Total RNA was extracted from hybridoma cell line secreting MAb HAb18, and Fd and light chain genes were amplified by RT-PCR. After PCR products were ligated into pMD18T vector, positive clones were screened and DNA sequences were tested and analysed by relative softwares. Then, light chain and Fd genes were sequential cloned into phage display vector pComb3. After recombinant vector was transformed into E.coli XL1-blue, recombinant vector was rescued by helper phage M13K07 and the specificity of phages to antigen was detected by indirect ELISA. Results: The size of amplified Fd and light chain genes was separately 665 bp and 668 bp. The results of sequence analysis showed that both VL and VH contained 2 characteristic cystines and CH1 was IgG1 classes and CL was ?. ELISA result identified that expressed Fab antibody could specially bind to corresponding antigen. Conclusion: Fd and light chain genes of MAb HAb18 were successfully cloned, which lay a good foundation for constructing a diversity of engineering antibody.

Objective: Study on the growth character of B16 melanoma cell which can express angiostatin. Methods: Angiostatin gene was constructed from human plasminogen cDNA by deletion mutation. A B16 melanoma cell clone named BAG28 which stably expresses angiostatin was established by introducing gene into it. Results: BAG28 in vitro had no changes in proliferation rate and the ability of clone formation in soft agar. Study in vitro showed that the tumor weight had reduced about 87% ( P