Two fragments G1 and G2 of the glycoprotein G gene of bovine respiratory syncytial virus(BRSV) were selected for expression in Escherichia coli based on the analysis of glycoprotein G by DNA Star software.Then the two fragments of glycoprotein G were amplified by PCR with synthesized G gene of BRSV as the template.The amplified fragments G1 and G2 are 570bp and 308bp in length,respectively.The PCR products were cloned into pET30a vector and expressed in soluble form in E.coli after induction of cultured E.coli with IPTG.Both of the recombinant proteins G1 and G2 were purified by immobilized Ni ion affinity chromatography under native conditions.Then the purified proteins were analysed by Western blotting.The results showed that the purified recombinant protein G1 retained good antigenicity and specificity.But the purified recombinant protein G2 didn't possess biological activity.Antibodies against BRSV were detected in suspected bovine serum samples in China by using indirect ELISA and Western blotting with the purified recombinant protein G1.The purified recombinant protein G1 might be used as antigen for establishing serological methods for diagnosis of BRSV infection.And the purified recombinant protein G1 might also be used for preparing polyclonal and monoclonal antibodies for research on biological functions of glycoprotein G of BRSV.

Objective To investigate the clinical states of enuresis children companied with spina bifida occulta(SBO)and study the efficient way of treatment.Methods The children with SBO were check out by X ray from a total of 121 children with bedwetting.Their parents were asked to complete the enuresis questionnaires.Urine routine test and B-ultrasound examination about kidney,bladder and ureter were also asked to be done.The clinic data of the 49 children were attained and analyzed.They were randomly divided into 2 groups,and given the controlled treatment.Group A[used 1-deamino-8-D-arginine vas-opressin(DDAVP)only] and group B(used DDAVP plus oxybutynin plus bladder training)treated for 12 weeks.Results There were totally 49 bedwetting children companied with SBO,and most of them(44 cases,89.8%)were severe type(bedwetting times≥7 times/week).Some of them coexisted with frequency,urgency,gentle urgency incontinence and microscopic hematuria(22 cases).Thirty cases were found the functional bladder capacity(FBC)decrease by B-ultrasound.The cure rates were 58.3%(group A)and 88.0%(group B)respectively.The relapse rates were 36.8%(group A)and 12.5%(group B)respectively after stopping treatment for 3 months.Conclusions SBO accounts for considerably higher rate in enuretic children.It might cause the disability of bladder function.The treatment plan with DDAVP plus oxybutynin plus bladder training can not only increase the cure rate but also lower the relapse rate.

Objective To analyze the relationship between different rectal volume,bladder volume and dose of organs at risk (OARs) in intracavitary brachytherapy for cervical cancer.Methods A total of 47 patients with cervical cancer were selected.All of them were treated with high dose rate (HDR) intracavitary brachytherapy with a 600 cGy dose for the dosage point.The effects of different volume of rectum bladder and small intestine for corresponding exposure dose under the standard planning were evaluated using a dose-volume histogram (DVH).According to bladder volume,patients were divided into three groups,＜ 80 cm3 group,80-120 cm3 group and ＞ 120 cm3 group.And according to rectum volume,patients were divided into ＞ 60 cm3 group and ≤ 60 cm3 group.The relationship between the volume and dosage were analyzed.The ANOVA test and t test were used for analyzing D1 cm3,D2 cm3,D30％ and D50％.Results Compared with the group with ＜ 80 cm3 bladder volume,D30％,D50％ value of bladder in groups with 80-120 cm3 and ＞ 120 cm3 of bladder volume increased (F =5.074,5.088,P ＜ 0.05).The difference of D1 cm3 and D2 cm3 value of the small intestine between 80-120 cm3 and ＞ 120 cm3 bladder volume groups were not statistically significant (P ＞ 0.05).D1 cm3 of rectum in groups with ≤ 60 cm3 rectum volume was decreased than that of ＞ 60 cm3 group (t =-2.045,P ＜ 0.05).Conclusions Keeping an appropriatly full bladder and reducing rectal volume in cervical cancers treated with intracavitary brachytherapy can make the exposure dose of bladder,rectum and small intestine relatively small,and reduce the adverse reactions of radiotherapy.

BACKGROUND & OBJECTIVE; survivin, a member of inhibitor of apoptosis protein (IAP) gene family, expresses in various human cancer tissues, and may facilitate tumor cell evasion from apoptosis, and promote aberrant mitotic progression. This study was to investigate cell proliferation and apoptosis status of human breast carcinoma cell line MCF-7 after knockdown of survivin. METHODS; Small interfering RNA was transfected into MCF-7 cells to inhibit expression of survivin. mRNA and protein levels of survivin were detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. Proliferation inhibition rate of MCF-7 cells was analyzed by MTT assay. Cell apoptosis was assayed by flow cytometry (FCM). RESULTS; The expression of survivin in siRNA-transfected group decreased by 64% in comparison to untransfected group. After treatment of different concentrations of siRNA, proliferation inhibition rate and apoptosis rate of MCF-7 cells were increased. The highest proliferation inhibition rate was 60.9%, and the highest apoptosis rate was 29.0% after treatment of 200 nmol/L of siRNA. CONCLUSION; survivin siRNA might be a useful therapeutic agent for the treatment of breast carcinoma.

OBJECTIVETo investigate the inhibitory effects of bcl-xl antisense oligodeoxynucleotides (ASODN) on growth and gene expression of human nasopharyngeal carcinoma (NPC) in nude mice.

METHODSCNE-2Z cell line was implanted subcutaneously into nude mice. Twenty four h after implantation, bcl-xl ASODN and mismatch control oligonucleotides (SCODN) were injected subcutaneously into nude mice, respectively. The tumor volume and weight were measured twice weekly. The histopathological changes of the tumors were observed by HE staining. RT-PCR and Western-blot were performed for bcl-xl gene expression.

RESULTSGrowth of NPC was significantly inhibited in the ASODN therapy group as compared with that in the control group (P < 0.01). The growth inhibitory rate was 41.7% in the ASODN group and 19.0% in the SCODN group. The expression level of bcl-xl mRNA and protein was decreased in the ASODN group, whereas in the SCODN group there was no significant difference in contrast with saline-treated control group.

CONCLUSIONOur findings suggest that bcl-xl antisense oligodeoxynucleotides results in marked inhibition of NPC growth in nude mice. It may be a novel treatment approach for human nasopharyngeal carcinoma.

Animals ; Female ; Humans ; Male ; Mice ; Mice, Inbred BALB C ; Nasopharyngeal Neoplasms ; drug therapy ; pathology ; Neoplasm Transplantation ; Oligonucleotides, Antisense ; therapeutic use ; Proto-Oncogene Proteins c-bcl-2 ; antagonists & inhibitors ; genetics ; RNA, Messenger ; analysis ; Transplantation, Heterologous ; bcl-X Protein

Objective To explore the self-management action of patient suffered from mechanical heart valve post-operation. Methods 168cases of the patients suffered from the mechanical heart valve post-operation were separately divided into 86 cases as the interference group and 82 cases as control group at random. The interference group was given for the normal medical treatment and also objective self-management education but the control group used normal medical treatment and also health education. Two groups of the patients were tested by "Self-Management Action Measurement Form" separately to asseSS the self-management action and also were assessed by Chinese version of SF-36 Measurement Form for the assessment of two groups of the patients for the comparison of the patients of two groups in living quality(QOL) one year after treatment and nursing. Results The various items of assessments of interference group were higher than the comparison group in self-management action measurement form SF-36 Measurement Form with the difference of statistics meaning(P ＜0. 01) averagely for the comparison of two groups one year after the treatment and nursing. Conclusions The self-management education can help the patients to set up a good action mode and improve the patients' self-management disease ability, reduce the occurrence of the complication to improve health and reach to raise living quality of the patients suffered from mechanical heart valve post-operation for purpose.

BACKGROUND & OBJECTIVE: Recent studies showed overexpression of bcl-xL in human nasopharyngeal carcinoma (NPC) cell line CNE-2Z; it may play a pivotal role in tumorigenesis, metastasis, and drug resistance of NPC. This study was to explore inducing effect of bcl-xL short hairpin RNA (shRNA) on apoptosis of CNE-2Z cells. METHODS: After transfection of recombinant plasmid pmU6-RNAi expressing bcl-xL shRNA,apoptotic CNE-2Z cells were detected by fluorescent staining and flow cytometry (FCM). mRNA levels of bcl-xL, bcl-2, survivin, and caspase-3 was detected by reverse transcription-polymerase chain reaction (RT-PCR); while protein levels of Bcl-xL, Caspase-3, and P53 were detected by Western blot.RESULTS: When treated with pmU6-RNAi for 24 h, an obvious apoptotic peak of CNE-2Z cells appeared; cell shrinkage, chromatin condensation, and nuclear fragmentation were observed in most cells under fluorescent microscope. RT-PCR analysis showed that pmU6-RNAi down-regulated mRNA levels of bcl-xL, bcl-2, and caspase-3, but had little or no effect on mRNA level of survivin; Western blot analysis showed an obvious reduction in protein levels of Bcl-xL and Caspase-3, and a great increase in protein level of P53.CONCLUSIONS: bcl-xL shRNA can induce apoptosis of CNE-2Z cells, which may be closely related to down-regulation of bcl-2, caspase-3 and p53. bclxL shRNA may be helpful for developing gene therapy for NPC.

Objective:To investigate the effect of inhibiting of HOXB7 gene expression on proliferation and apoptosis of colon cancer cells.Methods:The synthetic negative control siRNA (negative control group) and HOXB7-siRNA (HOXB7 transfection group) were transfected into human colon cancer SW480 cells by LipofectamineTM2000 liposome mediated method,untreated cells as blank group.RT-PCR and Western blot were used to detect the mRNA and protein expression of HOXB7 after transfected 48 h respectively;cell proliferation was detected by CCK8 assay after transfected 24,48,72,96 h;cells apoptosis was detected by flow cytometry after transfected 48 h;the expression of apoptosis related proteins Bcl-2,Bax and Notch1 signaling pathway Notch1 and Hes1 were detected by Western blot.Results:The mRNA and protein expression of HOXB7 in HOXB7 transfected group was significantly lower than that in blank group(P<0.05);OD value was no statistical significance in the three groups after transfected 24 h(P>0.05), while after transfected 48,72,96 h,compared with the control group,OD value in HOXB7 group was significantly decreased(P<0.05);compared with the blank group,the apoptosis rate in HOXB7 transfection group increased significantly,and the expression of Bcl-2, Notch1 and Hes 1 proteins was down regulated,and the expression of Bax protein was up-regulated(P<0.05).Conclusion:RNA inter-ference in the expression of HOXB7 gene in colon cancer can inhibit the proliferation of cancer cells and induce apoptosis by inhibiting of the Notch1 signaling pathway.

Objective To investigate the cell morphology and expression of 5-hydroxytryptamine transporter(5-HTT) in hippocampal CA1 region of depression rats induced by adolescent and post-adult stress,and to observe the inter-vention effect of modified Sini San (MSS). Methods One hundred and thirty-two Wistar rats were randomly divided into blank group,model group,JSS group,and fluoxetine group,33 rats in each group. And then the rats in each group were randomly subdivided into adolescent group, adult group, post-adult group acaccording to the age day, 11 rats in each subgroup. Age day 44,56 and 78 were used as the sampling time points for adolescent group,adult group,post-adult group respectively. Chronic unpredictable mild stress(CUMS)rat model was used. From day 21 to 44 and from day 57 to 78, the rats were modeled and given medication, but from day 44 to 55, the rats were fed normally. The rat general condition and body mass of various groups were observed,the cell morphology of hippocampal CA1 region was observed by hematoxylin-eosin (HE) staining , and the distribution of 5-hydroxytryptamine transporter (5-HTT)positive cells in CA1 region of hippocampus was observed by immunohistochemical staining. Results The general condition of the rats at different age stages in the model group was poor,while that in MSS group and fluoxetine group was improved obviously. The body mass of rats at different age stages in the model group was obviously decreased (P<0.01 compared with the blank group). After adulthood stage,the body mass of rats in model group, MSS group, and fluoxetine group was lower than that of the blank group(P < 0.01), but there was no difference between the 3 groups (P > 0.05). In aspect of cell morphological manifestation in hippocampal CA1 region, rats in the adolescent model group had more deeply-staining atrophy neurons, with unclear hyperchromatic nucleus and cytoplasm. The morphological manifestations in modeled rats at adult stage and post-adulthood stage showed progressive aggravation,manifested as a large amount of neurons stained deeply with unclear nucleus and cytoplasm, and a small amount of glial cells proliferated. Compared with the model group at the same stage,the neuronal atrophy and deeply staining decreased in fluoxetine group and MSS group. The average optical density value of 5-HTT expression in the model group was decreased significantly at the adult stage and after adulthood stage(P<0.05 or P<0.01 compared with the blank group). Compared with the model group, the average optical density value of 5-HTT expression in MSS group after adulthood stage, and in the fluoxetine group at the adult stage and after adulthood stage were increased (P<0.05 or P<0.01). Conclusion Rats suffering CUMS in adolescence presents depressive behavior, and post-adult stimulation can aggravates depression. 5-HTT expression in hippocampus may be an important pathway for MSS to achieve the therapeutic efficacy.

OBJECTIVETo study the inhibition of the expression of bcl-xL gene induced by RNA interference in CNE-2Z cell line in addition to the inhibition of its proliferation and apoptotic induction.

METHODSSmall interfering RNAs targeting bcl-xL gene were synthesized by using web design software provided by Amnion and the silencer short interfering RNA (siRNA) construction kit; fluorescein-labeled siRNAs were done by FAM-silencer siRNA labeling kit; siRNAs were transfected into CNE-2Z cells by using lipofectamine 2000 reagent; siRNA transfection efficiencies were analyzed by fluorescent microscopy; down-regulation of bcl-xL was detected by RT-PCR; thiazolyl blue (MTT) assay was used to assess the cell growth; apoptosis of CNE-2Z cells was analyzed by flow cytometry.

RESULTSGreen fluorescence in the cells was seen clearly in FAM-labeled siRNA transfected group under the fluorescent microscope while none in the untransfected group. Different down-regulations of bcl-xL mRNA expression were found in the transfected groups. The expression of bcl-xL mRNA decreased by 10% - 70% in the siRNAs transfected CNE-2Z by RT-PCR scan analysis. The inhibitory rate of cell proliferation depended on time and concentrations to some extent. Different cell apoptosis could be induced by different concentrations of siRNA4.

CONCLUSIONSThe synthesized siRNAs in vitro were able to down-regulate the expression of bcl-xL There were different capabilities of the specific siRNAs down-regulation. The transient transfected bcl-xL siRNA4 could effectively inhibit the growth of the cancer cells and induce theirs apoptosis. It was suggested that the siRNA technique provide not only an extremely powerful tool for the functional analysis of genome but also a new method for anti-nasopharyngeal carcinoma gene therapy.

Apoptosis ; Cell Line, Tumor ; Cell Proliferation ; Gene Expression Regulation, Neoplastic ; Humans ; Nasopharyngeal Neoplasms ; genetics ; pathology ; RNA Interference ; RNA, Messenger ; genetics ; Transfection ; bcl-X Protein ; genetics