Objective To construct prokaryotic expression vector of XTP3 gene and induce the expression of fusion protein in E.coli,and prepare XTP3-specific rabbit polyclonal antibody,detect the specificity of the antibody in hepatic carcinoma tissue and normal liver tissue.Methods DNA fragment of XTP3 amplified by PCR was inserted into pET-32a(+) to construct prokaryotic expression vector pET-32a(+)-XTP3.After identified by sequencing,pET-32a(+)-XTP3 was transformed into E.coli BL21 and induced with IPTG.After analyzed by SDS-PAGE and Western blotting,the induced expression product was purified and renatured by Ni+ affinity column chromatography.The purified protein was used to immunize New Zealand rabbits to gain polyclonal antibody,and the polyclonal antibody was then detected by ELISA,immunohistochemistry and Western blotting.Results Prokaryotic expression vector pET-32a(+)-XTP3 was successfully constructed,and the XTP3 fusion protein of about 52kD was highly expressed in E.coli.DS-PAGE showed that the protein product was mainly in inclusion body.The purified protein and polyclonal antibody were obtained successfully.It was manifested by ELISA that the titer of polyclonal antibody was over 1∶128 000.Immunohistochemistry showed that XTP3 antibody presented membrane-positive in hepatic carcinomous tissue.Conclusions The recombinant XTP3 protein and polyclonal antibody have been obtained successfully.These results lay a foundation for studying the immuneogenicity and bionomics of XTP3 protein.

Objective To clone and identify human genes transactivated by homo sapiens HCV FTP2 by constructing a cDNA subtractive library with suppression subtractive hybridization tech- nique.Methods Suppression subtractive hybridization(SSH)and bioinformatics techniques were used for screening and cloning of the target genes transactivated by HCV FTP2.The mRNA was iso- lated from HepG2 cells transfected pcDNA3.1(-)-HCV FTP2 and pcDNA3.1(-)empty vector re- spectively,and SSH method was employed to analyze the differentially expressed DNA sequence be- tween the two groups.After digestion with restriction enzyme Rsa I,small-size cDNAs were ob- tained.Then tester cDNA was divided into two groups and ligated to the specific adaptor I and adap- tor 2 respectively.After tester cDNA was hybridized with driver cDNA twice and underwent two times of nested PCR and then was subcloned into T/A plasmid vectors to set up the subtractive library. Amplification of the library was carried out with E.coli strain DH5?.Futhermore,the cDNA was se- quenced and analyzed in GenBank with Blast search after PCR.Results The subtractive library of genes transactivated by HCV FTP2 was constructed successfully.The amplified library contains 71 positive clones.Colony PCR shows that 56 clones contain 200～1000 hp inserts.Sequence analysis was performed in 24 clones randomly,and the full length sequences were obtained with bioinformatics method.Altogether 20 coding sequences in total were obtained,consisting of 19 known and 1 un- known.Conclusion The obtained sequences may be target genes transactivated by HCV FTP2,and some genes coding proteins involved in cell cycle regulation,metabolism and cell apoptosis.

Animals ; Creatine Kinase ; blood ; Lactate Dehydrogenases ; blood ; Lipid Peroxidation ; Male ; Rabbits ; Superoxide Dismutase ; blood ; Weight-Bearing

Animals ; Creatine Kinase ; blood ; Energy Metabolism ; Lactate Dehydrogenases ; blood ; Male ; Pressure ; adverse effects ; Rabbits ; Serum ; enzymology

Objective To screen promoter binding protein of cyclin B2 by using human liver cDNA library, and investigate the expression and regulation mechanism of cyclin B2 gene. Methods By using cyclin B2 biotinylated promoter DNA as the selective molecule, the T7select human liver cDNA library was biopanned and positive clones were selected. After screening, positive plaques were performed to amplify for inserted DNA fragment, and the DNA fragment was cloned into pGEM-Teasy vector. Results Sequence analysis was performed in 20 positive plaques, and the full length sequences were obtained with bioinformatics method and searched for homologous DNA sequence from GenBank, altogether 6 coding sequences were obtained, all of which were known ones. Conclusion Cyclin B2 promoter binding proteins were screened. The results will be useful for further study the expression and regulation mechanism of cyclin B2.

Objective:To explore the effects of dexmedetomidine on the blood glucose,tumor necrosis factor-α (TNF-α) and interleukin-6(IL-6) in perioperative patients with neurosurgery.Methods:50 patients with meningioma who were ready to accept craniotomy in department of neurosurgery of our hospital from February 2016 to November 2016 were enrolled,they were randomly divided into research group and control group,with 25 patients in each group.The research group was given DEX 1.0 μg/kg at 10 min before anesthesia,which was added in 0.9％ sodium chloride injection 50 mL,the injection was finished in 10 min,then the DEX change to the rate of 0.5 μg/kg/h.Control group was given 0.9％ sodium chloride injection with constant rate and volume.Record heart rate (HR),mean arterial pressure (MAP) and blood glucose,serum TNF-α,IL-6 levels of the two groups at 10min before anesthesia(T1),after the anesthesia (T2),the beginning of surgery(T3),1 hour after the surgery beginning(T4),the end of surgery(T5).Results:Compared with T1,HR of the two groups at T2 and T3 raised significantly,but research group was lower than control group(P＜0.05);In research group,compared with T1,MAP at T3 raised significantly;in control group,compared with T1,MAP at T3 and T4 raised significantly,but MAP of research group at T3 and T4 were lower than control group (P＜0.05).Compared with T1,blood glucose of the two groups at T2,T3,T4 and T5 raised sig-nificantly,but the research group at same points were lower than control group (P＜0.05).Serum TNF-α,IL-6 levels of control group at T2,T3 and T4 were significantly higher than T1 (P＜0.05),which in research group at different time points had no significant change (P＞0.05),and were lower than control group at T2,T3 and T4 (P＜0.05).Conclusion:DEX could maintain stable hemodynamics,antiinflammation,inhibit elevated blood glucose,so as to reduce the inflammatory response and stress response in patients with neurosurgery,then it could promote these patients postoperative recovery.

Objective Through analyzing and summarizing the main pathogens of bacterial infections of children in Sichuan Province and trends of drug resistance in 2013 to provide a reference for the clinical use of antibiotics.Methods The pediatric pathogen were collected by member of Sichuan province in china antimicrobial resistance surveillance system.Results A total of 22 470 clinical bacterial were isolated from the members,in which Staphylococci,Escherichia coli,Streptococcus pneumoniae,Kleb-siella pneumoniae,Hemophilus influenza,Moraxella catarrhalis were the most common Bacteria.The resistance rates of Staphylo-coccus aureus and Coagulase-negative staphylococci to oxacillin were 1 7.7% and 71.1%.4 strains of Staphylococcus aureus and 20 strains of coagulase-negative staphylococci were teicoplanin resistance.Escherichia coli and Klebsiella pneumoniae were clearly re-sistant to the third generation cephalosporins,ampicillin and ampicillin/sulbactam,with the exception of ceftazidime.Carbapenems remained highly active against all the target bacteria.The resistance rate of Streptococcus pneumoniae to Penicillin was 3.9% .All antibiotics excepted cotrimoxazole remained highly active against the haemophilus influenzae.All antibiotics except macrolide anti-biotics remained highly active against the moraxella catarrhalis.Conclusion Penicillin-insensitive Streptococcus pneumoniae,mac-rolides-resistant gram-positive cocci,cephalosporin-resistant Enterobacteriaceae ,Oxacillin-resistance coagulase-negative staphylo-cocci were revealed to be the most serious problems in terms of bacteria resistance for children in Sichuan province.

OBJECTIVE To investigate the pathogens in the early period after heart transplantation and analyze their drug-resistance.METHODS The pathogens in the early period after heart transplantation were identified and their drug-resistance was analyzed.RESULTS From all of the 121 pathogens,the rate of G-bacilli was 73.6%,the rate of G+ cocci was 17.4%and the rate of fungi was 9.1%;G-bacilli mainly consisted of Klebsiella pneumoniae,Enterobacter cloacae,Acinetobacter baumannii,Pseudomonas aeruginosa and Escherichia coli.G-bacilli showed higher sensitive rates to sulbactam/cefoperazone,cefepime,piperacillin/tazobactam and ceftazidime than to cefotaxime.All G-bacilli showed sensitive to imipenem except Pseudomonas aeruginosa.G+ cocci mainly consisted of negative coagulase Staphylococcus,Staphylococcus aureus and Enterococcus.Fungi mainly consisted of Candida,and they were sensitive to fluconazole,itraconazole and amphotericin B.CONCLUSIONS To observe the pathogens in the early period after heart transplantation and analyze their drug-resistance are important to control and prevent the infection efficiently for the heart transplantation recipients.

Objective To study the biomechanical strength of calcium phoshphate cement (CPC) in fixating distal radial fractures. Methods A total of 18 human distal radius fracture models were made and randomly divided into three groups: fixed with Kirschner wire (Kirschner group), CPC (CPC group) or CPC combined with Kirschner wire (combination group). Compression test was performed to measure compression rigidity and rotational angle of distal fragment at 100 N and anticompression strength and maximum compression strength at 2 mm displacement. Results Compression rigidity, distal fragment rotation angle in coronal or sagittal plate had no significant statistical difference at 100 N of compression in three groups. Compression strength at 2 mm displacement in CPC group (average 375 N) and combination group (average 386 N) exceeded that in Kirschner group (average 116 N), with significantly statistical difference (P

Aim To investigate the effects of silencing MALAT1 gene on cell proliferation inhibition and apop-tosis induced by Melittin in human hepatocellular car-cinoma HepG2 cells. Methods The inhibitory rate of cell proliferation treated with Melittin in HepG2 cells was examined by MTT assay. Apoptotic rate was detec-ted by flow cytometry. The MALAT1 expression level in HepG2 cells was measured by qPCR. Specific siR-NAs were utilized to silence MALAT1 expression. The rates of cell proliferation inhibition and apoptosis in HepG2 cells treated with siRNA and Melittin were compared with those of Melittin alone. Results Melit-tin significantly suppressed the growth of HepG2 and induced cell apoptosis in a dose-dependent manner. Compared with normal liver cell lines, MALAT1 was highly expressed in HepG2 cells ( P<0. 05 ) . The ex-pression of MALAT1 in HepG2 cells was inhibited by Melittin, and the inhibitory rate increased with the in-crease of concentration. The rates of cell proliferation inhibition and apoptosis in HepG2 cells treated with siRNA and Melittin were significantly higher than those treated merely with Melittin. Conclusion Melittin can reduce the expression of MALAT1 and silencing MALAT1 can effectively promote proliferation inhibi-tion and apoptosis in HepG2 cells induced by Melittin.