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 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.

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

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

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.

Objective To study correlation of the retinal nerve epithelium layer thickness measured with different optical coher-ence tomography (OCT) in vivo with histological measurement. Design Experimental study. Participants 15 rabbit eyes. Methods The retina measurement position of 15 rabbit eyes were marked by laser, and then were scanned by OSE-1800 OCT and Stratus OCT. Reti-nal nerve epithelium layer thickness was measured in retinal histological shdes of rabbit eyes. The results measured with three methods were compared and linear regression analyses were done with SPSS11.5 software. Results The average retinal nerve epithelium layer thickness measured with OSE-1800 OCT, Stratus OCT and histological method were 119.5±7.4, 118.0±5.6, and 116.3±8.8μm respec-tively(P=0.292). Retinal nerve epithelium layer thickness measured with both OCT instruments had the best correlation (r=0.914, P= 0.000), and the thickness measured with Stratus OCT and histological method had the better correlation (r=0.872, P=0.001), and the thickness measured with OSE-1800 OCT and histological method had the significant correlation (r=0.833, P=0.002). Conclusions The retinal nerve epithelium layer thickness measured with different OCTs in vivo correlate well with histomorphometry, and the measure-ment of both OCT instruments are accurate. (Ophthalmol CHN, 2009, 18: 239-242)

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.

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.

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.

Objectives To evaluate the effect of control measures and provide the basis for development of preventive measures though dynamical monitoring of the operation of water improvement projects and know The prevalent trends of endemic fluorosis.Methods In accordance with the provisions and requirements of the Surveillance Scheme of Drinking-Water-Borne Endemic Fluorosis(Trial),136 monitoring counties were selected in national 27 provinces(autonomous regions,municipalities) and Xin Jiang Production and Construction Corps,and 10 water improvement projects were selected in each county.The operation of water improvement projects was investigated,and water fluoride content was tested.Three epidemic villages were selected as fixed monitoring villages in each county.In the villages,the fluoride content in drinking water was detected by Standard Testing Method of Drinking Water.Dental fluorosis was diagnosed by Dean's method of all 8 to 12-year-old students in the villages.Stratified random sampling by age was used to select 50 adults over 25-year-old in each monitoring village,and skeletal fluorosis was examined by X-ray according to the Diagnosis Standard of Endemic Skeletal Fluorosis.Results ①A total of 1398 water improvement projects were monitored.Normal and intermittent operation and discarded projects accounted for 93.35％(1305/1398),5.29％(74/1398) and 1.36％(19/1398),respectively.The qualification rate of water fluoride content was 69.96％ (978/1398).② In the 327 surveyed villages with water improvement projects,the qualification rate of water fluoride content of the projects accounted for 76.15％(249/327).In the 84 surveyed villages without water improvement projects,villages of water fluoride ＞ 1.2 mg/L and ≤2.0 mg/L,＞ 2.0 mg/L,and ≤4.0 mg/L,and ＞ 4.0 mg/L,accounted for 57.14％ (48/84),32.14％(27/84) and 10.71％(9/84),respectively.③ In the 195 villages with water improvement projects and the projects were in normal operation with qualified water,the detection rate of dental fluorosis and the defect type of dental fluorosis in children were 25.03％ (3851/15 387) and 1.88％ (289/15 387),respectively,and for the 82 villages with fluoride content exceeded the national standard,the two rates were 56.27％ (3384/6014) and 10.92％(657/6014),respectively.④ In the 195 villages with water improvement projects and the projects were in normal operation with qualified water,the detection rate of X-ray skeletal fluorosis was 19.11％ (1465/7667).In the 79 villages with fluoride content exceeded the national standard,the detection rate of X-ray skeletal fluorosis was 17.47％ (634/3630).Conclusions The operation of water improvement projects and water quality need to be further improved,and the measure of changing water plays a certain role in control of drinking-water-borne endemic fluorosis,which needs a long-term consolidation.