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 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 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.

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 explore the clinical features of lissencephaly and the detection ofLISI gene.MethodsThe characteristic of clinical features, laboratory examination and gene detection in one case of lissencephaly was retrospectively analyzed. Meanwhile, the related literatures were reviewed.ResultsA 5-month-old female child diagnosed with epilepsy 20 days ago was hospitalized for convulsive seizure more than 30 times in 3 days. The manifestations were eyes staring, and turning upward, cyanosis of lips and face, froth at the mouth, extremities rigidity and loss of consciousness, and the symptoms can spontaneously remitted in 2-3 minutes. Laboratory examination showed that peripheral blood white cell count was 13.67×109/L, hemoglobin 108 g/L, red blood cell count 3.90×1012/L, lymphocyte 10.26×109/L; maocardial enzyme and hepatic and renal function were normal; blood ammonia was 23 μmol/L and lactic acid 2.11 mmol/L. Long-range video EEG showed highly arrhythmia, and frequent partial epilepsy, and sometimes secondary generalized epilepsy. Head MRI showed lissencephaly. The child was treated with oral administration of Keppra 27 mg/(kg·day), Topiramate 6.5 mg/(kg·day), currently no seizure. The detection ofLIS1 gene found that heterozygous mutation of c.232delG, which lead to protein shift mutation (p.E78NfsX25). No mutation was found in her parents.ConclusionsChild with lissencephaly may combine with epilepsy which may cause by mutation inLIS1 gene. And there was no information about point mutation of c.232delG inLIS1 gene being reported at home and abroad so far.

Objective: To investigate the clinical efficacy of treating herniation of lumbar intervertebral disc with electroacupuncture on Jiaji(Ex-B 2) points plus 3-D traction. Methods: To allocate 90 cases randomly into three groups and adopt therapies of 3-D traction, electroacupuncture on Jiaji(Ex-B 2) points and comprehensive method (combination of electroacupuncture on Jiaji(Ex-B 2) points and 3-D traction) and then compare the pre-treatment and post-treatment result with scores of clinical symptoms and clinical efficacy. Results: After 4-week treatment, the group of comprehensive therapy showed better effect than the other two groups. Conclusions:Electroacupuncture on Jiaji(Ex-B 2) points plus 3-D traction has positive effect on herniation of lumbar intervertebral disc.

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.