Objective To establish drug resistant models of temporal lobe epilepsy induced by amygdala kindling,and to investigate the changes of cAMP response element binding protein (CREB) and phosphorylated cAMP response element binding protein (p-CREB) expression in the hippocampus tissues in order to explore their roles in drug resistant epileptogenesis.Methods Eighty adult male SD rats were randomly divided into control group (n =10) and model group (n =70).The 70 rats were used to prepare the amygdaloid kindled model of epilepsy by chronic stimulation of amygaloid basal lateral nucleus.The successful kindled models were randomly selected as drug resistant epileptic group (n =10) and drug sensitive epileptic group (n =10) according to their response to the phenytoin and phenobarbital.On the basis of behavioral observation,electrophysiology,pathological HE staining,CREB and p-CREB expression changes,we verified the reliability of the models and explored the differences among the three groups above.The changes of CREB and p-CREB expression were detected by immunohistochemical method and Western blotting assay.Results In control group,the electroencephalogram (EEG) frequency was (8.700 ±1.494) Hz;in drug sensitive epileptic group,the EEG frequency was (14.700 ± 1.159) Hz;in drug resistant epileptic group,the EEG frequency was (19.800 ± 1.686) Hz.The frequency differences among the three groups were statistically significant (F =144.202,P =0.000).By immunohistochemical staining,a large number of CREB and p-CREB positive cells were observed in drug resistant epileptic group.As compared with the control group (CREB 0.197 ±0.058,p-CREB 0.260 ±0.176),the expression levels of CREB and p-CREB were increased in drug sensitive epileptic group (CREB 0.361 ±0.151,p-CREB 0.656 ±0.234) and in drug resistant epileptic group (CREB 0.591 ± 0.150,p-CREB 1.077 ± 0.400).The difference among the three groups had statistical significance (F =24.206,20.376,both P ＜ 0.01).Conclusions The expressions of CREB and p-CREB were significantly increased in drug resistant epileptic rats.These findings indicate that the expressions of CREB and p-CREB may play certain roles in the drugresistant epileptogenesis.

Objective To study the effects of A?25-35 on the expression of gene P21,CDK4,E2F1 and BAX in cultured PC12 cells.MethodsPC12 cells were treated with 25 ?mol/L A?25-35,the relation between cell cycle redistribution and apoptosis was analyzed by flow cytometry(FCM).Protein and mRNA expression of P21,CDK4,E2F1 and BAX were detected by RT-PCR and Western-blot respectively.Results About 90% PC12 cells were found arrest on G0/G1 by FCM being deprived serum.Treated with 25 ?mol/L A?25-35 for 8,16,24 h,the percent of S phase cells was raised remarkably(P

BACKGROUND:Gene therapy is the direction of spinal cord injury(SCI) therapy,the key of which is construction of targeting gene and vector. OBJECTIVE:To construct the recombinant adenovirus vector carrying human brain-derived neurotrophic factor(hBDNF) marked enhanced green fluorescent protein(EGFP). DESIGN,TIME AND SETTING:A single sample observation was completed in the First Affiliated Hospital of Fujian Medical University from September 2007 to June 2008. MATERIALS:Competent E. coli DH-5? was obtained from the American Stratagene Company. Plasmid pDC316-hBDNF,pDC316-mCMV-EGFP,pBHGlox_E1,3Cre and package system AdMax and 293 package cell strain were purchased from the Canadian Mixcrobix-Biosystems Company. METHODS:The hBDNF gene was constructed by PCR with plasmid pDC316-BDNF as template. With enzyme digestion,the hBDNF gene was inserted into the vector pDC316-mCMV-EGFP and the shuttle plasmid pDC316-hBDNF-mCMV-EGFP was constructed,which was cotransfected with the adenovirus skeleton plasmid pBHGlox_E1,3Cre into 293 cells to obtain the produced replication defective recombinant adenovirus vector Ad-hBDNF-EGFP. The recombinant adenovirus was propagated by repeat infection of 293 cells and purified by ion exchange method,then the virus particles were counted and the purity and titer were determined. MAIN OUTCOME MEASURES:①PCR identification of plasmid pDC316-hBDNF. ②Construction and identification of the shuttle plasmid pDC316-hBDNF-mCMV-EGFP. ③Packing,amplification and purification of recombinant adenovirus vector Ad-hBDNF-EGFP. ④PCR identification of the recombinant adenovirus. ⑤Titer of recombinant adenovirus. RESULTS:PCR amplification,restriction analysis and sequencing identified that both recombinant shuttle plasmid pDC316-hBDNF-mCMV-EGFP and recombinant adenovirus vector Ad-hBDNF-EGFP were correctly constructed. After amplification and purification,the virus particle count,A260/A280 and titer of recombinant adenovirus were 2.4?1011 VP/mL,2.0 and 0.8?1010 CCID50/mL,respectively. CONCLUSION:Recombinant adenovirus vector Ad-hBDNF-EGFP is successfully constructed,which laid a foundation for further study regarding gene function and therapy.

OBJECTIVE To explore the characteristics of the pathogens in ventilator-associated pneumonia (VAP) following severe craniocerebral trauma. METHODS The 220 cases with severe craniocerebral trauma were analyzed retrospectively. RESULTS The incident rate of VAP was 24% in these patients. A total of 169 pathogens were isolated,including 146 ( 86%) strains of bacteria and 23 (14%) fungal isolates. The main Gram-negative bacteria (114 strains) were Pseudomonas aeruginosa,Escherichia coli,Klebsiella pneumoniae,etc. The Gram-positives (32 strains) were Staphylococcus aureus and twenty-four (75%) of them were meticillin-resistant. CONCLUSIONS The incident rate of VAP is high following severe craniocerebral trauma. Most of the isolated pathogens are drug resistant. These cases have mixed infection occurred commonly. To prevent and treat VAP,we should pay attention to disinfection and aseptic manipulation,and make the rational use of antimicrobial agents.

Aim To study the effective mechanism of curcumin on abnormal cell cycle and apoptosis of serum-deprived PC12 cells induced by ?-amyloid peptide 25-35(A?25-35).Methods Synchronized PC12 cells were pretreated with 5 ?mol?L-1 Cur for 1 h,and then 25 ?mol?L-1 A?25-35 for 24 h.Protein and mRNA expression of p21,CDK4,E2F1 and bax were detected by RT-PCR and Western blot respectively.Results After synchronized PC12 cells being pretreated with 5 ?mol?L-1 Cur for 1 h,the mRNA and protein expression of p21 gene were increased gradually,while CDK4,E2F1 and bax gene were decreased.Conclusion Cur maybe effects the redistribution of cell cycle and apoptosis of serum-deprived PC12 cells induced by A?25-35,through increasing the mRNA and protein expression of p21,and decreasing the mRNA and protein expression of CDK4,E2F1 and bax gene.

Objective To investigate the clinical value of antibody D5F3 and Survivin in patients with non small cell lung cancer.Methods 200 paraffin embedded specimens of patients with non small cell lung cancer tested by RT-PCR(including EML4-ALK mutant and wild type)from October 2012 to June 2014 were selected.The gene protein expression were tested by ALK new antibody D5F3 and the sensitivity and specificity were compared by negative and positive(+～3+).ResultsTest results show(+)the coincidence rate is 15.78％,(+ +)the coincidence rate is 27.27％and(+ + +)compliance rate was 87.5％,the difference was statistically significant(P<0.05).The expression of Survivin protein in NSCLC tissues was correlated with clinicopathological features.The positive rate of Survivin protein expression was correlated with the degree of differentiation(P<0.05),but not with other clinical and pathological features,with the decrease of differentiation,the positive rate of Survivin protein expression was significantly increased.Conclusion D5F3 and Survivin antibodies are highly sensitive and specific in patients with NSCLC,with the screening value,save social resources,for the majority of patients with lung cancer services.

Objective To investigate the clinical distribution and drug resistance of multidrug-resistant (MDR)Pseudomonas aeruginosa to provide reference for clinical rational use of antibacterial drugs.Methods The strains were originated from bacterial culture in the samples of sputum,urine and wound secretion collected from the inpatients admitted in our hospital from January 2009 to December 2013.Results 200 strains of Pseudomonas aeruginosa were isolated during this period,in which 51 strains were MDR,accounting for 25.5%.The isolated MDR Pseudomonas aeruginosa strains were mainly distributed in the the respiratory de-partment(80.4%)and the surgical department(17.9%).MDR strains isolated from the sputum specimens accounted for 64.7%, followed by the wound secretion(19.6%)and urine(11.8%).Pseudomonas aeruginosa was seriously resistant to ampicillin-sulbac-tam with the resistance rate of was 100.0%,but had the low resistance rate to amikacin,imipenem,meropenem,piperacillin/tazobactam,ciprofloxacin,levofloxacin,polymyxin E,which were 28.9%,29.4%,25.0%,30.4%,34.5%,34.5% and 8.3%;the resistance rate to commonly used antibacterial drugs was 39.0%-58.3%.14 strains were pan-drug resistant Pseudomonas aerugi-nosa(PDR-PA);The detection rate was 7.0%.Conclusion The drug resistance of MDR Pseudomonas aeruginosa is more serious, the clinical application of anibiotics should be perfromed based on the result of drug susceptibility test.

BACKGROUND:Inactivated autologous replantation in repair of bone defects after bone tumor resection has obvious advantages. Boiling, alcohol soaking, cryogenic freezing, microwave, radiation and other methods have been used for inactivation; however, they al have shortcomings. OBJECTIVE:To study the effect and feasibility of high temperature and high pressure inactivated autologous bone in repair of large segmental bone defects. METHODS: Bone defect models of bilateral distal radius were established in New Zealand white rabbits. Bone defect at the right side was repaired by high temperature high pressure inactivated autologous bone via in situ replantation, as experimental group. Bone defect at the left side was repaired byin situbone replantation, as control group. The general observation of bilateral radius, X-ray detection, bone radionuclide scan test and histological examination were conducted at the 6th, 12th and 24th weeks after surgery. RESULTS AND CONCLUSION: At the 24th week after surgery, X-ray films showed normal bone healing in these two groups. At the 24th week after surgery, bone radionuclide scan test showed that in the experimental group, the radiation on the repaired bone segments was uneven; the concentrations of nuclear elements were stil slightly higher on both ends, but decreased in the middle area; and the concentration in the control group was closed to normal. At the 24th week after surgery, histological observation showed that there were a majority of trabecular bone tissues in the bone defect area of the experimental group, and some woven bone tissues were immature, which was similar to the performance of the control group at the 12th week after surgery; and normal bone was visible in the control group. These results demonstrate that high temperature and high pressure inactivated autologous bone can be used to repair long segmental bone defects, but can result in delayed bone healing.

BACKGROUND: Pilose antler polypeptides (PAP) have been proved to promote the proliferation of condrocytes cultured in vitro and expressions of glycosaminoglycan (GAG), type Ⅱ collagen and Aggrecan protein in the extracellular matrix.OBJECTIVE: To investigate the feasibility of chondrogenic phenotype differentiation of rabbit bone marrow-derived mensenchymal stem cells (BMSCs) in a defined medium and then to study the effect of PAP on chondrogenic differentiation of BMSCs in vitro.METHODS: The third passage BMSCs from rabbits were randomly divided into control group cultured in ordinary medium, induced group cultured in defined medium, and PAP group cultured in defined medium containing 10 mg/L PAP. An equal volume of articular chondrocytes were selected from rabbits as articular cartilage group. The cellular morphological and functional characteristics were observed after 1, 2, 3 weeks in centrifuge tubes by histological, biochemical and reverse transcription-polymerase chain reaction (RT-PCR) technique. RESULTS AND CONCLUSION: Cell masses in the control group gradually crumbled after 2 weeks, and hematoxylin-eosin staining could not be done. Cell masses in the induced and PAP groups were semitransparent, but slightly contracted. A part of these cells were round or oval with a high density distribution at the surface. The content of GAG and mRNA expression of type Ⅱ collagen in the induced and PAP groups were increased with culture time, and higher than those in the control group at different time points (P < 0.05). The content of GAG and mRNA expression of type Ⅱ collagen in the PAP group were higher than those in the induced group, but lower than those in the articular cartilage group (P < 0.05). The results indicated that BMSCs can differentiate into chondrogenic phenotype in the defined medium, and PAP can significantly enhance chondrogenic phenotype differentiation of BMSCs. But the quality of cultured cartilage tissue is poorer than that of the articular cartilage.

Objective To investigate the expression and clinical significance of zinc finger gene 217 (ZNF217) in human papillary thyroid carcinoma.Methods The expressions of ZNF217 mRNA and protein were detected by quantitative realtime PCR and Western blot in papillary thyroid carcinoma tissues (n =20) and adjacent normal tissues (n =20),and the data were analyzed.Results The expressions of ZNF217 mRNA and protein in papillary thyroid carcinoma were significantly higher than those in adjacent normal tissues (96.72 ± 44.19 vs 4.86 ±3.55,0.994 ± 0.172 vs 0.195 ± 0.061,both P＜0.01),being higher in the papillary thyroid carcinoma with capsule invasion compared with that without capsule invasion (P＜0.01).The expressions of ZNF217 mRNA and protein in papillary thyroid carcinoma were not related to gender,age,tumor size,TNM stage or lymph node metastasis (all P＞0.05).Conclusions The overexpression of ZNF217 may be associated with the oncogenesis and progress of papillary thyroid carcinoma and capsule invasion,and thus is expected to become a new target for prevention and treatment of papillary thyroid carcinoma.