Objective To construct an infectious full-length cDNA clone of enterovirus 71(EV71)and develop a technological platform for study on vaccine development as well as molecular virology of EV71.Methods According to the nucleotide sequence of EV71 strain 085 isolated in China,four pairs of primers were designed for amplification of four end to end overlapping subgenomic cDNA fragments,the cDNA fragments were directional cloned into pBluescript SK(+)vector,and the virus genome cDNA clone was obtained by ligation orderly.The rescued virus of parental strain 085 from RNA transfected host cells was identified by RT-PCR,IFA,titration as well as transmission electron microscope(TEM)after the transcription of the full-length cDNA clone in vitro.Results The full-length cDNA clone was constructed successfully,and the typical CPE was observed after its transcription into Vero cells.The rescued virus with 20-30 nm in diameter can not only be neutralized by EV71 special anti-serum but also react with anti-EV71 monoclonal antibody that virus infected cells stained with FITC can be detected by IFA.After amplification from the total RNA extraction of virus infected cells by RT-PCR with EV71 special primers,the 226 bp products can be detected.The growth curve showed that the rescued virus can propagate in Vero cells stably with a titer of 4.5 ～6.0 lgCCID50/ml during 8 passages.The plaque formed by rescued virus is identical as parental virus in morphology but smaller in size.Conclusion An infectious full-length clone of EV71 was developed successfully,which will be used for further study on pathogenesis and vaccine development of EV71.

To evaluate the clinical effects of rotary nickel-titanium instruments Mtwo in root canal therapy in the aged patients. Using step-back technique,80 teeth with pulpal and periapical involvement were instrumented by Mtwo in the M group, and by K file in the K group. Mtwo could keep the original curvature and flow of the root canals. No transportation, apical blockage, ledge or perforation was found in the M group. There was more complications in the K group than in the M group.The operative time was shorter and posttreatment pain seldom occurred in the M group. With rotary NiTi instruments Mtwo for seniles' root canals treatment, root canals can be prepared effectively and quickly,and is worth of clinical application.

Objective:To establish an in vitro root canal model infected by Enterococcus faecalis and to observe the morphology, distribution and relative position of Enterococcus faecalis in infected root canals.Methods: Ten human healthy premolars extracted for orthodontic reasons were collected. Following sterilization , a total of 5 specimens were aseptically transferred to separate Eppendorf tubes containing 1. 5 mL brain-heart infusion broth (BHI) inoculated with 0.1 mL Enterococcus faecalis suspension that had been adjusted to Mcfarland 5, and were incubated at 37℃ for 21 days. The other 5 specimens were as controls. The roots of all specimens were then split into two halves along the mesiodistal axis. One half was processed with light microscopic ( Brown & Brenn stain) to check the bacteria in dentinal tubules, and the other was observed with SEM to investigate the bacterial status in infected root canals. Results: Enterococcus faecalis could penetrate into the dentinal tubules about 330-1 000μm. A dense bacterial aggregation composed of Enterococcus faecalis and amorphous matrix was observed in the apical third of the root canals, whereas Enterococcus faecalis were seen free-floating or planktonic in the crown and middle third of the root canals . No microorganisms were found in the root canals of the controls. Conclusion:Enterococcus faecalis could form bacterial biofilm on the root canal walls and penetrate into the dentinal tubules. The in vitro model designed was simple, and had good practicability to make a further comparative evaluation of various antimicrobial methods in the reduction of intracanal bacteria.

Objective:To investigate the patterns of microbial infection on the apical external root surfaces of treated and untreated teeth associated with chronic apical periodontitis and to study bacteria in the biofilm in order to find out the species,constitution and origination of bacteria in periapical biofilm.Methods: Ten teeth with chronic apical periodontitis from patients of the Department of Stomatology of People's Hospital,Peking University: 5 untreated teeth with a radiographically visible chronic periradi-cular lesions and 5 teeth with extensive carious lesions,radiolucent lesions of varying sizes and attached periradicular tissues were selected for study.Using aseptic techniques and sterile instruments,bacterial samples of the root canals were taken,inoculated and separated according to usual practice.After extraction,ten teeth were fixed and the apical 5 mm portion of one root was sectioned.Root tips were dehydrated,sputter coated with gold,and then examined for the occurrence of bacteria on the apical root surfaces using scanning electron microscope.Five healthy teeth with vital pulp were used as controls.Results: Microbial study showed that ten specimens yielded bacterial growth.The most prevalent bacteria were P.micros and F.nueleatum.In the 5 untreated teeth,bacterial cells were usually observed close to the apical foramen in only 1 specimen.Morphologically,these bacteria consisted of cocci.In the 5 treated teeth,a dense bacterial aggregation composed mainly of cocci and rods was observed surrounding the apical foramen of all specimens.Besides rods,other bacterial morphological types were recognized,including coaggregations of cocci and filaments,characterizing a fully developed "corn-cob".No microorga-nisms were found in the healthy controls.Conclusion:Bacterial biofilm was always present in teeth with post-treatment endodontic disease.The presence of apical bacterial biofilm is clinically important,and it may cause failure of endodontic treatment as a consequence of persistent infection.

Objective:To establish an in vitro root canal model infected by Enterococcus faecalis and to observe the morphology,distribution and relative position of Enterococcus faecalis in infected root canals.Methods: Ten human healthy premolars extracted for orthodontic reasons were collected.Following sterilization,a total of 5 specimens were aseptically transferred to separate Eppendorf tubes containing 1.5 mL brain-heart infusion broth(BHI) inoculated with 0.1 mL Enterococcus faecalis suspension that had been adjusted to Mcfarland 5,and were incubated at 37 ℃ for 21 days.The other 5 specimens were as controls.The roots of all specimens were then split into two halves along the mesiodistal axis.One half was processed with light microscopic(Brown & Brenn stain) to check the bacteria in dentinal tubules,and the other was observed with SEM to investigate the bacterial status in infected root canals.Results: Enterococcus faecalis could penetrate into the dentinal tubules about 330-1 000 ?m.A dense bacterial aggregation composed of Enterococcus faecalis and amorphous matrix was observed in the apical third of the root canals,whereas Enterococcus faecalis were seen free-floating or planktonic in the crown and middle third of the root canals.No microorganisms were found in the root canals of the controls.Conclusion: Enterococcus faecalis could form bacterial biofilm on the root canal walls and penetrate into the dentinal tubules.The in vitro model designed was simple,and had good practicability to make a further comparative evaluation of various antimicrobial methods in the reduction of intracanal bacteria.

Objective: To compare and analyze the antimicrobial efficacy of three different mechanical preparation techniques in single infected root canals. Methods; Forty-five single root canals with chronic periapical periodontitis were selected. The specimens were divided into three groups randomly, 15 root canals per group. Croup A: preparation with stainless steel K-files (step-back technique), Croup B: preparation with HERO 642 NiTi rotary files (crown-down technique) and Group C: preparation with Mtwo NiTi rotary files ( modified crown-down technique). The sterile normal saline was used as irrigation. Samples were taken before and after canal preparation. The difference of CFU was calculated as well as the bacterial species. Results; All groups were effective to reduce bacteria within the infected root canals greatly(P<0.01). Croup A and Group C were statistically better than Group B(P<0.05). Group A was more effective than Group C but there was no statistically difference between them(P>0.05). Conclusion; Mechanical preparation can greatly reduce the intracanal bacteria, but can not obtain bacteria-free canals. The mechanical preparation must be aided by chemical irrigation to improve the success of root canal therapy.

Objective Curative effect of 2 drugs which present elinie used in atrium filament trembles,provides the elinieal experience.Methods Choice outpatient service atrial fibrillation trembles the patient 344 eases,plants stochastically according to sickness divides into two groups,takes the rope separately Sotalol and Amiedarone,the adjustment medication dosage treats and observes for 12 months,carries on the material summary.Results The rope Sotalol group and the Amiedarone group each is 172 people,the rope Sotalol group atrial fibrillation trembles transfers sinous rhythm 45 peoples,transfers rate 26.2%,recurs 5 patients,after transfers sinous rhythm maintenance rate 88.9% ; Amiodarone,group atrial fibrillation trembles transfers sinus rhythm 41 people,transfers rate 23.8%,recurs 2 patients,after transfers sinous rhythm maintenance rate 95.1% ; Conclusion Sotalol group treats atrial fibrillation trembles rate to be higher than the Amiedarone group,after transfers sinous rhythm maintenance rate Amiedarone group to be higher than Sotalol group.

Aim To investigate the effect of chronic intermittent hypobaric hypoxia ( CIHH) on the paeonol induced vasomotion of isolated rat ’ s thoracic aorta rings and its underlying mechanisms. Methods Spra-gue-Dawlay ( SD ) rats were randomly divided into 2 groups: control group ( CON ) and CIHH treatment group ( CIHH) . CIHH rats were exposed to hypoxia in a hypobaric chamber simulating 5 000 m altitude, 6 hours daily for 28 days. CON rats lived in the same en-vironment as CIHH animals except hypoxia. Organ bath technique was used to observe the effect of pae-onol on isolated thoracic aorta rings of rats. Results There were no significant differences of noradrenaline ( NE )- and KCl-induced contraction in thoracic aorta rings among CIHH and CON rats;CIHH enhanced va-sodilative effects of paeonol on isolated thoracic aorta rings of rats; the vasodilative effects on CIHH rats could be partly decreased by β-receptor blocker prop-ranolol,ATP-sensitive potassium channel ( KATP ) bloc-ker glibenclamide and NO synthase inhibitor L-NAME. Paeonol significantly inhibited NE-induced intracellular and extracellular calcium-dependent contraction in CIHH rats. Paeonol didn ’ t inhibit NE-induced con-traction by intracellular calcium release and its inhibi-tory effect couldn ’ t be blocked by glibenclamide in CON. Vasodilative effects of paeonol couldn ’ t be re-versed by indomethacin, a cyclooxygenase inhibitor, in CIHH and CON rats. Conclusion CIHH significantly enhances vasodilative effects of paeonol on isolated tho-racic aorta rings of rats. Besides promoting the signa-ling pathway of paeonol in CON, CIHH significantly enhances vasodilative effects of paeonol via activating KATP and inhibiting Ca2+ release from sarcoplasmic re-ticulum.

BACKGROUND: Cristata L flavonoid is a kind of plant estrin, which possesses multiple physiological function, has no toxicity and adverse effect, and is effective in treating and preventing osteoporosis.OBJECTIVE: To study the effect of cristata L flavonoid on bone morphogenetic protein, urine inorganic salt and content of lysozyme of rats with diabetes mellitus (DM).DESIGN: Randomized grouping design and controlled study.SETTING: Pharmaceutical Laboratory of Xinxiang Medical College.MATERIALS: The experiment was completed in the Xinxiang Medical College from September to December 2003. Totally 24 health male SD rats were randomly divided into three groups: normal control group, diabetes mellitus (DM) group, DM + cristata L flavonoid group with 8 in each group.were injected intraperineally with 60 mg/kg streptozotocin, and 48-72 hours later, blood of rear caudal vein was collected to measure total blood glucose.Rats were determined as DM if the blood glucose was ≥ 16.7 mmol/L; oth erwise, 60 mg/kg streptozotocin was injected once more. After modeling,cristata L flavonoid and rats in normal control group were given the same was measured with atomic absorbency method and content of urine sodium histochemistry of bone morphogenic protein-2 (BMP2) in bone was detected with streptavidin tagged by peroxidase and immunohistochemic expression lysozyme reflected reabsorption function of renal tubule was measured with with t-test. MAIN OUTCOME MEASURES: Comparison between content of calcium, sodium, kalium in urine and lysozyme and immunohistochemic expression of BMP2 10-week intervention later.calcium and sodium in urine in DM model group were obviously higher than those in normal control group, but content of kalium in urine was obviously lower (P ＜ 0.05-0.01). Contents of calcium in urine in DM +cristata L flavonoid group were obvioualy lower than those in DM model group and normal control group were obviously higher than those in DM model group, and content of lysozyme in urine in those groups was obviously lower than that in DM model group (P ＜ 0.05-0.01).CONCLUSION: Expression of BMP2 of DM rats is decreased, but after supplement of cristata L flavonoid compound, the expression is increased.In addition, output of urine calcium and sodium in DM rats is decreased,and reabsorption function of renal tubule is increased.

Objective To establish rabbit model of remnant carcinoma after RFA therapy, and to observe pathomorphological changes of remnant carcinoma in different time. Methods Forty-eight New Zealand white rabbits underwent ultrasound-guided percutaneous inoculation with VX2 carcinoma, then RFA therapy was performed to made models of remnant carcinoma. These models were averagely divided into 6 groups randomly (each n=8). Rabbits in each group was killed and pathologically observed before RFA and 1 d, 3 d, 1 week, 2 and 3 weeks after operation, respectively. Results The expression of MVD,VEGF and PCNA in remnant VX2 carcinomas tissues decreased significantly, but increased 2-3 weeks after RFA. The remnant VX2 carcinomas tissues were in inhibitory state 2 weeks after RFA. Conclusion The growth of remnant carcinoma could be inhibited in short term after RFA. Further therapy is necessary.