Objective To investigate the effect of non-structural protein amino acid mutations(NS1-G53D,NS3-E250V)on viral proliferation and virulence of Dengue virus(DENV)type 4 Ban18HK20 strain.Methods The mutant sites in Ban18-HK20 strain were determined by sequence alignment with an attenuated strain of DENV(PDK53). Point mutation plasmids pSPTM-Ban18HK20(G53D),pSPTM-Ban18HK20(E250V)and pSPTM-Ban18HK20(G53D+E250V)were constructed by using homologous recombination technique,and identified by enzymatic cleavage and sequencing. Viral RNA was obtained by in vitro transcription and then electrotransfected to Vero cells to rescue the point mutant virus. The biological characteristics of point mutant virus were identified by plaque assay,indirect immunofluorescence assay,virus genome sequencing,growth kinetics test,CCK-8 test and mouse neurotoxicity test.Results Enzyme digestion and sequencing confirmed that the point mutation plasmids were constructed correctly. The results of immunofluorescence assay and virus sequencing showed that the virus was rescued successfully. The point mutation virus showed smaller plaque than the wildtype virus. The virus titers of Ban18HK20(G53D),Ban18HK20(E250V)and Ban18HK20(G53D+E250V)reached the peak on the 4th day,which were 7. 67,7. 84 and 7. 78 LgPFU/mL,respectively. The titer of the wild-type virus reached the peak value 7. 68 LgPFU/mL on the 5th day. The proliferation activity of BHK21 cells infected with Ban18HK20(G53D)was significantly higher than those infected with the wild-type virus(P = 0. 003 6)and the proliferative activity of C6/36 cells infected with Ban18HK20(G53D)and Ban18HK20(G53D+E250V)was significantly higher than those infected with the wild-type virus(P < 0. 000 1). Both the point mutation virus and wild-type virus had no neurotoxicity to 4-week-old BALB/c mice. The neurotoxicity of point mutation viruses Ban18HK20(E250V)(LD_(50)=8. 51 PFU)and Ban18HK20(G53D+E250V)(LD_(50)= 0. 69 PFU)on 3-day-old BALB/c neonatal mice was lower than that of the wildtype virus(LD_(50)= 0. 69 PFU). The survival rate(60%)of nude mice injected with point mutation virus Ban18HK20(G53D +E250V)was higher than those injected with the wild-type virus(0)after intracerebral injection of the same amount of virus. The point mutation virus was genetically stable,and no recurrent mutation occurred in Vero cells after passage to the10th generation.Conclusion The non-structural protein NS1-G53D amino acid mutation weakens the cytotoxicity of Ban18HK20 strain to C6/36 and BHK21 cells,and the two-point combined mutation could weaken the neurotoxicity of Ban18HK20 strain in nude mice,which laid a foundation of the study of DENV virulence sites and the development of the vaccine.

Objective To discuss the therapeutic efficacy of budesonide and compound ipratropium bromide inhalated in treatment of infant bronchiolitis.Methods From January 2005 to February 2007,150 infants with bronchiolitis were randomly divided into treated group and control group.Both the two groups were treated with comprehensive treatment and antiviral therapy;control group were treated with budesonide suspemion and compound ipratropium bromide inhalated.Clinical efficacy of both two groups was evaluated.Results Cure rate of treatment group was 91%,cure rate of control group was 71%,the clinical therapeutic efficacy were compared between the two groups,the difference of which had statistical significance.Conclusion The result of experiment indicates that the therapeutic efficacy of budesonide and compound ipratropium bromide inhalated in treatment of infant bronchiolitis is better method.

Objective To observe the therapeutic effect of Famotidine combined with Hemocoagulase treating the neonatal stress ulcer complicating upper gastrointestinal hemorrhage. Methods 72 patients of stress ulcer complicating upper gastrointestinal hemorrhage were randomly divided into two groups:study group(35 cases) were treated with Famotidine plus Hemocoagulase;control group(37 cases)were cured by Famontidine only. The different therapeutic effects were compared. Results 35 cases of study group(100%)and 28 cases of control group(75.68%) were cured successfully. Compared the excellent rate of study group and control group,the former was significantly higher than the latter(P＜0.01). Conclusion Famotidine combined with Hemocoagulase treating the neonatal stress ulcer complicating upper gastrointestinal hemorrhage has notable therapeutic effect.

Objective:To investigate the injury on isolated cadaver testes induced by ischemia-reperfusion (I/R).Methods:13 isolated cadaver testes contributed by 13 persons were preserved under 0℃～4℃ hypothermia and then reperfused under 37℃.Histology and histochemical changes were observed.Results:4℃ cold ischemia in 18 hours only could induce trivial swelling and vascular degeneration of endothelial cells(ECs).Obvious pathologic changes occurred after 24 hours,including detachment of ECs,separation between basement membrane and seminiferous epitelium,malalignment of spermatogenous cell and edema of mesenchyme.Injury was worse along with the prolongation of cold preservation time.Changes of LDH and SDH activities were found by histochemical staining.Reperfusion following 6 hours ischemia induced tissue injury and unusual enzyme activity.All changes were more obvious after reperfusion following 12,18,24 or 36 hours of cold ischemia.Conclusion:HCA can preserve testes for 18 hours.Testes can't keep normal histological and histochemical structure beyond 24 hours of preservation under 4℃ hypothermia,and 37℃ reperfusion can make injury worse.

Professor LI De-lin is sophisticated in the clinical experiences of treating diabetes mellitus(DM)with traditional Chinese medicine(TCM).Based on clinical observations for many years,the key pathogenesis of DM resulted from internal organs'deficiency,endogenous heat exuberance,liquid and qi's consumption as well as blood density and turbid fluid thickness.From the knowledge above,professor LI advocated to treat DM and diseases accompanied by it with the combination of treatment based on disease and syndrome differentiation to improve the curative effects.During the specific treatments,professor LI emphasized that treatment based on disease differentiation should benefit qi,nourish yin and strengthen yang;treatment based on syndrome differentiation should strengthen vital qi and eliminate pathogenic factor.Strengthening vital qi includes to benefit qi ,nourish blood,tonify yang and supply yin.Eliminating pathogen includes to treat heat-toxicity,phlegm,dampness and depression at the same time.He advocated firstly to invigorate spleen and kidney,secondly to clear heat fire and disperse stagnated liver qi,lastly to moisten and invigorate lung.We received satisfactory curative effects both on the clinical treatments and experiments in DM and diseases accompanied by it when adopting the TCM methods of treatment based on the theory by Professor LI De-lin.

Objective To study the effect of the Riche-Cannieu anastomosis, an anomalous anastomosis between the deep branch of ulnar nerve and the recurrent branch of median nerve in the palm of the hand, on the diagnosis of patients with carpal tunnel syndrome. Methods Twelve patients (2 male, 10 female, mean age 49.8 years) with carpal tunnel syndrome with the presence of Riche-Cannieu anastomosis were tested in this study. Their clinical data including the symptoms and signs, electrophysiological findings and diagnosis were collected and discussed in relation to an anatomical review of these nerves. Results Due to the existence of the Riche-Cannieu anastomosis, the patients with carpal tunnel syndrome spared part or all of the function of the abductor pollicis brevis (APB) muscle. It may result in an unconformity of the patients′lesion degree caused by carpal tunnel syndrome and its clinical symptoms and electrophysiological findings. Conclusion Knowledge of the Riche-Cannieu anastomosis is of crucial importance in the clinical evaluation, diagnosis and treatment of carpal tunnel syndrome, as well as in avoiding errors in interpreting the electrophysiological data of the patients.

Objective;To investigate whether the traditional Chinese medicine Yisheng Injection can reduce the injury on isolated rat heart induced by cold ischemia. Methods ;18 isolated female closed colony SD rats were divided into two groups at random. In the control group, the isolated rat hearts were preserved in 0.9% sodium chlorine solution at 4℃ , while in the experimental group ,the hearts were preserved in 0.9% sodium chlorine solution containing Yisheng injection at 4℃. The specimens were harvested 2,6,12, 24,48 and 72 hours after preservation. Histochemical changes were observed in two groups. Results ;In the control group, the activities of alkaline phosphatase ( ALP) and NADPH diaphorase (NADPHD) decreased after cold ischemia for 24 hours. Lactic dehydro-genase (LDH) decreased slightly, while ALP and NADPHD decreased evidently, after cold ischemia for 48 hours. Succinic dehydro-genase ( SDH) and cytochrome oxidase ( CCO) decreased slightly, and NADPHD disappeared after cold ischemia for 72 hours respectively. In the experimental group, NADPHD, LDH and CCO decreased slightly after cold ischemia for 24, 48 and 72 hours. ALP decreased slightly after cold ischemia for 48 hours, however, SDH showed no changes, after cold ischemia for 72 hours. Conclusion : Cold ischemia can induce cold ischemia injury in isolated rat hearts, especially the vascular endothelial cells being more sensitive. Yisheng injection can reduce this kind of injury to some degree.

0.05),however,by the 12th week,there was a significant difference(3.6?0.48 vs 2.4?0.48,P0.05)in the graft vasculopathy between two groups.Conclusion: It is practicable to establish a cardiac chronic graft failure model between closed colony SD rats,and ischemia reperfusion injury,as an alloantigen independent factor,would accelerate the development of rat cardiac chronic graft vascular disease,which can save the time for establishing models.

Objective To investigate the clinical and electrophysiological characteristics of entrapment syndrome involving the median nerve's recurrent branch. Methods Fourteen cases (10 male, 4 female, mean age 34.7) of medial nerve recurrent branch entrapment syndrome were retrospectively analyzed in this study with regard to their clinical and electrophysiological data. Results Normal sensory conduction velocity ( SCV ) and normal amplitude of the sensory nerve action potential ( SNAP) were found, but there was a delay of the distal motor latency (DML) in all 14 cases. The compound muscle action potential (CMAP) amplitude was decreased in 11 cases, and normal motor conduction velocity (MCV) was demonstrated in 12 cases. Fibrillation potentials and/or positive sharp waves were detected in the abductor pollicis brevis muscle in 12 cases and in the opponens pollicis muscle in 14 cases. EMGs of the flexor digitorum superficialis and pronator quadratus muscle were normal in all 14 cases. Conclusion Electrophysiological examination could provide objective and reliable data for diagnosing and differentially diagnosing median nerve recurrent branch entrapment syndrome.

Objective To investigate the assessment value of serum Lp-PLA2 ,hs-CRP and D-D dimer on the severity of cor-onary artery lesion .Methods A total of 136 patients with coronary heart disease acute myocardial infarction (AMI) group 46 ca-ses ,stable angina pectoris (SAP) group 38 cases and unstable angina pectoris (UAP) group 52 cases from the inpatient of cardiolo-gy department in our hospital were selected as observation group ,and 135 healthy subjects were selected as control group ,serum Lp-PLA2 was detected by enzyme kinetic method ,Serum hs-CRP and plasma D-D dimer were detected by immune turbidimetric method ,of the severity of coronary artery lesion was evaluated by the Gensini integrations .Results Serum Lp-PLA2 ,hs-CRP ,D-D dimer level and Gensini score of observation group were significantly higher than those of control group ,above indexes of AMI group were higher than those of SAP group and UAP group ,UAP group were significantly higher than SAP group ,with statistical significance between the two groups(P<0 .05) .Serum Lp-PLA2 ,hs-CRP ,D-D dimer level and Gensini score of patients with heart functional class Ⅱwere significantly lower than those of Ⅲ and Ⅳ patients ,above indexes of patients with heart functional class Ⅲwere significantly lower than Ⅳ patients ,the difference between two groups was significant(P<0 .05) .Compared with severity of coronary artery lesion ,serum Lp-PLA2 hs-CRP and D-D dimer levels of mild coronary artery lesions were significantly lower than those of moderate and high coronary artery lesions ,and moderate coronary artery lesions was obviously lower than high coronary artery lesions (P<0 .05) ,Pearson correlation analysis also found that ,Lp-PLA2 ,hs-CRP ,D-D dimer levels had positive correlation with Gensini score (r=0 .642 ,0 .651 ,0 .608 ,P<0 .05) .Conclusion The assessment of serum Lp-PLA2 ,hs-CRP and D-D dimer level in the severity of coronary artery lesion has important significance .