ObjectiveTo explore the purification effect of hepatitis A virus(HAV)with a new type of composite medium Capto Core 400 chromatography packing.MethodsHAV crude liquid cultured with human embryonic lung diploid cells(2BS strain)was purified using Capto Core 400 chromatography media. The optimum conditions for purification of HAV were determined from flow rates(60,150,300 and 450 cm/h),pH of loading buffer(6. 6,7. 0 and 7. 4),salt ion concentration of loading buffer(0,0. 15,0. 3 and 0. 45 mol/L)and capacity[0. 5,1,2 and 4 column volume(CV)],respectively. The purification effect was compared with that of traditional Sepharose 4FF chromatography media.Results The optimum purification conditions of Capto Core 400 were preliminarily determined as follows:flow rate of 150 cm/h,loading buffer pH of 7. 0 ～ 7. 4,salt ion concentration of 0. 15 mol/L,loading volume of 2 CV. The average recovery rate of virus was 92. 61%,and the average removal rate of protein was 84. 25%. The average recovery rate of HAV antigen purified by Sepharose 4FF chromatography media was 76. 03% and the average protein removal rate was 73. 21%. The purification effect of Capto Core 400 chromatography media was better than that of Sepharose 4FF chromatography media.ConclusionThe Capto Core 400 composite chromatography media had a high recovery rate for HAV purification and a relatively simple operation,which was suitable for HAV purification.

BACKGROUND: To tonify qi, benefit spleen and primary qi, strengthen tendons and bones and promote circulations in meridians and collaterals are the classical principles in Chinese medicine for anti-kinetic fatigue.OBJECTIVE: To observe the efficacy of compound Chinese herbal medicine on anti-kinetic fatigue.DESIGN: Randomized controlled experiment based on animals.SETTINGS: Animal Experimental Center of Wenzhou Normal University and Zhejiang College of Traditional Chinese Medicine.MATERIALS: The experiment was performed in Animal Experimental Center of Zhejiang College of Traditional Chinese Medicine from February to June 2001, in which, 80 ICR mice of either sex, body weighted(22±2) g and 40 SD male rats, body weighted(194±7) g were employed.ETHODS: ① Eighty mice were randomized into two groups, of which, 40 mice was gastric-infused with Chinese herbal fiang li fang(experimental group) and 40 mice was with physiological saline (the control) . After bred for 15 days, they were randomized into 3 groups for anti-fatigue experiment ( n = 15, swimming time), anti-hypoxia ability( n = 15) and determination of hemoglobin(n=10). ② Determination of testosterone: Forty rats were randomized into 4 groups, named the control with physiological saline infusion, herbal infusion group, exercise group, exercise-herbal infusion group. In herbal infusion group, the rats were infused once a day. In exercise group and exercise-herbal infusion group, the rats exercised once a day, swimming till being exhausted at the tolerance of 3% body weight at 28 ℃ of water temperature. After bred for 20 days, heart blood 2 mL was collected with chest opened and the serum was centrifuged and placed in refrigerator at 4 ℃. Minigamma 1275γ counter and VCS method were applied for determina tion.MAIN OUTCOME MEASURES: ① Determination of swimming time of mice in every group. ③ Time of anti-hypoxia. ③ Levels of hemoglobin and testosterone in rats.RESULTS: Forty rats and 80 mice were all entered the result analysis. ①Observation of swimming time in mice: The time was remarkably prolonged in experimental group compared with the control[ (125.2±13.7), (108.9 ±12.6) minutes, P＜0.01] . ② Anti-hypoxia ability: It was much improved in experimental group compared with the control[(25.0±l.5),(23.1 ±1.8) minutes, P ＜0.05] . ③ Level of hemoglobin: The level of hemoglobin in experimental group was higher than that in the control, but the difference was not significant[(162.1 ±5.2), (159.3 ±5.3) g/L] . ④Level of testosterone: That in exercise group of rats was remarkably lower than exercise-herbal infusion group[(4.69 ±0.92), (6.03 ±0.77) nmol/L,P ＜ 0.05 ] . The difference in testosterone level was not significant between the control with infusion of physiological saline and herbal infusion group [(6.59 ±0.91), (6.98 ±0. 88) nmol/L, P ＞ 0.05].CONCLUSION: Under fatigue state, fian lifang can prolong the time of swimming in mice, improve work time and efficacy, increase hemoglobin content and benefit anti-hypoxia ability, which indicates rather strong anti-fatigue action. The combination of Chinese herbal jian lifang and exercise training can much' significantly increase serum testosterone level in rats compared with exercise group, which explains that fian li fang can increase testosterone level in rats, prevent testosterone decrease induced by exhausted exercise and accelerate the recovery of physical function.

Objective To contrast clinical effect of normal frequency mechanical ventilation vs .nasal continuous positive air‐way pressure ventilation in the treatment of neonatal respiratory failure .Methods Sixty children diagnosed as neonatal respiratory failure were enrolled in our hospital from January 2013 to October 2014 ,and the children were randomly divided into observation group (n=31) and control group (n=29) .Observation group were given nasal continuous positive airway pressure ,children with the control group received normal frequency mechanical ventilation .The X‐ray of the two groups before treatment and 2 d after treatment were observed and scored ,the 3 d survival rate ,blood gas analysis index ,ventilator associated pneumonia ,time of opera‐tion ,and incidence of pulmonary hemorrhage were observed in the two groups .Results The X‐ray results showed that :after treat‐ment ,the two groups of children with X‐ray film scores were gradually decreased ,compared with before treatment ,the difference was statistically significant (P<0 .05);the observation group decreased significantly after treatment 24 h compared with the control group ,the difference was statistically significant (P<0 .05);the survival rate of 3 d in the observation group(93 .50% ) was signifi‐cantly higher than that in the control group(82 .76% ) ,the difference was statistically significant (P<0 .05) .The pH in observation group was less than the control group ,PO2 was greater than the control group ,the difference was statistically significant (P<0 . 05) ,the PCO2 in observation group was less than the control group ,but the difference was not statistically significant (P>0 .05) . Time ,incidence of pulmonary hemorrhage and incidence of ventilator associated pneumonia in observation group were less than those in control group ,the difference was not statistically significant (P>0 .05) .Conclusion The normal frequency of mechanical ventilation and nasal continuous positive airway pressure can be effective reducing the relevance of respiratory pneumonia in the treatment of neonatal respiratory failure ,short‐term effect of nasal continuous positive airway pressure is slightly better than normal frequency mechanical ventilation .

Objective To analysis the effect of vitamin, folic acid combined with Ginkgo biloba extract on cognitive function in elderly patients with stroke.Methods 56 patients who were diagnosed with acute cerebral infarction were collected.All patients were randomly divided into experimental group and control group, 28 cases in each group.Control group was given conventional treatment for improving circulation, nutrition brain cells, anti platelet aggregation, experimental group was given vitamin B6 tablets(20 mg,one time per day), folic acid tablets(20 mg, one time per day) and Ginkgo Biloba Extract Tablets(80mg, two times per day) on the basis of conventional treatment.After treatment, the levels of plasma Hcy, serum BDNF, UCP2 level and cognitive function were detected in all patients.ResuIts After treatment, compared with control group, the plasma Hcy level was significantly lower in experimental group(P<0.05).The serum BDNF level in experimental group was significantly higher (P<0.05) .The serum UCP2 level in experimental group was significantly higher (P<0.05) .The MMSE and MoCA scores of the patients in the experimental group were significantly higher (P<0.05).ConcIusions Vitamin, folic acid combined with Ginkgo biloba extract can reduce the level of plasma Hcy in elderly patients with stroke, improve serum BDNF, UCP2 levels, improve the cognitive function.

Objective To analyze the clinical,demographic and pathologic features of symmetric acral keratoderma.Methods Totally,62 outpatients with symmetric acral keratoderma were collected at the Dongguan Hospital for Chronic Disease Prevention and Control from May 2003 to June 2012.Data on these patients,including demographic information,family history,physical and pathological examination findings,were reviewed.Results There were 55 males and 7 females among these patients.The age at onset and disease duration varied from 4 to 53 years (average: 24.02 years) and from 15 days to 10 years (average: 26.65 years) respectively.Characterized manifestation was brown keratinized patches on the dorsum of hands and digits,wrists,elbows,knees,ankles,and along the lateral margin of palms,which became white 3 to 6 minutes after immersing in water.Lesions usually resolved spontaneously in winter.Microscopic examination of lesions for fungal elements was negative.Histopathology revealed epidermal hyperkeratosis,acanthosis and mild papillomatous hyperplasia as well as dermal infiltrates with a few lymphocytes.Conclusion Symmetric acral keratoderma is a skin disease characterized by symmetric acral keratosis with apparent seasonality.

The assessment of right heart function plays an important role in diagnoses, therapeutic decision-making, curative effects and prognoses of many congenital and acquired heart diseases. Recently, a lot of new techniques of echocardiography have been used to evaluate right heart function, such as DTI, MPI, 3-DE, AQ and CUS, which all have their own advantages and disadvantages.

Objective To explore the application effect of critical thinking applied in the intravenous infusion.Methods To give nurses who worked in the Neurosurgery Department from January to April 2012 training lessons of critical thinking so that they could analyze,judge,formulate and implement corresponding nursing measures during the intravenous infusion process through their critical thinking.Then the successful rate of venepuncture for patients and the patients' satisfaction degree with nursing before and after the critical thinking training lessons were observed.Results The successful rate of venepuncture for patients and the patients' satisfaction degree of nursing were apparently improved after the nurses received critical thinking training.The difference was of statistical significance.Conclusions After the implementation of critical thinking training during the intravenous infusion process for nurses,nurses puncture skills is enhanced,infusion swelling and complications are reduced,and the quality of clinical nursing care and patients' satisfaction degree of nursing is improved.

In recent years, with the continuous increasing incidence and mortality of lung cancer, traditional Chinese medicine (TCM) researches have made some progress. This article reviewed the research progress in the relationship between TCM syndromes and modern medical staging of lung cancer, with a purpose to provide references for the follow-up related work.

Objective To culture the acute leukemia cells in vitro,and to prepare cancer vaccine expressing heat shock protein 70 (HSP70) of Bacille Calemette-Geérin(BCG) onto the cell surface,so as to study its anti-tumor effect and mechanism.Methods Acute myeloid leukemia (AML) cells were cultured in a serum-free Stemspan(H) culture supplemented with cytokines [stem cell factor(SCF),flt-3 ligand (FL),interleukin (IL)-3 and IL-6] in vitro.And B-lineage acute lymphoblastic leukemia (B-ALL) cells were cultured in a Iscove modified medium(IMDM) culture supplemented with cytokines (SCF,FL,IL-3 and IL-7) in vitro.Cellular morphology was observed by the microscopy and immunophenotype determination was used to verify the biological characteristics of acute leukemia cells after culture.Lipofectamine 2000 was used to transfect the pDisplay-HSP70 plasmid into acute leukemia cells.The expression of HSP70 on the cell surface was detected by fluorescene microscope.Then the immunogenicity of the leukemia cells expressing HSP70 were detected.The experimental groups were divided into 3 subgroups:the wide-type acute leukemia cells (wt-LC group),the pDisplay-leukemia cells (pDisplay-LC group),and the pDisplay-HSP70-leukemia cells (HSP70-LC group),respectively.The leukemia cells in different groups were cultured with autologous peripheral blood T cells for 72 hours.The proliferation indices of T cells were assayed by carboxyfluorescein diacetate succinimidyl ester (CFSE)-staining method,and the contents of interferon-γ(IFN-γ) were tested by enzyme-linked immunosorbent assay (ELISA).The leukemia cells in different groups were cultured with autologous peripheral blood T cells,and after 6 days,the fresh acute leukemia cells were added [in the different ratios of cytotoxicity T lymphocyte (CTL):leukemia cells were 10 ∶ 1,20 ∶ 1,40 ∶ 1 and 80 ∶ 1] and continued to be cultured for another 12 hours.Cytotoxicity assay was measured by lactate dehydrogenase (LDH) release.Results After short term culture in vitro,the leukemia cells were in colony-like suspension and maintain the proliferation characteristics were maintained.The cell proliferation was rapidly cultured for about 10 days and then was gradually slowed down.But there was no difference between the day 10 and day 0 in the expressions of CD13 and CD33 in fifteen cases of AML cells (P ＞ 0.05).Equally,there was no difference between the day 10 and day 0 in the expressions of CD19,CD10 and CD22 in fifteen cases of B-ALL cells (P ＞ 0.05).After BCG HSP70 gene transfection,the yellow-green fluorescence on the leukemia cells surface was observed under the confocal microscope.Detection of the immunogenicity:(1) Autologous T cell proliferation:the most significant T cell proliferation was observed in the group of HSP70-transfected leukemia cells (t =17.89,19.58,all P ＜0.05).There was no difference between the wt-LC group and pDisplay-LC group (P ＞ 0.05).(2) The contents of cytokines:the IFN-γ level in the group of HSP70-transfected leukemia cells was higher than those of wide-type acute leukemia cells and the pDisplay-transfected ones (t =24.72,24.81,all P ＜ 0.05).(3) Cytotoicity of CTL:the killing rate in HSP70-transfected leukemia cells was significantly higher than those of wide-type acute leukemia cells and pDisplay——transfected ones(F =13.66,P ＜ 0.05).And with the increase of the ratio from 10 ∶ 1 to 80 ∶ 1,the inhibiting activity of CTL in the HSP70-LC group was raising(F =19.69,P ＜ 0.05).Conclusions Fresh acute leukemia cells can be successfully cultured in vitro.Short-term culture can significantly increase the number of leukemia cells,but has little effect on surface antigen expression.So,the biological characteristics of the leukemia cells can be maintained.The leukemia cells vaccine expressing BCG HSP70 onto its surface was successfully prepared,and gene transfection of BCG HSP70 can significantly enhance the immunogenicity of leukemia cells.

Objectives To prepare the HL-60 cell vaccine expressing heat shock protein 70 (HSP70) of Bacille calmette-Guérin (BCG), so as to study its anti-tumor effect and mechanism. Methods The whole BCG HSP70 gene was amplified from BCG genome by polymerase chain reaction (PCR) and sub-cloned into the polyclone endonuclease sites in pDisplay. The recom-binant vector of pDisplay-HSP70 was verified by sequencing. Then the HL-60 cell vaccine expressing the protein onto the cell surface was prepared by lipofectamine transfection. To detect the immunogenicity of HL-60 cells expressing HSP70, the test groups were divided into three subgroups, HL60-wt, HL60-pDisplay, and HL60-HSP70 respectively. Each group was cultured with peripheral blood T cells for 72 h, then the proliferation indices of T cells were assayed by CFSE-staining method, and IFN-γwere tested by enzyme-linked immunosorbent assay (ELISA). The HL-60 cells of different groups were cultured with peripher-al blood T cells for 6d. The wild-type HL-60 cells were added and co-cultured for another 12h. Cytotoxicity assay was measured by LDH release. Results (1) The fragment of BCG HSP70 was consistent with the theoretical value. DNA sequencing showed that the recombinant vector of pDisplay-HSP70 was correctly constructed. (2) BCG HSP70 expressed onto the HL-60 cells sur-face. (3) Detection of the immunogenicity: ①The most significant T cell proliferation was observed in the group of HSP70-transfected HL-60 cells (P<0.05). There was no difference between the HL60-wt group and HL60-pDisplay group (P>0.05).②The contents of IFN-γof the HSP70-HL60 group was the highest.③The inhibiting activity of CTLs on HL-60 cells in the group of HSP70-transfected HL-60 cells was more significant than that of wide-type and pDisplay--transfected HL-60 cells. And with the increase of the E:T ratio, the inhibiting activity of CTLs in the HSP70-HL60 group was rising. Conclusions The recombinant eukaryotic expression vector (pDisplay-HSP70) of BCG HSP70 was successfully constructed. And the HL-60 cell vaccine expressing BCG HSP70 onto its surface was successfully prepared. The results showed that gene transfection of BCG HSP70 could significantly enhance the immunogenicity of HL-60 cells.