Construction and characterization of △sseK2 mutant strain of Salmonella typhimurium SL1344 / 中华微生物学和免疫学杂志

Objective To analyze the biological characteristics of a mutant strain of Salmonella ty-phimurium SL1344 with sseK2-deletion (SL1344△sseK2) in order to provide reference for further study of safe and effective live vaccines. Methods The mutant strain SL1344△sseK2 with a deletion of 1047 bp in sseK2 gene was constructed through a two-step allelic exchange using recombinant suicide plasmid. Its com-plemented strain, SL1344C△sseK2, was also constructed. Biological and immunological characteristics of the mutant strain were detected. Results PCR, double-enzyme digestion and sequencing analysis showed that the mutant strain SL1344△sseK2 and the complemented strain SL1344C△sseK2 were successfully con-structed. The serotype of the mutant strain was 1,4,[5],12:i:1,2, identical to the parent strain SL1344. In addition, the mutant strain showed no significant change in biochemical characteristics or growth rate and was genetically stable in vitro. Compared with the parent strain SL1344, the virulence of SL1344△sseK2 was attenuated in BALB/ c mice. The median lethal dose of SL1344△sseK2 for 6-week-old BALB/ c mice was 3. 44×108 colony-forming units (CFU), which was 1620 times lower than that of SL1344. Oral immuniza-tion with SL1344△sseK2 protected 62. 5％ of the mice against challenge with wild Salmonella typhimurium strains on 17 d after vaccination. The levels of serum IgG antibody peaked on 14 d after immunization. No significant difference in biological characteristics was observed between the complemented and the parent strains, indicating that the mutant strain was basically complemented to the wild-type strain.Conclusions The mutant strain SL1344△sseK2 was constructed successfully and genetically stable with sig-nificantly attenuated virulence and good immunogenicity. This study suggested that sseK2 gene played an im-portant role in regulating the virulence of SL1344, which might provide reference for further study of its func-tion and for assessing its potential as a candidate live attenuated vaccine.

Taurine Transporter dEAAT2 is Required for Auditory Transduction in Drosophila / 神经科学通报·英文版

Drosophila dEAAT2, a member of the excitatory amino-acid transporter (EAAT) family, has been described as mediating the high-affinity transport of taurine, which is a free amino-acid abundant in both insects and mammals. However, the role of taurine and its transporter in hearing is not clear. Here, we report that dEAAT2 is required for the larval startle response to sound stimuli. dEAAT2 was found to be enriched in the distal region of chordotonal neurons where sound transduction occurs. The Ca imaging and electrophysiological results showed that disrupted dEAAT2 expression significantly reduced the response of chordotonal neurons to sound. More importantly, expressing dEAAT2 in the chordotonal neurons rescued these mutant phenotypes. Taken together, these findings indicate a critical role for Drosophila dEAAT2 in sound transduction by chordotonal neurons.

Analysis of unmet supportive care needs in patients with thyroid cancer during 13I treatment / 中华核医学与分子影像杂志

Objective To investigate the unmet needs of thyroid cancer patients undergoing 131Itreatment and their relationship with psychological distress and quality of life.Methods From October 2014 to March 2016,a total of 338 patients (106 males,232 females,average age (35.50±9.98) years) with thyroid cancer who were receiving 131I treatment were enrolled.The 34 items of Supportive Care Needs Survey Short Form (SNCS-SF34) were used to investigate the unmet care needs of patients.Patients' perceptions of distress and main causes of distress were measured by Distress Thermometers (DT) and issue list.The 12-item Short-Form Health Survey (SF-12) was used to measure patients' quality of life.Pearson correlation analysis was used to analyze the data.Results The degrees of patients' unmet care needs from high to low were as following:health information needs (65.38％,221/338),psychological needs (47.93％,162/338),patients' care and support needs (44.08％,149/338),physical and daily needs (36.09％,122/338) and sexual needs (34.32％,116/338).There were 47.93％(162/388) patients with DT score ＞ 4.The top three causes of patients' distress were fears,fatigue and memory deterioration.Degree of unmet needs for patients had positive correlation with the degree of psychological distress (r values:0.232-0.462,all P＜0.01).The unmet physiological and daily needs and psychological needs of patients had negative correlations with several dimensions of quality of life (r values:from-0.367 to-0.202,all P＜ 0.05).Conclusions The unmet care needs of patients with thyroid cancer have correlations with their psychological distress and quality of life.Evaluation of the unmet care needs of patients with thyroid cancer could be beneficial for providing the targeted care.

High-performance liquid chromatographic analysis of felotaxel, a novel anti-cancer drug, in rat plasma and in human plasma and urine.

A high-performance liquid chromatographic method was developed for the determination of a new derivative of docetaxel, felotaxel, in rat plasma and human plasma and urine. The separation of felotaxel was performed on a Dikma C18 column with 0.2% formic acid and acetonitrile (50:50) as the mobile phase. The flow rate was 0.8 mL/min and the column effluent was monitored by an ultraviolet detector set at 275 nm. The method was validated and found to be linear in the range of 5-1,000 ng/mL. The lower limit of quantification was 5 ng/mL based on 100 µL of plasma. The variations for intra-day and inter-day precision were less than 6.9%, and the accuracy values were between 87.3 and 107.4%. The extraction recoveries were more than 80.5%. These data confirm that the developed method has satisfactory sensitivity, accuracy and precision for the quantification of felotaxel in rat plasma and in human plasma and urine. The method was successfully applied to a pharmacokinetics study of felotaxel after intravenous doses of 5 mg/kg in rats.

Analysis of the Problem of Informed Consent in Clinical Trial / 中国医学伦理学

Objective:By observing the process of informed consent in clinical trials of one top three hospital, to disclose the non - standard phenomenon existingin the process of informed consent in clinical trials, and put for-ward that there should be special protection mechanism for vulnerable groups to participate in clinical trials. Meth-ods:Combined with the actual situation of one top three hospital, we comprehensively analyzed the problems exist-ing in the process of informed consent of clinical trials in our center, put forward strict solutions, and also formula-ted strict standards for the informed consent process of vulnerable groups. Results:Through the strict requirements of the informed consent process of vulnerable groups, the test process of vulnerable groups participating in the clini-cal trial was standardized; meanwhile the vulnerable groups were given special protection. Conclusion:Informed consent is an important guarantee that protects vulnerable groups to participate in clinical trials.

Effects of peer education on treatment among patients using 131I with thyroid cancer after surgery / 中华护理杂志

Objective To investigate the effects of peer education on treatment among patients using 131I with thyroid cancer after surgery.Methods A total of 80 patients using 131I therapy after surgery of thyroid can cerhospitalized from June 2014 to May 2016 were enrolled and randomly divided into two groups.The control group were given routine health care including hospitalized treatment,education,and follow-up examination after discharge from hospital,while the experimental group were given peer education,and the intervention was provided from admission to one month after discharge.Distress level,complication rates and knowledge mastering were compared between two groups.Results The differences of distress level,complication rates and knowledge mastering between two groups were statistically significant(P＜0.05).Conclusion Patients using 131I with thyroid cancer after surgery suffer from significant distress.Peer education can greatly reduce patients' distress,complications after 131I treatment,and improve mastering knowledge.

A qualitative study on the experience of thyroid cancer patients about diagnosis and post-surgical radioactive iodine treatment / 中国实用护理杂志

Objective To explore the experience of thyroid cancer patients about the diagnosis and post-surgical radioactive iodine treatment. Methods A qualitative, descriptive study was used. Thirty-eight patients with thyroid cancer who were undergoing post-surgical radioactive iodine treatment participated in this study. Dairies about the participants′ experience were collected from them. Interpretative phenomenological analysis was used. Results Three superordinate themes emerged: the psychological process from distress to acceptance; the experience about post-surgical radioactive iodine treatment; and personal growth related to the cancer. Conclusions Understanding the psychological process of patients with thyroid cancer after the diagnosis and their experience about post-surgical radioactive iodine treatment can contribute to the clinical staff to improve the care quality for this group.

Analysis of Standardization of Clinical Trial Contract Audit from the Perspective of Subject Protection / 中国医学伦理学

Carrying out clinical trial contract audit can audit the authenticity,legality and effectiveness of the contract and timely avoid the potential risks,clarify the responsibilities,rights and obligations of all parties participating in the clinical trials and clear the responsibility of each party in protecting the subjects,guarantee that the contracts include contents of subject protection and put them into practice.From the perspectives of establishing relevant documents,defining the main implementer,making the process,and confirming the core elements,this paper introduced how Xijing Hospital standardized clinical trial contract audit and expounded its significance in promoting the protection of subjects.

Comparative Study on the Preventive Use of 3 Kinds of Cephalosporins for 1 066 Patients with Breast Lesion Resection in Shaanxi Province / 中国药房

OBJECTIVE:To compare the effects of prophylactic application of 3 different antibiotics on prognosis in patients underwent breast lesion resection. METHODS:1 066 patients with breast lasion resection from 12 hospitals of Shaanxi province were divided into trial group(360 cases),control group A(352 cases)and control group B(354 cases)according to random num-ber table. Trial group was given first generation cephalosporin cefazolin;control group A was given second generation cephalospo-rin cefuroxime;control group B was given third generation cephalosporin cefoperazone sodium and tazobactam sodium. The dosage regimens of 3 groups were as follows:relevant drug 2 g added into 0.9%Sodium chloride injection 100 ml,ivgtt,0.5 h before sur-gery,medication course≤24 h after surgery in trial group. Those indexes of 3 groups were observed,such as post-operative ADR, incision healing,infection,hospitalization duration,phamaceutical costs per capita. RESULTS:There was no statistical signifi-cance in the rate of incision healing and the rate of post-operative infection among 3 groups(P>0.05). The incidence of post-opera-tive ADR,hospitalization duration and phamaceutical costs per capita in observation group were significantly lower or shorter than in control group A and B,with statistical significance(P<0.05). CONCLUSIONS:Cefazolin is better than cefuroxime and cefo-perazone sodium and tazobactam sodium to reduce the postoperative adverse reaction,antibiotics cost per capita and hospital drug cost per capita,shorten the hospitalization duration.

Construction and characterization of an attenuated recombinant Listeria monocytogenes vector vaccine delivering HPV16 E7 / 生物工程学报

Listeria monocytogenes (L. monocytogenes, LM) is an excellent tumor vaccine vector. In this study, recombinant LM vaccine candidate expressing human papillomavirus type 16 (HPV16) E7 protein was constructed and its charactericts were determined. Through homologous recombination, E7 gene was cloned in frame with the LM4 Phly promoter-signal sequence, and introduced into the chromosome of LM4. The recombinant strain named LM4△hly::E7 with the plasmid-free and antibiotic-resistant gene-free was constructed. LM4△hly::E7 could express and secrete E7-LLO fusion protein; its size is 66 kDa and has immunological activity. Furthermore, LM4△hly::E7 could multiply in RAW264.7 macrophages by confocal laser scanning microscope. Additionally, LM4△hly::E7 could induce specific antibodies against E7 in immunized mice in ELISA. Also, the 50% lethal dose (LD₅₀) of LM4△hly::E7 strain was 3.863×10⁹ CFU (Colony-Forming Units) in C57BL/6 mice with intraperitoneal immunization, which was more attenuated than wild type LM4. Mice immunized with LM4△hly::E7 did not show obvious pathological change. These data show that LM4△hly::E7 expressing E7-LLO fusion protein has good safety, which may provide the materials for research of antitumor effect and would be a promising vaccine candidate for cervical cancer.

Characterization of a Salmonella typhimurium SL1344 cya mutant strain / 中国免疫学杂志

Objective:To explore the function of the cya gene and the preliminary mechanism of attenuated strain.Methods:The biological characteristics of cya mutant in acid and alkali resistant,salt resistance,motility,biofilm components,poisonous to the cells of epithelial cell adhesion,invasion were analysis.Results:The mobility capabilities,acid and alkali resistance and salt tolerance of cya mutant were significantly lower than the parent strain;the composition testing revealed that the cya mutant did not produce cellulose,curli and biofilm;at the same time the adhesion and invasion to epithelial cells of cya mutant had a prominent depression,and the toxicity to HeLa cells was weaker than the parent strain.Conclusion:The function of cya gene is closely related to athletic ability, penetration of cell membrane, the formation biofilm and virulence.It will provide a theory reference to the functional research of Salmonella typhimurium cya gene and the mechanism of attenuated strain.This will contribute to the development of oral vaccine using attenuated Salmonella typhimurium as vector.

Construction and characterization of type III secretion system of attenuated Salmonella typhimurium / 生物工程学报

In order to develop a recombinant attenuated Salmonella typhimurium as oral live vaccine vector, we constructed recombinant plasmid pYA-sopENt100 by replacing the trc promoter with the sopE promoter and secretion signal sequence sopENt100 of Salmonella typhimurium on the basis of plasmid pYA3493. Then, the complementary plasmid pYA-sopENt100 was transformed into ΔcrpΔasdSL1344 by electroporation to generate attenuated Salmonella typhimurium type III secretion system ΔcrpΔasdSL1344 (pYA-sopENt100). We further characterized ΔcrpΔasdSL1344 (pYA-sopENt100). We also constructed a recombinant strain ΔcrpΔasdSL1344 (pYA-sopENt100-egfp) that harbored the reporter gene-enhanced green fluorescent protein (egfp) gene. Vero cells were infected with ΔcrpΔasdSL1344 (pYA-sopENt100-egfp) and the ability of delivery foreign antigens was tested via Western blotting analysis. The results of PCR, enzyme digestion and sequencing showed that the ΔcrpΔasdSL1344 (pYA-sopENt100) type III secretion system was constructed successfully. The serotype of ΔcrpΔasdSL1344 (pYA-sopENt100) was identical to ΔcrpΔasdSL1344 and SL1344. Compared with wild strain SL1344, the biochemical characteristics of ΔcrpΔasdSL1344 (pYA-sopENt100) had obvious change, but it was basically the same with ΔcrpΔasdSL1344. The growth speed was much slower than that of the wild strain SL1344. The chicken virulence test (LD₅₀) showed that the virulence of ΔcrpΔasdSL1344 (pYA-sopENt100) was 7×10⁴ times lower than SL1344. In addition, we observed the 37 kDa SopENt100-egfp protein in the cultured supernatant of ΔcrpΔasdSL1344 (pYA-sopENt100-egfp) strain by Western blotting analysis. However, both the 37 kDa SopENt100-egfp protein and 27 kDa EGFP protein were detected in ΔcrpΔasdSL1344 (pYA-sopENt100-egfp)-infected Vero cells. These results demonstrated that the recombinant Salmonella typhimurium type III secretion system ΔcrpΔasdSL1344 (pYA-sopENt100) was successfully constructed, and it should be used as a live vaccine vector for expressing foreign genes.

Construction and characterization of invasion protein B gene deleted mutant asd host-vector balanced lethal system / 中国免疫学杂志

Objective: To develop an oral live vaccine vector which stably carries exogenous genes.Methods:SL1344ΔsipBΔasd host-vector balanced lethal system was constructed by the method of recombinant suicide plasmid-mediated allelic exchange on the basis of attenuated Salmonella typhinurium SL1344ΔsipB.Then,the biological characteristics of SL1344ΔsipBΔasd was analyzed.Results:The results showed that the mutant was stabile with the Δasd gene in vitro;the serotype and growth rate of SL1344ΔsipBΔasd strain was almost same as the parent SL1344ΔsipB and SL1344 strain.And the mutant strains remain swim ming zones.Virulence test in mice showed that the virulence of SL1344ΔsipBΔasd which carried complementary plasmid pYA3493 by electro-transformation decreased by 1.4%compared with SL1344.Conclusion: These results showed that the SL1344ΔsipBΔasd mutant was successfully constructed.It is likely that this mutant should be used as a live vector to express foreign genes.

Efficacy of the emergency endoscopic sclerotherapy and tissue adhesives embolization of bleeding esophageal varices / 中国综合临床

Objective To investigate clinical value of the emergency endoscopic sclerotherapy and tissue adhesives embolization on patients with gastroesophageal varices bleeding. Methods A retrospective analysis of 52 cases of esophageal variceal bleeding in patients with clinical data was performed. Of which,22 patients were given conservative treatment alone and they were served as conservative treatment group. Another 30 patients were given the emergency endoscopic sclerotherapy and/ or tissue adhesive embolization besides the basis of conservative treatment,and they served as endoscopic treatment group. The data of hospitalization and mortality,rebleeding rates were recorded during hospitalized time and 1 year off hospitalization. Results The indices of hospitalized periods in endoscopic treatment group was( 16. 2 ± 1. 5)d,shorter than that in conservative treatment group((17. 6 ± 2. 1)d,P = 0. 53). The mortality and rebleeding rate in endoscopic treatment group in followed up periods were 3. 3% and 6. 6%,lower than those in conservative treatment group (9. 1%,13. 6%),and the comparison of relative risk RR of two groups of mortality and rebleeding rate were 2. 76 and 2. 06. Conclusion The therapy of emergency endoscopic sclerotherapy and tissue adhesives embolization of bleeding esophageal varices shows a good effect and it is worth recommending.

Social skills training intervention in popularization and application of children's behavioral problems at school / 中华行为医学与脑科学杂志

Objective To investigate the effects of social skills training on behavior problems of children aged 6-13 years.Methods 168 children with behavior problems were selected as the sample,who received social skill training by health education teachers for 12 weeks.The Rutter Parent and Teacher Scale and PHCSS(Children' s self-concept Scale) were used to evaluate the effect before and after intervention.Results After intervention,the total scores,A and N scores of the Rutter Parent Scale decreased significantly((5.42±6.23) vs (0.62± 1.77),(1.68±2.63) vs (0.50±2.74),(2.04±4.27) vs (0.36±0.65),P＜0.05).The total scores,A and N scores of the Rutter Teacher Scale decreased significantly((4.35±4.11) vs (0.62± 1.77),(1.56±3.65) vs (0.21 ±0.44),(1.32±2.48) vs (0.36±0.65),P＜0.05).The total score and different factor scores of PHCSS increased significantly(P＜0.05).The clinical effectiveness of intervention group was better than control group.Conclusion Social skill training by teacher is acceptable and helpful to improve behavior problem,psychosocial competence and selfconcept for children with behavior problems.

Pharmacokinetic Study on Single Dose and Multiple Dose of Triflusal Capsule in Healthy Volunteers / 中国药房

OBJECTIVE:To study the pharmacokinetic characteristics of triflusal capsule in healthy volunteers. METHODS:In ran-domized test,36 healthy volunteers were randomly divided into 3 groups. They were given low-dose,medium-dose and high-dose of Triflusal capsule(300 mg,600 mg and 900 mg),qd,for one day,and then pharmacokinetic study of single dose of Triflusal capsule was conducted;Triflusal capsule medium-dose group was continuously given medicine for 13 days,and then pharmacokinetic study of multiple dose of Triflusal capsule was conducted. The plasma concentration of triflusal was determined by LC-MS/MS,and Zorbax SB-C18 column was used with methanol-0.2% formic acid (80:20,V/V) at the flow rate of 0.2 ml/min. ESI was adopted in MRM mode,negative ion detection was carried out,quantitative analysis m/z 247.1→161.1(triflusal),m/z 294.0→250.0(internal standard, diclofenac sodium). Pharmacokinetic parameters were calculated by using WinNonlin 6.2 software,and the difference of them were compared. RESULTS:The linear range of triflusal were 0.05-20 μg/ml. The main pharmacokinetic parameters of triflusal capsules high-dose,medium-dose and low-dose groups were as follows:t1/2 were (0.45 ± 0.20),(0.47 ± 0.10),(0.43 ± 0.20) h;tmax were (0.56±0.20),(0.60±0.20),(0.47±0.40)h;cmax were(3.30±0.98),(10.65±3.26),(13.96±4.88)μg/ml;AUC0-8 h were(3.99±0.93), (13.29±1.72),(19.62±6.78)μg·h/ml;within dose of 300-900 mg,linear relationship was found between cmax,AUC0-8 h and dose(R2=0.954,0.986). When reaching stable state of multiple dose,average blood concentration was(0.71±0.20)μg/ml;main pharmacokinetic parameters were as follows:AUCs(17.10±4.82)μg·h/ml,t1/2(0.49±0.10)h,tmax(0.85±0.62)h,cmax(11.58±3.99)μg/ml,AUC0-8 h (16.99±4.84)μg·h/ml,AUC0-∞(17.08±4.81)μg·h/ml;accumulation factor(1.28±0.40). tmax and t1/2 of single dose were similar to those of multiple dose. CONCLUSIONS:LC-MS/MS can determine the content of triflusal in human plasma rapidly and accurately, and accumulation phenomena exist in healthy Chinese volunteers,which shows linear pharmacokinetic characteristics.

Analysis of the curative effect and safety of estrogen as an adjunctive treatment of senile vaginitis / 中华老年医学杂志

Objective To analyze the curative effect and safety of the estrogen as an adjunctive treatment of senile vaginitis.Methods 100 patients with senile vaginitis in our hospital from June 2011 to September 2012 were divided into observation group (n =50) and control group (n =50).Observation group was given estrogen plus metronidazole and control group was given metronidazole only.Results 45 cases were cured and 4 cases had effective treatment in observation group,and 21 cases were cured and 17 cases had effective treatment in control group.The efficacy rate was 98.0％ (49 cases)in observation group versus 76.0％(38 cases) in control group (x2=11.50,P＜0.01).After the treatment,the recurrence rate was 6.1％ (3 cases)and 26.3 ％ (10 cases)in observation and control groups respectively (x2 =10.66,P＜0.05).After treatment,vaginal health scores,symptom scores of vaginitis,and vaginal PH were significantly higher in the observation group than in the control group (all P＜0.05).In addition,the improvements of signs and symptoms were significantly better in the observation group than in the control group (all P＜0.05).However,there was no significant difference in adverse reaction rate between the two groups (P＞ 0.05).Conclusions Estrogen as an adjunctive treatment of senile vaginitis has better curative effect and safety.It is worthy of generalization in clinical medicine.

Diagonsis establishment of fluorescen quantitative PCR assay for pseudorabies wild-type virus and vaccine virus / 生物工程学报

We designed two pairs of primers and their corresponding TaqMan probes according to gH, gE gene of PRV. By optimizing the probe's concentration, Mg2+ concentration, primers concentration and sample DNA extraction, real-time fluorescent quantitative PCR (FQ-PCR) which can quickly identity field virus and vaccine virus of PRV was established. According to our results, the dynamic range of the FQ-PCR assay is between 10 x 10(1) copies/microL and 10 x l0(8) copies/microL, and the detection limit of FQ-PCR is 1.0 x 10(1) copies/microL, which is 100 fold higher than that of conventional PCR. We detected 60 doubtful tissue samples using the FQ-PCR assay, serum neutralization and conventional PCR. In conclusion, the FQ-PCR method is rapid, sensitive, specific and accurate, and can be used to detect field strains of PRV rapidly. The closed-tube format of the assay minimized the risk of contamination of subsequent reaction and the assay can be performed in 2 h or less. Development of real-time quantitative PCR provides the basis for the early and rapid detection and analyzing quantitatively the infectious degree of PRV.

Assessment on Bioequivalence of Tegaserod Maleate Dispersible Tablets in Human Body / 中国药房

OBJECTIVE: To study the bioequivalence of tegaserod maleate dispersible tablets in healthy volunteers.METHODS: A single oral dose of 6mg test or reference preparations of tegaserod maleate was given to 22 healthy volunteers in a randomized crossover study.The plasma concentrations of tegaserod were determined by LC/MS/MS assay.RESULTS: The main pharmacokinetic parameters of test and reference products were as follows: tmax(0.86? 0.22) and(1. 01? 0.24) h;Cmax(2.21? 0.69) and(2.05? 0.64) ng? mL1;AUC0～ 17(6.35? 2.48) and(6.47? 1.99) ng? h? mL-1,AUC0～ ∞(6.69? 2.59) and(6.70? 2.03) ng? h? mL-1,respectively.The relative bioavailability of test to reference preparation was(98.2? 22.1) %.CONCLUSION: The reference preparation and the test preparation are bioequivalent.