The E. coli BL21(DE3) strain bearing the plasmid(T_7lac/His6-tag) with the h?TNF?recombinant gene was grown in LB medium containing SO?g/ml kanamycin, at 37 ℃ , up to the late logarithmic phase(OD590 ~ 0.5)then induced with IPTG (final concentration lmmol/L)for 5 hours. SDS-PAGE analysis revealed that expression level of the product(His6-?TNF?was up to 45% of the total bacterial proteins and was mainly as insoluble inclusion bodies ( IBs) . After cell lysis, the IBs was separated by centrifugation, dissolved in 7mol/L urea, then purified by Ni column ( Ni~(2+) -Sepharose 6B) . And the purity of more than 95% and the recovery of about 90% were obtained. The purified product was refolded with the renaturation buffer under low temperafure(

Objective To ascertain the pressure ulcer prevalence among secondary and tertiary general hospitals in different areas of Guangdong Province, and describe pressure ulcer related nursing measures in hospitalized patients. Methods A cross-sectional descriptive design was used in the present study. 5 secondary and tertiary general hospitals including 25164 hospitalized patients from different regions in Guangdong Province were recruited. There were 267 pressure ulcer patients among them. Minimum Data Set (MDS) was used to measure prevention measures of hospitalized patients. Pressure ulcer patients′questionnaire was used to measure medical treatment of pressure ulcer patients. Results The overall prevalence rates of pressure ulcers in the 25 hospitals range from 0 to 3.49%, with an average of 1.26%. The prevalence rate in the secondary general hospitals was 1.8%(41/2266), which was significant higher than that of in the tertiary general hospitals(1.0%, 226/22998), χ2=13.48, P<0.01. For the hospitalized patients at risk, 42.0%(1408/3353) of patients didn′t use any systemic support surface, 45.8%(1537/3353) of patients didn′t use any local support surface. In the pressure ulcer patients, 35.2%(94/267) of patients didn′t use any systemic support surface, 14.2% (38/267)of patients didn′t use any local support surface, and 61.4%(164/267) of pressure ulcer patients didn′t use any skin care products. Conclusion The prevention measures of pressure ulcer in Guangdong province were not effective, further improvements are still required.

Brown-Sequard Syndrome ; etiology ; Cervical Vertebrae ; Humans ; Intervertebral Disc Displacement ; complications ; surgery ; Male ; Middle Aged

Objective To investigate the distribution of 16S rRNA methylase genes in Klebsiella pneumoniae strains producing Klebsiella pneumoniae ealbapenenase type 2(KPC-2).Methods A total of 37 Klebsiella pneumoniae isolates producing KPC-2 were collected.The minimal inhibitory concentrations (MICs)of these strains to amikacin,gentamyein and netilmicin were determinated by agal dilution method.Six 16S rRNA methylase genes(armA,rmtA,rmtB,rmtC,rmtD and npmA)were detected by PCR.Results The resistant rates to amikacin,gentamycin and netilmicin were 97.3%(MIC50≥1024μg/mL).Among those resistant strains,8 were armr/A positive,25 were rmtB positive,4 were both armA and rmtB positive.and no other 16S rRNA methylase genes were found.The total positive rate of 16S rRNA methylase genes was 78.4%(29/37).Conclusion 16S rRNA methylase genes armA and rmtB ale prevalent in Klebsiella pneumoniae strains producing KPC-2.

OBJECTIVE: To develop a rapid DNA detection kit for DNA extraction and PCR identification of Panax ginseng C. A. Mey. METHODS: The classical DNA extraction and PCR identification methods for Panax ginseng C. A. Mey were modified, and the compositions and reaction conditions of the kit were determined. In addition, the specificity, stability, sensitivity, and repeatability of the kit were evaluated. The genomic DNAs of genuine and counterfeit ginseng goods were extracted by the kit and PCR was performed to identify the authenticity. The purity of the extracted DNA was detected by UV spectrophotometry. Finally, commercially available ginseng samples were verified. RESULTS: The purity of the genomic DNA extracted by the kit was (1.73 ± 0.13) (OD260/OD280), and a fragment between 150 and 200 bp could be amplified only from authentic Panax ginseng C. A. Mey. The specificity of the kit was 100%. The repetitive experiments showed that the average intra-assay CV% and inter-assay CV% of the kit were 2.38% and 2. 62%, respectively. The DNA in solutions diluted by 200 times could still be detected. Stability experiment proved that repeated freeze-thawing for 20 times had no significant effect on the activity of this kit and the test sample could be stored at - 20℃ for one year. The specificity test confirmed that 8 samples among the 10 commercial products were genuine, and 2 were counterfeit. CONCLUSION: The nucleic acid extraction and purity of the DNA detection kit can meet the requirement for identification of Panax ginseng C. A. Mey. The kit has good specificity, high sensitivity, and good stability, so it is suitable for the rapid detection of Panax ginseng C. A. Mey.

Objective To establish the quality control standard for Shuanggen Qingnao Granules.Methods TLC was used to identify Yiyiren SEMENCOICIS,Yujin RADIXCURCUMAE,Banlangen RADIX ISATIDIS. HPLC was used to determine the content of Puerarin in Shuanggen Qingnao Granules.Results The TLC spots developed were fairly clear,and the bland test showed no interference.Puerarin showed a good linear relationship in the range of 5.184～40.50?g/mL,the average recovery of Puerarin was 99.93%,and RSD=1.81%.Conclusion The method is accurate and quick,and can be used for the quality control of Shuanggen Qingnao Granules.

Objective To investigate precipitants of epileptic seizure, and to explore the correlation between various precipitants and relationship between precipitants and clinical features of epilepsy.Methods Data were collected from 154 patients attending a tertiary-care epilepsy clinic of Nanjing Brain Hospital between April 2015 and April 2016.The patients with epilepsy were older than 16 years, had a clinical history of one year or more, and one seizure at least a year and one seizure at least in the latest three months.An enclosed questionnaire was combined with open interview to identify and characterize seizure precipitants and clinical characteristics of patients.Patients were asked respectively whether there were some precipitants three months before and during last three months.Correlation between seizure precipitants and relationship between precipitants and clinical characteristics, such as age, gender, course, seizure frequency and so on, were calculated.Results A total of 125 (81.2%) participants reported at least one precipitant.Common precipitants (in descending order) were as follows: emotional stress (56.0%), sleep disorder (38.4%), fatigue (27.2%), missed medication (20.0%).There were one to six different precipitants for one patient, and 60.8% of patients had two or more precipitants.There was a correlation between emotional stress and sleep disorders as well as fatigue (χ2=4.665, 8.668;P<0.05).Patients with idiopathic epilepsy were more sensitive to sleep disorders.There was no relationship between total precipitants and clinical features such as age, gender, age of onset, duration, type of seizure, seizure frequency, number of drug taking and so on.Conclusions Seizure precipitants were found widespread.The most common precipitants were found to be emotional stress, sleep disorders, fatigue and missed medication.There existed a correlation between emotional stress and sleep disorders as well as fatigue.There was no connection between total precipitants and patient′s demographic characteristics as well as clinical features.However, the type of seizure precipitants was different in patients with different demographic and clinical characteristics.

Objective Through the analysis of cases,to improve the cognition of clinicians on extracranial vas-cular embolism diseases(VED)in children.Methods The clinical information included incidence trend,thrombophi-lia,clinical features,treatment and prognosis of 29 children with VED,who were admitted from January 201 1 to Decem-ber 201 5 in Nanjing Children′s Hospital Affiliated to Nanjing Medical University,and retrospectively analyzed.Results The annual rate of VED increased from 0 to 1 .49 per 1 0 000 hospital admissions from January 201 1 to December 201 5.In 29 cases,there were 20 male and 9 female,with an average age of 6.03 years old (2 h -1 3.83 years old).In-fants accounted for >20%(6 /29 cases).The average age of the female,non infected group,rheumatic disease group and venous thrombosis group was higher than that of the male,the infection group,the non rheumatic disease group and the arterial embolization group,but there was no significant difference(all P >0.05).Thrombophilia were as follows:in-fection disease,heart disease,kidney disease syndrome,rheumatoid disease,tumor,and twins embolization syndrome. Risk factors included central venous line,immobility,dehydration,glucocorticoids use,etc.Heart disease was the leading cause of non thrombotic embolism,while other thrombophilic diseases resulted in thrombotic embolism.Pulmonary em-bolism was mainly seen in patients with severe pneumonia,especially mycoplasma pneumoniae pneumonia (MPP ).The older the age,the more typical clinical manifestations,and significantly elevated D dimer,fibrin/fibrinogen degradation products tips VED.Treatment depends on the degree of risk.Thirty -seven point five percent(3 /8 cases)of children with arterial embolism were less than 1 year old,and the mortality 25.00% (2 /8 cases)was significantly higher than that of venous thrombosis 1 7.65%(3 /1 7 cases)and 1 1 .76% (2 /1 7 cases).Conclusion Heart disease mainly cause arterial embolism and infection,while nephritic syndrome and rheumatic disease mainly cause venous embolism.Pulmona-ry embolism is mainly seen in children with severe pneumonia,especially MPP.The risk of arterial embolization is higher than that of venous embolism.

Objective To analyze the effect of N-acetyl-L-cysteine(NAC)on endoplasmic reticulum stress mediated HepG2 cells apoptosis and evaluate the role of NAC in the treatment of liver injury.Methods HepG2 cells were treated with thapsigargin(TG)to establish the model of oxidative endoplasmic reticulum stress mediated apoptosis,and NAC was used to intervene in apoptosis.To evaluate the apoptosis,various methods such as MTT assay,flow cytometry,DNA ladder and Western blot were performed.Results After treated with 2 μmol/L TG for 0,24,36 and 48 hours,the vitality of HepG2 cells decreased.The ratio of apoptotic cells increased along with the prolonged treatment duration of TG(0.7%±0.5%,27.6%±6.3%,29.7%±3.3%,47.9%±3.5% respectively,P<0.05),and the production of reactive oxygen species(ROS)also increased in time-dependent manner(14.0%±0.5%,36.1%±3.0%,38.2%±6.0%,48.3%±12.4%,P<0.05).The HepG2 cells showed typical morphologic change of endoplasmic retieulum stress induced by 2 μmol/L TG after 36 h and 48 h.DNA ladder was observed at the same concentration and time point correspondingly.Endoplasmic reticulum stress mediated-apoptosis was confirmed by Western blot.Both 10 mmol/L and 20 mmol/L NAC could protect ceils from apoptosis.The ratio of apoptotic cells decreased to 14.0%±1.3% and 11.0%±0.3%,respectively.The production of ROS decreased to 34.7%±0.8% and 31.5%±2.9%,respectively.The effect was related to the concentration of NAC.Conclusions As a Ca2+-adenosine triphoshatase inhibitor,TG may disrupt intracellular calcium homeostasis,which can induce endoplasmie reticulum stress and apoptosis.NAC,the precursor of the synthesis of-SH,can directly inhibit the ROS reaction and alleviate liver damage,which may play a role in the treatment of liver failure.

Objective To investigate the molecular epidemiology of carbapenem resistant Acinetobacter baumannii from Zhejiang Province using multilocus sequence typing.Methods Three hundred and two imipenem or meropenem resistant A.baumannii isolates were collected from eleven tertiary hospitals of eleven different regions of Zhejiang Province in 2009 and 2010.Multilocus sequence typing was used for molecular typing.eBURST was used to analyze the results of multilocus sequence typing.PCR was used to detect the OXA-type carbapenemase genes.Results Seventeen sequence types (STs) were identified from three hundred and two A.baumannii isolates.eBURST analysis revealed eleven STs belonged to clonal complex 92 (CC92) and corresponded to European clone Ⅱ lineage.CC92 was the predominant carbapenem resistant A.baumannii clone of Zhejiang Province,which accounted for 94.4％ (285/302) of all isolates and distributed in all 11 hospitals.blaOXA-23 gene was identified in 97.4％ (294/302) of all isolates.Conclusions Carbapenem resistant A.baumannii CC92 were clonal disseminated among multiple hospitals of Zhejiang Province.blaOXA-23 was the most popular carbapenemase gene in carbapenem resistant A.baumannii.(Chin J Lab Med,2013,36:303-307)