Objective To construct a lentiviral vector carrying angiopoietin-1 (Ang-1) and DsRed gene, and to package a virus particles. Methods The Ang-1 lentiviral vector with DsRed (PLV.Ex3d.P/puro-CMV>Ang-1>IRES/DsRed-Express2) was constructed by Gateway technology, and identiifed by PCR and gene sequencing. The lentiviral vector was mixed with helper vector pLV/helper-SL3, pLV/helper-SL4 and pLV/helper-SL5 by Lipofectamine 2000 to prepare DNA-Lipofectamine?2000 complexes. The complexes were then added to transfect 293FT cells and package virus. The virus titers and infection ef-ifciency were determined by lfuorescence expression. Results Ang-1 lentiviral vector PLV.Ex3d.P/puro-CMV>Ang-1>IRES/DsRed-Express2 was constructed successfully as identified by PCR and gene sequencing. Lentivirus with high-efficiency infection was produced by transfection to 293FT cells and the virus titer was 5×108 TU/ml. Conclusions The recombinant len-tiviral vector for Ang-1 was successfully constructed by Gateway technology and the lentivirus with high-efifciency infection packaging can be used for further experiment of Ang-1 gene.

Objective The aim of this study was to investigate the diagnostic impact of polymerase chain reaction (PCR) assays for fungal pathogens in fluid samples,and to evaluate the feasibility of fast PCR diagnostic method for the invasive fungal infection.Methods The sterility body fluid samples from 60 cases hospitalized immunocompromised patients with clinical underlying diseases and suspect of invasive infections with fungi between January 2008 and December 2011 were processed for microscopy and cultures.Applying fungal ITS4 and ITS86 universal primers to amplify pathogenic fungi genes from the sterility body fluid with method of rapid PCR.The results were compared between the standard and PCR methods.Results Humoral direct clinical specimens by PCR amplification of DNA fragments with the scan results were similar.Positive rate of PCR test with clinical body fluid samples and the traditional fungal cultivation was similar.There was no significant statistical difference [38.3％ (23/60) vs 33.3％ (20/60),P ＞ 0.05].Conclusions PCR test is feasibility with clinical fungal diagnosis from directly humoral specimens.To amplify the clinical sterility body fluid samples with ITS fungal universal primers and PCR method might provide an accurate and rapid approach to detect the pathogenic fungi.Its methods on early diagnosis and prognosis of invasive fungal infections are of guiding significance.

Objective To investigate the status of the using and knowledge about hazard in reusing needles of insulin pen. Methods The self-designed questionnaire about insulin pen using was used. A total of 358 patients using insulin pens were surveyed. Results Only 10. 84% patients knew the hazards of reusing needles of insulin pen. The influencing factors were incomes, educator, experience and risk education.Conclusions At present, the safety usage of insulin pens needles in the diabetic patients is not optimistic. The risk related health education should be stressed.

OBJECTIVEAn ion chromatographic method was developed to analyze sulfur dioxide in traditional Chinese medicine (TCM).

METHODThe samples were extracted with alkaline solution and reacted with formaldehyde to form hydroxymethyl sulfonic acid. The extracts were cleaned by C18 column and SO3(2-) was determined with ion chromatography equipped with a conductance detector. In the method, AS11-HC chromatographic column was used and the mobile phase was 15 mmol x L(-1) KOH.

RESULTThe calibration curve was in good linearity in the range of 1 to 100 microg (r =0.9992). The detection limit was 0.53 mg x kg(-1) and the average recovery was 103.7%.

CONCLUSIONThe method is simple, sensitive, precise, short-time-consuming, and the random error is smaller, which is applicable to the testing of large quantity batches of TCM.

Alkalies ; chemistry ; Chromatography, Ion Exchange ; methods ; Drug Contamination ; prevention & control ; Drugs, Chinese Herbal ; analysis ; Plants, Medicinal ; chemistry ; Quality Control ; Sulfur Dioxide ; analysis

Objective To investigate the ability of invasion and migration of breast cancer MDA-MB-231 cells under serum starvation and hypoxia, and the effect of antiangiogenic drugs, rh-endostatin and bevacizumab, on the ability of invasion and migration of breast cancer cells under serum starvation and/or hypoxia, in order to explore the potential risk of antiangiogenic therapy in clinics.Mte hods The cells were randomized into 4 groups, i.e., group A:10%fetal bovine serum ( FBS) group;group B:hypoxia +10%FBS group;group C:serum starvation group;group D: hypoxia +serum starvation group; each group was further divided into three subgroups as blank control, treated with rh-endostatin and bevacizumab, respectively.Cell counting kit-8( CCK-8) was used to assess the inhibition rate of cell growth induced by endostatin and bevacizumab, in order to determine the proper working concentration and time of the two drugs.Transwell assay was conducted to detect the cell invasion and migration in vitro.The expressions of c-Met and MMP-9 were detected by Western blot.The cells treated with rh-endostatin or bevacizumab under serum starvation were tested by hybridization using Exiqon miBase 18.0 microarray.The miRNAs which exibited significant differences ( P<0.05) in miRNA hybridization were verified by real-time PCR assay. Results CCK-8 assay showed that the inhibition rates of MDA-MB-231 cells cultured with 800 mg/L rh-endostatin for 48 h and 24 h were (32.2±2.5)%and (27.0±1.3)%, respectively, showing a significant difference ( P=0.023) .The inhibition rates of MDA-MB-231 cells cultured with 80 mg/L bevacizumab for 48 h and 24 h were (30.5±1.4)%and (26.1±2.4)%, respectively , showing also a significant difference (P=0.015).The Transwell assay showed that in the starvation blank group, the number of invaded and penetrated cells were 28.8 ±2.2 and 31.4 ±1.5, respectively, significantly different from that in the rh-endostatin and bevacizumab groups (P<0.05).The relative expressions of c-Met and MMP-9 were 0.213± 0.017 and 0.542±0.048, respectively, with a significant difference from those of the groups treated with each drug ( P<0.05 for both) .The numbers of penetrated cells in the Transwell assay treated with rh-endostatin in hypoxia were 17.5±2.1 and 16.5 ±2.8, respectively, and the numbers of penetrated cells in the Transwell assay treated with bevacizumab were 16.3±3.5 and 17.5±2.4, respectively, showing no significant difference among them ( P>0.05 for both ) .The ability of migration and invasion of MDA-MB-231 cells and the expression of c-Met and MMP-9 were not impacted by hypoxia ( P>0.05) .Real-time PCR assay showed that only the levels of miR-2355 and miR375 were significantly and stably decreased in the cells which had increased ability of invasion and migration.The relative expression levels of miR375 and miR-2355 in the serum starvation blank group were 0.550±0.036 and 0.852±0.121, respectively, significantly lower than that in the groups treated with rh-endostatin or bevacizumab (P<0.05).In the serum starvation group, the expression levels of miR375 and miR-2355of cells treated with rh-endostatin were 02.95±0 .012 and 0.253±0.011, and the expression levels of cells treated with bevacizumab were 0.234±0.020 and 0.309±0.02,2 respectively, ( P>0.05 for all).Compared with the serum starvation blank group, the expression levels of miR2355 and miR375 were significantly decreased when cells were treated with rh-endostatin/bevacizumab under serum starvation, but no significant difference was found between the two drugs ( P>00.5 ).However, hypoxia did not affect the expressions of miR2355 and miR375 ( P>0.05).Conclusions The results of this study suggest that serum starvation can increase the ability of invasion and migration of breast cancer cells.Furthermore, both rh-endostatin and bevacizumab may enhance their invasion and penetration ability under serum starvation condition.

Objective To investigate the ability of invasion and migration of breast cancer MDA-MB-231 cells under serum starvation and hypoxia, and the effect of antiangiogenic drugs, rh-endostatin and bevacizumab, on the ability of invasion and migration of breast cancer cells under serum starvation and/or hypoxia, in order to explore the potential risk of antiangiogenic therapy in clinics.Mte hods The cells were randomized into 4 groups, i.e., group A:10%fetal bovine serum ( FBS) group;group B:hypoxia +10%FBS group;group C:serum starvation group;group D: hypoxia +serum starvation group; each group was further divided into three subgroups as blank control, treated with rh-endostatin and bevacizumab, respectively.Cell counting kit-8( CCK-8) was used to assess the inhibition rate of cell growth induced by endostatin and bevacizumab, in order to determine the proper working concentration and time of the two drugs.Transwell assay was conducted to detect the cell invasion and migration in vitro.The expressions of c-Met and MMP-9 were detected by Western blot.The cells treated with rh-endostatin or bevacizumab under serum starvation were tested by hybridization using Exiqon miBase 18.0 microarray.The miRNAs which exibited significant differences ( P<0.05) in miRNA hybridization were verified by real-time PCR assay. Results CCK-8 assay showed that the inhibition rates of MDA-MB-231 cells cultured with 800 mg/L rh-endostatin for 48 h and 24 h were (32.2±2.5)%and (27.0±1.3)%, respectively, showing a significant difference ( P=0.023) .The inhibition rates of MDA-MB-231 cells cultured with 80 mg/L bevacizumab for 48 h and 24 h were (30.5±1.4)%and (26.1±2.4)%, respectively , showing also a significant difference (P=0.015).The Transwell assay showed that in the starvation blank group, the number of invaded and penetrated cells were 28.8 ±2.2 and 31.4 ±1.5, respectively, significantly different from that in the rh-endostatin and bevacizumab groups (P<0.05).The relative expressions of c-Met and MMP-9 were 0.213± 0.017 and 0.542±0.048, respectively, with a significant difference from those of the groups treated with each drug ( P<0.05 for both) .The numbers of penetrated cells in the Transwell assay treated with rh-endostatin in hypoxia were 17.5±2.1 and 16.5 ±2.8, respectively, and the numbers of penetrated cells in the Transwell assay treated with bevacizumab were 16.3±3.5 and 17.5±2.4, respectively, showing no significant difference among them ( P>0.05 for both ) .The ability of migration and invasion of MDA-MB-231 cells and the expression of c-Met and MMP-9 were not impacted by hypoxia ( P>0.05) .Real-time PCR assay showed that only the levels of miR-2355 and miR375 were significantly and stably decreased in the cells which had increased ability of invasion and migration.The relative expression levels of miR375 and miR-2355 in the serum starvation blank group were 0.550±0.036 and 0.852±0.121, respectively, significantly lower than that in the groups treated with rh-endostatin or bevacizumab (P<0.05).In the serum starvation group, the expression levels of miR375 and miR-2355of cells treated with rh-endostatin were 02.95±0 .012 and 0.253±0.011, and the expression levels of cells treated with bevacizumab were 0.234±0.020 and 0.309±0.02,2 respectively, ( P>0.05 for all).Compared with the serum starvation blank group, the expression levels of miR2355 and miR375 were significantly decreased when cells were treated with rh-endostatin/bevacizumab under serum starvation, but no significant difference was found between the two drugs ( P>00.5 ).However, hypoxia did not affect the expressions of miR2355 and miR375 ( P>0.05).Conclusions The results of this study suggest that serum starvation can increase the ability of invasion and migration of breast cancer cells.Furthermore, both rh-endostatin and bevacizumab may enhance their invasion and penetration ability under serum starvation condition.

OBJECTIVETo investigate the incidence, pathogens, and clinical features of infection in consecutive cases from 2010 to 2012 in Peking Union Medical College Hospital.

METHODThe incidence, pathogen, treatment, and outcomes of patients with hematological diseases who had positive findings of bacterium in their samples from 2010 to 2012 were retrospectively analyzed.

RESULTSThere were 449 positive samples (5.8%) from 4 890 patients during this period, among which 388 were proved to be with pathogenic bacteria. Samples separated from patients with community-aquired infections accounted for 8.4% of all positive samples. Most community-aquired infections were caused by Gram-negative bacteria (75%), although no multidrug-resistant bacteria was observed. Samples separated from patients with nosocomial infections accounted for 91.6% of all positive samples. Respiratory tract (49.4%) and peripheral blood (32.6%) were the most common samples with positive results. Skin soft tissues (10.4%), and urine (3.7%) were less common samples. Most of the pathogenic bacteria of the nosocomial infections were Gram-negative (66.9%). The most common Gram-negative bacteria included Escherichia coli (13.8%), Pseudomonas aeruginosa (12.1%), and Klebsiella pneumonia (12.1%), while Staphylococcus aureus (10.4%), Enterococcus faecium (7.0%), and Staphylococcus epidermidis (5.1%) were the most common Gram-positive bacteria. Gram-negative bacteria consisted of most of sputum samples and peripheral blood samples. Samples from the surface of skin wound and anal swab were composed largely by Gram-positive bacteria (63.8%). The detection rates of extended-spectrum beta-lactamase-producing Klebsiella pneumonia/Klebsiella oxytoca, Escherichia coli, and Proteus mirabilis were 24.0%, 87.9% and 38.4%, respectively. The resistance to Acinetobacter baumannii was serious. Multidrug-resistant, extensive drug resistant and pan drug resistant A. baumannii acountted for 74% of all A. Baumannii infections. Stenotrophomonas maltophilia showed low resistance to sulfamethoxazole/trimethoprim, levofloxacin and minocycline. Also, 22 methicillin-resistant Staphylococcus aureus and 9 methicillin-resistant Staphylococcus Epidermidis were detected, which were only sensitive to vancomycin, teicoplanin, and linezolid. All patients were treated in the haematology wards and most of them were under agranulocytosis or immunosuppression. Finally, 22 patients reached clinical recovery through anti-infective therapy, whereas 49 patients died. Among those deaths, 42 patients attributed to severe infections and infection-associated complications. Fourteen of all the deaths might be infected with drug-resistance bacteria. There were 61 samples proved to be bacteria colonization. Nonfermenters such as Acinetobacter baumannii and Stenotrophomonas maltophilia made up for a large amount of bacteria colonization.

CONCLUSIONSThe pathogens of nosocomial infections in the hematology ward are mainly Gram-negative bacteria. The incidences and pathogens vary from different infection sites. Nosocomial infection still has a higher mortality rate. Once nonfermenters are detected positive, the pathogenic or colonial bacteria should be distinguished.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Bacteria ; isolation & purification ; Bone Marrow Transplantation ; Cross Infection ; microbiology ; Female ; Hematologic Diseases ; complications ; microbiology ; Hematology ; Hospital Departments ; statistics & numerical data ; Humans ; Male ; Middle Aged ; Retrospective Studies ; Young Adult