A temperature-sensitive mutant strain was isolated after transposon mutagenesis with Tn5 and named MT54.PCR was carried out with primers designed according to the sequence of transposon,the PCR products showed that the MT54 carried transposon in the genome. The mutant grew well at 30℃in minimal medium(MM)containing p-nitrophenol(PNP)as sole carbon source,while it cannot grow at 37℃in the same medium,NO_2.detection results also proved that.Comparing the degradation rate of PNP and hydroquinone of MT54 and DLL-E4 at different temperature,it was speculated that the mutant site locate in the PNP degradation related genes.

OBJECTIVETo study the synergistic effects of hypothermia and hypoxia on the damage of pulmonary microvascular endothelial cells (PMVEC) in rat.

METHODSPrimary PMVECs were obtained by complex phosphoesterasum digesting from isolated lung tissues of Wistar rats, the PMVECs were identified by phase contrast microscope and immunofluorescence studies for CD31 antigen and bandeiraea simplicifolia isolectin (BSI) binding test. Factorial design was adopted in trial according to hypothermia and hypoxia existing or not. Using corresponding kit measured the levels of lactate dehydrogenase (LDH) activity in cell medium. Level of nitric oxide (NO) concentration was measured by Griess Assay. RT-PCR was used to examine the expression of vascular endothelial growth factor (VEGF), endothelin-1 (ET-1) mRNA in PMVECs.

RESULTSThe monolayer of cultured PMVECs displayed the shape of pavingstone. CD31 antigen and binding BSI results by fluorescence microscope identified the cultured cells were PMVECs. Compared to the control group, LDH activity and VEGF, ET-1 expression levels were significantly increased in hypothermia group, hypoxia group and hypoxia combined with hypothermia group. And the levels of NO concentration were reduced in these three groups. The results of One-way ANOVA showed that there was a synergistic effect between hypothermia and hypoxia.

CONCLUSIONHypothermia and hypoxia both have an effect on PMVECs whether in altering the cell permeability or in releasing of vasoactive substances including NO and ET-1. In addition, there is a synergistic effect between hypothermia and hypoxia.

Animals ; Cell Hypoxia ; Cell Membrane Permeability ; Cells, Cultured ; Cold Temperature ; Endothelial Cells ; cytology ; metabolism ; Endothelin-1 ; metabolism ; Endothelium, Vascular ; cytology ; Lung ; blood supply ; Male ; Nitric Oxide ; metabolism ; RNA, Messenger ; genetics ; Rats ; Rats, Wistar ; Vascular Endothelial Growth Factor A ; metabolism

Objective To evaluate the diagnostic value of the heparin-binding protein (HBP),procalcitonin (PCT),C-reactive protein (CRP),white blood cell (WBC) in respiratory tract bacterial infection.Methods 66 respiratory tract bacterial infection patients,37 respiratory tract non-bacterial infection patients and 39 control group in the Third Xiangya Hospital from October 2015 to March 2017 was selected as objects in this prospective study.The levels of HBP,PCT and CRP in blood of the objects were tested with ELESA,immunofluorescence assay,immunoturbidimetry respectively;WBC counts were taken by Sysmex XE-5000 blood analyzer.The difference among the three groups was analyzed by Student's t test,one-way ANOVA or Wilcoxon test.Receiver operating characteristic curve was utilized to analyze the diagnostic value of HBP,PCT,CRP and WBC in respiratory tract bacterial infection.Results The plasma level of HBP were 36.30 (7.78-89.36) ng/ml,5.57 (4.37-8.23) ng/ml,2.84 (1.53-6.51) ng/ml in respiratory tract bacterial infection group,respiratory tract non-bacterial infection group and control group respectively.The socre of PCT were 0.08 (0.04-0.83) ng/ml,0.09 (0.04-0.30) ng/ml,0.04 (0.03-0.08) ng/ml.The socre of CRP were 56.20 (19.33-76.23) mg/L,34.40 (2.15-83.95) mg/L,(2.20 ± 0.99) mg/L.The socre of WBC count were (10.59 ±4.58) × 109/L,8.40 (5.80-11.88) × 109/L,(6.14± 1.31) × 109/L.There were statistically significant differences in HBP scores between respiratory tract bacterial infection group and respiratory tract non-bacterial infection group or control group (Z =-4.828,P ＜0.001;Z =-5.685,P ＜ 0.001).There were no statistically significant differences in PCT,CRP and WBC scores between respiratory tract bacterial infection group and non-bacterial infection group (F =0.045,P ＞ 0.05;F =0.100,P ＞ 0.05;F =2.417,P ＞ 0.05),but significant differences between respiratory tract bacterial infection group and control group (Z =-2.881,P ＜ 0.05;Z =-6.595,P ＜ 0.001;t =6.499,P ＜ 0.001).The area under curve (AUC) of HBP,PCT,CRP and WBC diagnosing respiratory tract bacterial infection was 0.89,0.69,0.95 and 0.85 respectively.The AUC of HBP differential diagnosising was 0.80.Conclusion HBP can be used as an efficient supplementary indicator for respiratory tract bacterial infection,the differential diagnostic value is superior to PCT,CRP and WBC.

Abstract：Objective To analyze the stabilities of neuraminidase（NA）in influenza vaccine at different temperatures and provide a reference for further complete understanding of overall shelf life of vaccines. Methods Monovalent bulks of influenza H1N1，H3N2 and B vaccines were stored at 4（low temperature），25（room temperature）and 37 ℃（changed temperature）for 0. 5，2，7，24 and 48 h separately，using that at 100 ℃（extreme temperature）for 1 h as control，and determined for NA activity by enzyme⁃linked lectin method. Results The NA activities of influenza H1N1 vaccines stored at 25 and 37 ℃ decreased significantly with the increasing of time. No significant decreases were observed in H3N2 and B vaccines even after storage at two non⁃storage temperatures for 48 h. However，all the NA activities of three vaccines decreased at 100 ℃. Conclusion Both H3N2 and B vaccines showed high stability at abnormal storage temperatures not more than 37 ℃，while H1N1 vaccine was relatively sensitive to the temperature for storage.

OBJECTIVETo explore the damage effects and expression of vascular endothelial growth factor (VEGF) exposed with different low-temperatures on rat dermal microvascular endothelial cells (DMVECs).

METHODSPrimary DMVECs were obtained by discontinuous Percoll gradient centrifugation. The DMVECs were identified by phase contrast microscope and immunofluorescence studies for CD31 antigen. Applied 28 degrees C, 12 degrees C and 0 degrees C to interfere with rat DMVECs as cold-exposure model. The changes of cells morphology were observed under invert microscope. The membrane integrity was determined by lactate dehydrogenase (LDH) activity. RT-PCR was used to examine the expression of vascular endothelial growth factor mRNA in cells.

RESULTSThe monolayer of cultured PMVECs displayed the shape of pavingstone. CD31 antigen and binding BSI results by fluorescence microscope identified the cultured cells were DMVECs. After 24 h cold exposure, the cell morphology of 0 degrees C group was shrunken, the other groups were "Fibroblast-like". The LDH activity (U/L) in the medium of 28 degrees C, 12 degrees C and 0 degrees C groups was 54.17 +/- 3.02, 64.66 +/- 3.03, 82.13 +/- 10.91 respectively, which was significantly higher than that of 37 degrees C group (12.23 +/- 3.0, P < 0.01). The VEGF mRNA expression level was up-regulated in 28 degrees C group and 12 degrees C group versus control group (P < 0.05), but unchanged in 0 degrees C group.

CONCLUSIONThe rat DMVECs injury severity are deteriorated with temperature decreasing, and VEGF might be involved in the regulation of membrane permeability in this period.

Animals ; Animals, Newborn ; Cells, Cultured ; Cold Temperature ; Dermis ; blood supply ; Endothelial Cells ; cytology ; metabolism ; Endothelium, Vascular ; cytology ; Rats ; Rats, Wistar ; Vascular Endothelial Growth Factor A ; metabolism

OBJECTIVETo study protective effect and pathogenesis of complex salvia miltiorrhiza (DanShen) on acute mercury poisoning in rabbits.

METHODSModels of acute mercury poisoning was made in rabbits. The effect of complex salvia miltiorrhiza on blood urea nitrogen (BUN), copper-protein (CP), lactate dehydrogenase (LDH), acid phosphatase (ACP) and the malondialdehyde (MDA) in plasma, superoxide dismutase (SOD) in erythrocyte and MDA, SOD in tissues homogenate were observed.

RESULTSThe administration of complex salvia miltiorrhiza after mercury injection 0.5 h and 9.5 h, decreased BUN, CP, MDA, LDH and ACP, and prevented the reduction of SOD. Compared with mercury poisoning group, the difference was statistical significant (P < 0.05, P < 0.01).

CONCLUSIONIt is suggested that acute mercury poisoning may result in renal damage but also multiple organ tissues, and complex salvia miltiorrhiza possesses protective effect, through stabilized membranes.

Animals ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Female ; Male ; Mercury Poisoning ; blood ; drug therapy ; Phenanthrolines ; pharmacology ; therapeutic use ; Rabbits ; Salvia miltiorrhiza

OBJECTIVETo investigate the protective effects of natakalim against rat aortic vascular endothelial cells (RAVECs) injuries induced by hypoxia and its mechanisms.

METHODSSelecting RAVECs as a cell model injured by hypoxia, these RAVECs were divided into 5 groups: i.e. control group, hypoxia group, natakalim low, medium and high group. The cell survival rate was determined by MTT assay, con was measured using Griess Assay, RT-PCR was used to examine t he expression of intercellular adhesion molecule-1 (ICAM-1), vascular endothelial growth factor (VEGF), endothelin-1 (ET-1) mRNA in RAVEC.

RESULTSNatakalim could reverse hypoxia-induced changes in endothelial cell function, including increased endothelial cell survival rate and level of NO concentration, significantly inhibited the hypoxia-induced endothelial ICAM-1, ET-1, VEGF mRNA expression levels increased.

CONCLUSIONNatakalim have protective effects on hypoxia-induced changes in endothelial cell function, increasing of permeation, excess expression of cell adhesion molecules.

Allyl Compounds ; pharmacology ; Animals ; Aorta ; cytology ; metabolism ; Cell Hypoxia ; Cells, Cultured ; Endothelial Cells ; metabolism ; Endothelin-1 ; metabolism ; Intercellular Adhesion Molecule-1 ; metabolism ; Male ; Propylamines ; pharmacology ; RNA, Messenger ; genetics ; Rats ; Rats, Wistar ; Vascular Endothelial Growth Factor A ; metabolism ; Vascular System Injuries ; metabolism

OBJECTIVETo compare the toxicity of Euphorbia pekinensis before and after being processed by vinegar on normal liver cells LO2, and discuss its possible mechanism.

METHODLO2 cells were cultured in vitro, and processed with different concentrations of crude and vinegar-processed E. pekinensis. MTT assay was used to measure the inhibitory effect of LO2 cell; Hoechst 33258 staining was used to observe the morphological changes in apoptosis cell; Annexin V-FITC flow cytometry was used to analyze the apoptotic rate of LO2 cell; PI staining flow cytometry was used to analyze its impact on cell cycle. The level or content of ALT, AST, LDH, SOD, MDA and GSH were observed as well.

RESULTCompared with the negative control group, crude E. pekinensis at all concentrations could obviously inhibit LO2 cell proliferation, induce LO2 cell apoptosis and cause cell arrest in S phase, with significant differences (P <0.05). E. pekinensis could significantly increase the levels of ALT, AST and LDH (P <0.05) in the supernatant of cell culture fluid, significantly decrease the level of SOD and the content of GSH (P <0.05) , and significantly increase the content of MDA (P <0.05). Compared with the crude E. pekinensis group, E. pekinensis after being vinegar-processed can significantly reduce cell apoptotic rate, cell cycle arrest, activities of ALT, AST, LDH in the supernatant of cell culture fluid (P <0.05) , and remarkably increase the level of SOD and the content of GSH, but reduce the content of MDA in the supernatant of cell culture fluid.

CONCLUSIONVinegar-processed E. pekinensis can release the cytotoxicity of LO2 cell. Its mechanism may be related to the decrease in the oxidative damage of LO2 cells, thereby reducing the cell cycle arrest and apoptosis.

Acetic Acid ; chemistry ; Apoptosis ; drug effects ; Cell Cycle Checkpoints ; drug effects ; Cell Line ; Cell Membrane ; drug effects ; metabolism ; Cell Proliferation ; drug effects ; Chemistry, Pharmaceutical ; Drugs, Chinese Herbal ; chemistry ; toxicity ; Euphorbia ; chemistry ; Humans ; Liver ; cytology ; Oxidative Stress ; drug effects

BACKGROUNDFoetal echocardiography has become a diagnostic method to detect foetal congenital heart disease with high probability. However, it is not only time consuming and but also difficult to visualize outflow tract of foetus early in the second trimester of pregnancy, even for an experienced obstetric ultrasonographer. Recently, many methods for screening foetal cardiac anomalies were explored, but much more work is needed to develop an effective and suitable screening method. The aim of this study was to investigate the clinical significance of utilising the ductus venosus (DV) Doppler examination and the four-chamber view of heart to screen for foetal cardiac malformation in early second trimester of pregnancy.

METHODSHeart and DV of 401 consecutive foetuses in early second trimester (12(+1) - 17(+6) weeks) in high risk pregnancies were examined with Acuson 128 xp/10 or Sequoia 512 ultrasound diagnostic systems. Absent or reversed flow during atrial contraction (A-wave) in the DV was defined as sufficiently abnormal to screen for foetal cardiac malformations. The foetal echocardiographic diagnosis was confirmed by postnatal echocardiography (or postmortem). The sensitivities of screening tests were compared among the three methods: DV Doppler examination, four-chamber view alone, and the combination of both techniques.

RESULTSSatisfactory examinations were obtained in 383/401 foetuses (95%). Thirty foetuses with cardiac abnormalities were confirmed by neonatal echocardiography (or postmortem). The sensitivity of DV Doppler examination or four-chamber view alone is 63% (19/30) and 60% (18/30), respectively. The sensitivity of combining information, DV Doppler flow waveform and four-chamber view, to screen for foetal cardiac malformation is 83% (25/30) and significantly better than that of either DV Doppler flow waveform or four chamber view alone (P < 0.05).

CONCLUSIONDoppler flow waveform of DV can be used to screen for foetal cardiac malformation early in the second trimester. Combining information from Doppler flow waveform of DV and four-chamber view will improve the overall sensitivity of the screening.

Female ; Heart Defects, Congenital ; diagnostic imaging ; Humans ; Pregnancy ; Pregnancy Trimester, Second ; Ultrasonography, Doppler ; Ultrasonography, Prenatal