ObjectiveExpression and purification of human tumor necrosis factorα(hTNFα)fusion protein with a stretch of six consecutive histidine residues(6×His) in E. coli.MethodshTNFα fusion proteins with 6×His at N and C terminus were expressed by using E. coli expression vectors pET- 28a(+) and pET-22b(+). The His6 tag allows the expression fusion protein purified in one step by immobilized metal Ni2+ chelation affinity chromatcgraphy in native state. Results The two construct expression vectors were expressed in E. coli respectively, the former with high level as insoluble protein, account for 45% of the total bacteria proteins and not purified; the later 8% as soluble protein, and characterized by SDS- PAGE, Westren-blot. By using affinity chromatography through Ni2+ - IDA Sepharose 6B, 100ml induction culture had 0.4mg hTNFα-6×His fusion proteins. Its purifity reached 90 %. ConclusionThe purification expression product can possess TNF activity and reach 5.42×104U/mg.

BACKGROUND: Decrease of regional cerebral blood flow (rCBF) perfusion of different degree exists in depressive patients, but the decreased ranges were different, different cerebral cortical and subcortical regions are involved.OBJECTIVE: To observe the types of abnormal rCBF perfusion in patients with depressive disorder.DESIGN: A diagnosis-based case-control analysis.SETTINGS: Department of Nuclear Medicine, Ruijin Hospital Affiliated to the Medical College of Shanghai Jiaotong University; Staff Room of Biomedical Engineering, Medical College of Shanghai Jiaotong University;Putuo District Mental Health of Shanghai City.PARTICIPANTS: The patients with depressive disorder were selected from the Putuo District Mental Health Center from January 1999 to December 2000, and 19 normal subjects matched by age were selected from the medical staff of the hospital as normal controls.METHODS: The patients with depressive disorder and healthy subjects were examined by single photon emission computerized tomography (SPECT), ethyl cysteinate dimmer (99Tcm-ECD) was performed, and the images were analyzed with the statistical parametric mapping (SPM). All the SPECT images were converted into DICOM format or ANALYZE-7 format at first, and then the images were standardized by taking MRI template as the template and smoothed, and the significantly different regions were evaluated by the statistical analysis and inferences with the Gaussian random theory.MAIN OUTCOME MEASURES: The differences of the cerebral blood flow perfusion in specific region and the changes of average rCBF images were mainly observed in the depressive disorder group and normal control group.RESULTS: All the 12 patients with depressive disorder and 19 normal healthy subjects entered the analysis of results. The SPM showed that the blood flow perfusion was decreased in bilateral frontal lobes, left superior temporal gyrus, left Brodmann area 20, left mamillary body of thalamus,right cingulated gyrus and right putamen in the patients with depressive disorder. The blood flow perfusion was increased in the right central frontal gyrus, right superior temporal gyrus, right occipital lobe and left central frontal gyrus.CONCLUSION: The rCBF is obviously changed in the frontal lobe, temporal lobe and limbic system of patients with depressive disorder.

Objective To establish a convenient ~(13)C-breath test system in live mice,and investigate the value of ~(13)C-methacetin breath test(~(13)C-MBT) in the diagnosis of acute liver damage of mice with domestically synthesized ~(13)C-methacetin. Methods Domestically synthesized ~(13)C.methacetin was prepared from aeamol by methylation. Abdominal injection of CCl_4 was adopted to duplicate acute liver damage of mice,then the mice were housed under normal laboratory condition for a whole month to gain recovery,which were indentified by hepatic pathological examinations and biochemical teats of liver function.After fasting, the mice were orally administered ~(13)C-methacetin,and the expired air was collected at various time points. Infrared spectrometer was employed, and delm over baseline(DOB) curves of ~(13)C-exhalation were drawn. Results Six to eight min after administration of ~(13)C-methacetin,the rate of ~(13)C-exhalation peaked in control group(51.9±2.04), and decreased thereafter. Sixteen min after administration of ~(13)C-methacetin,the rate of ~(13)JC-exhalation peaked in model group(26.37±5.74), and decreased thereafter.There were significant differences between these two groups(P<0.05).There was no significant difference in peak value and time to reach the peak on DOB curves of ~(13)C-methacetin breath test after the two groups of mice were housed under the same condition for a month(P>0.05).Conclution ~(13)C-MBT facilitates the collection and evaluation of ~(13)CO_2 in the expired air of live mice,and yields precise reflection of alterations of liver function in acute liver injury and functional recovery.