HCV RNA positive serum was first selected by RT-PCR test kit from several anti-HCV positive sera obtained from Xi'an.HCV RNA extrac ted from the elected sera was converted to cDNA by reverse transcription with ra ndom primer.Half-nested PCR was performed.The amplified product was 852 bp.The purified PCR product was digested by restriction endonucleases and then ligated to epressio vector pET-22b\++.Its nucleotide sequence was determined by dideoxy chain termination method.A comparison of the sequence with several isolates rep orted previously showed that the sequence belonged to HCV type Ⅱ.

Cloning, Molecular ; Escherichia coli ; genetics ; Hepacivirus ; genetics ; Humans ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; immunology ; Viral Nonstructural Proteins ; biosynthesis ; genetics ; immunology

OBJECTIVETo establish the national quantity standard of hepatitis B surface antigen according to the world health organization' s standard material and prepare the national liner reference panel for hepatitis B surface antigen.

METHODSSera from hepatitis B patients and health blood donors in different areas were collected and detected by domastic HBsAg kits, anti-HBs kits, HBeAg kits, anti-HBe kits, anti-HBc kits and anti-HCV, and then confirmed by the kits produced by Abbott, which was approved by WHO. One serum with high concentration of HBsAg was calibrated with the standard sample of WHO. And then it was diluted by 1.5 fold as the liner HBsAg reference panel.

RESULTSThe HBsAg concentration of one serum was 1226 IU/ml calibrated by 21 independent standardization measurements with 7 kinds of kits. The coefficient of variation of each calibration were less then 15%. A panel contained 8 serial dilutions was established as the national liner HBsAg reference panel. The permitted range of every dilution was stipulated and the stability of the panel was detected by accelerated test.

CONCLUSIONSThe national quantity standard of hepatitis B surface antigen was established and the national quantitative reference panel for HBsAg which contains eight liner serum was developed.

China ; Hepatitis B ; virology ; Hepatitis B Surface Antigens ; blood ; Humans ; Immunoenzyme Techniques ; methods ; standards ; Reagent Kits, Diagnostic ; standards ; Reference Standards

Objective To observe the change in cardiac shape and heart function and evaluate the effect of self-treatment on patients with Keshan disease by echocardiography. Methods To check the 31 patients with Keshan disease before the self-treatment, and follow them up in the 3rd and 6th months after self-treatment by echocardiography. The left atrium diameter(LAd), left ventricular end-diastolic diameter(LVEDd), the thickness of interventricular septum in end-diastolic(IVSTd), the thickness of LV posterior wall in end-diastolic (LVPWTd), left ventricular mass(LVM), left ventricular mass index(LVMI), left ventricular ejection fraction(LVEF) and mitral valve flow E/A ratio(E/A) were measured. Results The LAd[(35.8±5.1)ram] and LVPWTd[(9.3±1.0)mm] obviously decreased in the 3rd month after serf-treatment compared with prior self-treatment [ (37.0±5.0), (9.9± 1.2)mm](P<0.05). The LAd[(34.5±5.0)mini, IVSTd[(9.5±1.3)mm], LVEDd[(50.2±7.7)mm], LVPWTd [(8.7±1.1)mm], LVM[(196.1±87.2)g] and LVMl[(126.5±56.4)g/m2] obviously decreased in the 6th month after self-treatment compared with prior self-treatment [(37.0±5.0), (10.2±1.5), (51.3±8.1), (9.9±1.2)mm, (230.4±95.5)g, (144.0±54.6)g/m2] and in the 3rd month after self-treatment [(35.8±5.1)mm, (10.2±1.4) ram, (51.1±8.1)nun, (9.3±1.0)mm, (219.4±82.5)g, (136.8±50.0)g/m2] (P<0.05). The results of the mitral valve flow E/A ratio and LVEF in the 3nt month after self-treatment [1.0±0.5, (59.4±13.3)%] were increased compared with the prior self-treatment[0.9±0.5, (58.1±15.6)%], and the results in the 6th month after self-treat- ment[ 1.0±0.4, (60.7±13.6)%] were further inereased compared with before, but there was no signifieant differ- ence(P0.05). Conclusions Self-treatment of Keshan disease patients can improve the heart function by pre- venting left ventrieular remodeling and reversing. Echocardiography can be used as an essential technique to evalu- ate the effect of self-treatment on Keshan disease patients.

OBJECTIVETo evaluate anti-HEV diagnostic kits by experimental infecting rhesus monkeys with HEV.

METHODSEight rhesus monkeys were infected with genotype 1 and 4 HEV separately. The alanine aminotransferase (ALT) level of all monkeys were detected before and after the process of infection. HEV RNA in stool specimens was tested by reverse transcriptase-polymerase chain reaction (RT-PCR) assay. Anti-HEV IgG in serum was detected by GL-IgG and WT-IgG.

RESULTSHEV RNA presented in the stool of all the 8 monkeys after infection. The ALT level of 1 monkey infected with genotype 1 HEV and 2 monkeys infected with genotype 4 HEV appeared abnormally after infection. Tested by GL-IgG, 2 of the 4 monkeys infected with genotype 1 HEV and 1 of 4 monkeys infected with genotype 4 HEV seroconverted to anti-HEV IgG. However, when tested by WT-IgG, all the infected monkeys seroconverted to anti-HEV IgG. The anti-HEV IgG tested by WT-IgG was positive during the whole observation period,and the anti-HEV IgG measured by GL-IgG only remained 12 weeks after infection. Detected by GL-IgG and WT-IgG, seropositive conversion of the anti-HEV IgG happened almost at the same time.

CONCLUSIONBoth GL-IgG and WT-IgG could detect the anti-HEV IgG of experimentally infected rhesus monkeys but the WT-IgG had a higher sensitivity for detection of anti-HEV IgG than

Alanine Transaminase ; blood ; Animals ; Disease Models, Animal ; Genotype ; Hepatitis E ; immunology ; virology ; Hepatitis E virus ; genetics ; immunology ; Immunoglobulin G ; immunology ; Macaca mulatta ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVE To investigate whether Renqing Changjue has a protective effect on acute respiratory dis?tress syndrome (ARDS) induced by endotoxin lipopolysaccharide (LPS) in rabbits. METHODS Thirty-six healthy male New Zealand white rabbits were randomly divided into six groups: normal control group, model group, dexamethasone group, Renqing Changjue high, middle and low dose group, with six rabbits in each group. LPS was used to replicate the ARDS model after five consecutive days of gavage. Arterial pressure, respiratory rate and anal temperature blood were recorded for arterial blood gas analysis at 0, 0.5, 1, 2 and 4 h, respectively. At the end of the four-hour experiment, rab?bits were killed by bloodletting, and the lung tissue was quickly removed to determine the cytokines, SOD, MDA and pathological examination of rabbit lung. RESULTS Renqing Changjue can significantly reduce the pathological changes of lung in ARDS model group. The expression of AQP1 and MPO in rabbit lung was significantly decreased by immuno?histochemistry (P<0.05) ,reduce the lung wet/dry weight ratio, increase the ratio of PaO2/FiO2, inhibit the release of inflammatory factors and scavenge free radicals and antioxidant effects. CONCLUSION Renqing Changjue can effec?tively protect rabbits with acute respiratory distress syndrome induced by LPS, and may protect the lung by inhibiting the release of cytokines and anti-oxidation.

OBJECTIVETo study the impact of TNM staging and combined treatment mode on the survival of non-small cell lung cancer (NSCLC) patients.

METHODSFrom January 1997 to December 2002, 987 NSCLC patients were surgically treated in this hospital. Of those, 574 received combined modality therapy (surgery + chemotherapy/radiotherapy), while 413 underwent operation alone. Their clinicopathological data were retrospectively analyzed.

RESULTSThe 1-, 3-, 5-, and 10-year overall survival rates were 87.7%, 57.5%, 54.6% and 54.5%, respectively, for the whole group, which were 90.6%, 57.5%, 54.3% and 54.1% for the combined therapy group versus 83.8%, 57.6%, 55.2% and 55.2% for the group treated by surgical resection alone. The 1-year survival rate of the combined therapy group was significantly higher than that of the surgical resection alone group (90.6% vs. 83.8%) (P<0.01). With regard to the T factor, 5-year survival rate of the combined therapy group (surgery + radiotherapy) was higher than that of surgery alone group, especially in T4 cases (43.6% vs. 12.7%), with a significant difference between them (P<0. 05). As for the N factor, the 1-year survival rate of NO patients in the combined therapy group (surgery + chemotherapy/radiotherapy) was significantly higher than that of surgery alone group (94.4%, 97.9% vs. 90.0%) (P<0.05). The 1-year survival rate of N1 patients in the combined therapy group (surgery + chemotherapy or + chemotherapy and radiotherapy) was 91.7% and 100% versus 82.9% in the surgery alone group (P<0.01); The 1- and 3-year survival rates of N2 patients in the combined modality therapy group (surgery + chemotherapy) were 82.1% and 37.3%, while those of the surgery alone group were 69.4% and 26.5%, respectively, with a significant difference (P<0.05, P<0.01). All the severity of primary tumor, distance of lymph node involvement, and distant tumor metastasis significantly worsen the prognosis of the patients.

CONCLUSIONThe prognosis in NSCLC patients treated with combined modality therapy (surgery + chemotherapy/radiotherapy) is better than that with surgery alone. The larger the original tumor and the farther the lymph node and tumor metastases, the worse the prognosis is for NSCLC patients.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Carcinoma, Non-Small-Cell Lung ; pathology ; therapy ; Chemotherapy, Adjuvant ; Child ; Female ; Follow-Up Studies ; Humans ; Lung Neoplasms ; pathology ; therapy ; Lymphatic Metastasis ; Male ; Middle Aged ; Neoplasm Staging ; Pneumonectomy ; methods ; Radiotherapy, Adjuvant ; Retrospective Studies ; Survival Rate ; Young Adult

Objective To investigate the predictive factors of symptomatic intracranial hemorrhage(SICH) and the effect on outcomes after mechanical thrombectomy in patients with acute ischemic strokecaused by anterior circulation tandem occlusion. Methods From January 2014 to September 2018, patients with acute ischemic stroke caused by anterior circulation tandem occlusion and treated by endovascular mechanical thrombectomy from the Departments of Neurology in 5 general hospitals (the First Affiliated Hospital of Henan University of Science and Technology, the Affiliated Hospital of Yangzhou University,General Hospital of Eastern Theatre Command of PLA, Yijishan Hospital of Wannan Medical College,Xinqiao Hospital of Army Medical University of PLA) were enrolled retrospectively. SICH was evaluated according to the criteria of Heidelberg Bleeding Classification. The functional outcome was assessed by the modified Rankin Scale score at 90 d after onset, and 0-2 was defined as good outcome. Multivariate logistic regression analysis was used to determine the effect of SICH on the outcome at 90 d and the independent risk factors for SICH. Results A total of 124 patients with acute ischemic stroke caused by anterior circulation tandem occlusion and treated by endovascular mechanical thrombectomy were enrolled in this study. Among them, 19 (15. 3％) had SICH, 60 (48. 4％) had good outcomes, and 28 (22. 6％) died. The incidence of poor outcomes (94. 7％ vs. 43. 8％; χ2 = 16. 708, P < 0. 001 ) and mortality (57. 9％ vs.16. 2％; P < 0. 001 ) in the SICH group were significantly higher than those in the non-SICH group.Multivariate logistic regression analysis showed that SICH was an independent risk factor for poor outcome(odds ratio [OR] 27. 78, 95％ confidence interval [CI] 2. 60-96. 70; P = 0. 006), while larger infarct core(low ASPECT score) was the only independent predictor of SICH (OR 2. 63, 95％ CI 1. 18-5. 88; P =0. 018). Conclusion In patients with acute ischemic stroke caused by anterior circulation tandem occlusion and treated by endovascular mechanical thrombectomy, SICH is associated with poor outcome and higher mortality at 3 months, and larger preoperative infarction core is an independent predictor of SICH.

Objective To evaluate the multiplex nucleic acid testing (NAT) assays for HBV,HCV and HIV in detecting HBV DNA in plasma samples. Methods 534 plasma samples collected form several areas were detected with Abbott Architect i2000 HBsAg, ani-HBs, HBeAg, anti-HBe, anti-HBc and anti-HBc IgM diagnostic kits. HBV DNA levels of those samples were detected with Roche COBAS AmpliPrep/ COBAS TaqMan HBV Test. Two kinds of multiplex NAT assays for HBV, HCV and HIV were used to test HBV DNA of those 534 samples. Results of serology-markers and quantitative HBV DNA levels with results of NAT were compared. Results HBV DNA was positive in all 81 HBsAg, HBeAg and anti-HBc positive samples,detected by both of NAT assays. HBV DNA was positive in 11 and 19 of 200 HBsAg negative samples when detected with the two kinds of NAT assays separately. Compared with the quantitative results detected by Roche COBAS AmpliPrep/COBAS TaqMan HBV Test, the HBV DNA positive rates were 96.9% and 94.3% in 193 samples of HBV DNA levels over 500 IU/ml while 40.2% and 45.3% in 117 samples of HBV DNA levels below 500 IU/ml while 99.3% and 96.0% in 151 samples of DNA negative HBV. Conclusion There are some occult low level HBV DNA carriers with HBsAg negative results in China. NAT assays for HBV, HCV and HIV may be useful to improve the transfusion safety.

OBJECTIVELcrV is an important component for the development of a subunit vaccine against plague. To reduce immunosuppressive activity of LcrV, a recombinant LcrV variant lacking amino acids 271 to 326 (rV270) was prepared by different methods in this study.

METHODSA new strategy that produced non-tagged or authentic rV270 protein was designed by insertion of rV270-thrombin-hexahistidine fusion gene into the vector pET24a, or by insertion of hexahistidine-enterokinase-rV270 or hexahistitine-factor Xa-rV270 fusion gene into the vector pET32a. After Co(2+) affinity chromatography, a purification strategy was developed by cleavage of His tag on column, following Sephacryl S-200HR column filtration chromatography.

RESULTSRemoval of His tag by thrombin, enterokinase and factor Xa displayed a yield of 99.5%, 32.4% and 15.3%, respectively. Following Sephacryl S-200HR column filtration chromatography, above 97% purity of rV270 protein was obtained. Purified rV270 that was adsorbed to 25% (v/v) Al(OH)₃ adjuvant in phosphate-buffered saline (PBS) induced very high titers of antibody to rV270 in BALB/c mice and protected them (100% survival) against subcutaneous challenge with 10⁶ CFU of Y. pestis virulent strain 141.

CONCLUSIONThe completely authentic rV270 protein can be prepared by using enterokinase or factor Xa, but they exhibited extremely low cleavage activity to the corresponding recognition site. Thrombin cleavage is an efficient strategy to prepare non-tagged rV270 protein and can be easily operated in a large scale due to its relatively low cost and high cleavage efficacy. The recombinant rV270 can be used as a key component to develop a subunit vaccine of plague.

Amino Acid Sequence ; Animals ; Antibodies, Bacterial ; blood ; Antigens, Bacterial ; genetics ; immunology ; Blotting, Western ; Cloning, Molecular ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli ; genetics ; Female ; Genetic Vectors ; Mice ; Mice, Inbred BALB C ; Molecular Sequence Data ; Plague ; immunology ; prevention & control ; Plague Vaccine ; genetics ; immunology ; Plasmids ; Pore Forming Cytotoxic Proteins ; genetics ; immunology ; Protein Engineering ; methods ; Recombinant Fusion Proteins ; genetics ; immunology ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Survival Analysis ; Vaccines, Subunit ; genetics ; immunology ; Yersinia pestis ; growth & development ; immunology