AIM: To obtain nerve growth factor subunit ?(NGF ?) gene from Chinese fetal hippocampus tissue, prove its sequence, clone the mature peptide sequence, and make it express in E.coli .METHODS: Total RNA was extracted, amplified by RT-PCR method. Its 650 bp DNA sequence was inserted into PCR Ⅱ vector. PCR/NGF ? vector was used as the template to amplfy the C-terminate mature peptide sequence, then subcloned it into PG5 vector. The recombinant was transferred into E.coli BL21. BL21 was cultured and induced by IPTG. The activity of the expressed product was measured after purified and refolded.RESULTS: A complete cDNA sequence was determined as 1 047 nucleotides. The cloned 636 bp encoding 212 amino acids was proved homological to Genbank by sequence analysis. The expressed mature peptide showed an clear band of the prospected 14 kD by SDS-PAGE electrophoresis, and was testified by Western blot. The expression level was about 10% of the total cell lysate. CONCLUSION: The chinese NGF ? gene was homological to the foreigners'. The recombinant NGF ? was efficiently expressed in E.Coli and the recombinant protein has high immunological activities.

Halomonas sp.BYS-1 was a moderately halophilic bacteria strain isolated from activated sludge,It could grow on MM with NaCl concentration from 0.1～2.6 mol/L and phenyl acetic acid as sole carbon souce. When BYS-1 grew in the media with different concentrations of NaCl, there was no obvious change in its intracellular Na+ contents , it accumulated K+, glutamic acid and betaine as osmoprotectants. Its intracellular contents of K+,glutamic acid and betaine increased by 1.9,2.4 and 13.6 times, respectively, when the concentration of NaCl increased from 0.1 mol/L to 2mol/L.

Acetamides ; pharmacology ; Animals ; Bridged-Ring Compounds ; poisoning ; Disease Models, Animal ; Female ; Male ; Rats ; Rats, Sprague-Dawley ; Troponin I ; blood

HIV-1 integrase (IN) is a key enzyme for the viral replication. The protein-protein interaction (PPI) between HIV-1 IN and a cellular cofactor lens epithelium-derived growth factor (LEDGF/p75) is a validated target for anti-HIV drug discovery. In order to build the platform for screening inhibitor against PPI between IN and LEDGF/p75, the vector containing the LEDGF/p75 protein cDNA was constructed and expressed in Escherichia coli and the function of the LEDGF/p75 protein was assayed. The LGDGF/p75 encoding gene optimized according to the preference codon usage of E. coli, was synthesized and cloned into the expression vector pGEX-4T-1 to form a recombined plasmid, then transformed into host cell E. coli BL21 (DE3). The recombined clones were identified and confirmed by BamH I/Sal I digestion and sequencing, the successfully recombined plasmid in the host cell was induced by IPTG and the condition of the expression was optimized. The expressed protein was purified by the Ni2+ affinity chromatography column and SDS-PAGE was used to analyze the molecular weight and specificity. In addition, ELISA assay was used to analyze the function of the recombinant protein. The recombinant LGDGF/p75 was soluble, and expressed highly and stably in E. coli. The protein was proved to enhance HIV-1 IN strand transfer activity in vitro by ELISA. It will be helpful to build the platform of screening inhibitors against PPI between IN and LEDGF/p75.
Cloning, Molecular ; Escherichia coli ; metabolism ; HIV Integrase ; metabolism ; HIV-1 ; physiology ; Humans ; Intercellular Signaling Peptides and Proteins ; biosynthesis ; Protein Binding ; Virus Replication

The pharmacoclynamics of Kuntaian was studied. The results showed that it exerts an inhibiting effect on normal uterine smooth muscle in a dosedependent manner. It can counteract the violent uterine contraction induced by pituitrin, promote the growth of uterus of young mice, dilate the capilaries and accelerate blood circulation. It also exerts analgesic antiphlogistic antioneotie effects, suggesting that this drug may be applicable to the treatment of dysmanorrhea, irregular menstruation and pelvic infection.

In order to mutate a carbofuran degrading strain CFDS-1,transposon of pSC123 was introduced into the genomic DNA of strain CFDS-1 by conjugation with E.coli DH5? (pSC123) as donor and strain CFDS-1 as recipient,6 mutants which lost carbofuran degrading ability were obtained with a kanamycin resistant gene in the middle of transposon and the disappearance of a red compound during the degrading of carbofuran as preliminary selecting marks,they were designated as CFDS-M1～CFDS-M6. UV scanning and GS assay results also proved their mutations. PCR was carried out respectively with primers designed according to the sequence of transposon and genomic DNA of 6 mutants as templates,restriction analysis of PCR products showed that the mutation of carbofuran degrading genes of these mutants was caused by transposon insertion.

OBJECTIVETo study the clinical effect of combination of kurarinone (KR) and interferon a-lb (IFNalpha-1b) in treating chronic hepatitis B.

METHODSOne hundred and forty-six patients with chronic hepatitis B were randomized into four groups. Under the basic conventional treatment, additional KR combined IFNa-lb was given to Group A, IFNa-lb to Group B and KR to Group C while none was given to Group D. The therapeutic course for them was 6 months and succeeded with 6 months of follow-up. Changes in liver histology, expression of transforming growth factor beta (TGF-beta) in liver tissue, and serum levels of TGF-beta, hyaluronic acid (HA), laminin (LN), collagen type IV (IVC), precollagen III (PCIII), as well as the negative conversion rate of HBeAg and HBV DNA, and normalization rate of alanine transaminase (ALT) before and after treatment were observed by immuno chemical method, RIA and ELISA.

RESULTSAfter treatment, in Group A, the scores of liver fibrosis and all the above-mentioned indexes significantly lowered, as compared with those in Group B and C, the difference was significant (P<0.05 or P<0.01). The negative conversion rate of TGF-beta in liver tissue was in accordance with the dynamic change of liver fibrosis. Comparison between Group B and C in those aspects showed insignificant difference (P >0.05).

CONCLUSIONKR combined with IFNalpha-1b shows better effect in treating chronic hepatitis B than that of using either of the two alone.

Adult ; DNA, Viral ; blood ; Drug Therapy, Combination ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Flavonoids ; therapeutic use ; Hepatitis B e Antigens ; blood ; Hepatitis B virus ; isolation & purification ; Hepatitis B, Chronic ; drug therapy ; Humans ; Interferon-alpha ; therapeutic use ; Male ; Middle Aged ; Phytotherapy

Objective To investigate the value in the diagnosis of tumor of the stomach by hypotonic water filling method com-bined with CT multi planar reconstruction (MPR).Methods CT image data of 21 5 cases with gastric tumor confirmed by operation and pathology in our hospital were analysed retrospectively.Conventional CT enhanced scan was obtained in patients with the stom-ach hypotonic water filling condition,and MPR CT characteristics of lesions were observed.Results In the 21 5 cases of gastric be-nign or malignant lesions,MPR showed 5 pathological types in 210 cases.In the conventional CT examination,the tumor diagnosis rate had obvious improvement in different gastric parts and types of the stomach tumors through CT MPR.Conclusion There is high detection rate in the diagnosis of gastric tumors using hypotonic water filling method with MPR,which can accurately display invasion and metastasis,and reduce the misdiagnosis and missed diagnosis in gastric tumor.

Objective To evaluate the image quality of 64-multi detector computed tomography (MDCT)and the clinical accuracy in detecting coronary artery lesions.Methods One hundred and five patients were studied by MDCT.The results were compared with invasive coronary angiography(ICA). Patients were excluded for atrial fibrillation,but not for high heart rate,coronary calcification,or obesity. MDCT was analyzed with regard to image quality and presence of coronary artery lesions.Results The data evaluation of the image quality was based on a total of 1365 segments(13 coronary segments for each patient),of which 1144 segments were considered to have diagnostic image quality,but 221 segments (16.2%)could not be sufficiently evaluated because of severe calcifications(153 segments)and motion artifacts(68 segments).The median calcium score[Agatston score equivalent(ASE)]was 154(range 0—1983).87 of the 105 patients had an ASE of less than 1,000[median 105(range 0—994)],and 18 patients had an ASE greater than 1000[median 1477(range 1115—1983)].For detecting lesions with 50% or greater narrowing(without any exclusion criteria),the overall sensitivity,specificity,positive predictive value,and negative predictive value were 85.7%,97.9%,93.0%,and 95.5%,respectively. When limiting the number of patients to those with a calcium score of less than 1000 ASE,the threshold- corrected sensitivity for lesions with 50% or greater narrowing was 96.0%;specificity,98.9%;positive predictive value,95.3%;and negative predictive value,99.0%.Conclusion Our results indicate high quantitative and qualitative diagnostic accuracy of 64-slice MSCT in comparison to QCA in a broad spectrum of patients.

Objective:To investigate the distribution of neural cholinesterase and elastic and collagen fibers in rat pancreatic tissues.Methods:Rats were decapitated and the pancreatic tissues were harvested,frozen and made into 8?m serial sections.A combined-staining method(K-R-VB-P-PA method,a combination of Karnovsky-Roots and Victoria blue-Ponceau-Picric acid) was used for staining and the distribution of neural cholinesterase and elastic and collagen fibers was observed.Results:The results showed that the neural cholinesterase was brown.the elastic fibers were cyan.the collagen fibers were light red.and the smooth muscle was yellow in the pancreatic acinar,islet of Langerhans and blood vessels.Conclusion:K-R-VB-P-PA combined staining method can demonstrate the distribution of neural cholinesterase,elastic and collagen fibers and smooth muscle fibers in the pancreatic tissues synchronously.