Nucleic acid sequence-based amplification(NASBA) is a sensitive,isothermal,transcription-based amplification system specifically designed for the detection of RNA targets,which could amplify templete RNA in 2h under isothermal condition at about 42?C and without any special equipment.NASBA is now widely applicated in diagnosis of many pathogenic microorganism.It is mainly about principles and applications of NASBA in viral diagnosis.

Acute Kidney Injury ; therapy ; Child ; Hemofiltration ; methods ; Humans

Objective:To explore the influence of ischemia/reperfusion (anoxia/reoxygenation)[FK(W16*2。142mmZQ1mm]on immunofunction of endothelial cells(ECs) and effect of intervention with 2-hydroxymethyl-3,5,6-trimethylpyrazine on it.Methods:Model of ECs induced by anoxia/reoxygenation was established to mimic ECs ischemia/reperfusion injury in vivo with human umbilical vein endothelial cell line ECV304.Then 2-hydroxymethyl-3,5,6-trimethylpyraxine was used to intervene the anoxia/reoxygenation process.Nuclear transcriptional factor-?B(NF-?B) was exhibited by fluorescent staining,HLA-ABC,HLA-DR,CD86 and CD54 were detected by flow cytometry.Mixed endothelial cell-lymphocyte reaction(MELR) was conducted to examine the proliferation of lymphocyte,production of IL-2 and percentage of apoptotic lymphocyte.Results:Anoxia/reoxygenation made the ECV304 cell became round,shrunk and abscised,with increased plasma NF-?B,and changed from positive cytoplasm to positive nucleus.HLA-ABC,HLA-DR and CD86 on surface of cells increased but CD54 showed unchanged.MELR showed the incorporation of ~3H-TdR and production of IL-2 increased significantly and the percentage of apoptotic lymphocyte decreased.After 2-hydroxymethyl-3,5,6-trimethylpyrazine intervention,the ECV304 cell shapes recovered,NF-?B expression didn’t down-regulated,but the percentage of positive cells decreased,changes to positve dominant.Besides,reversal changes were shown in other parameters.Conclusion:Anoxia/reoxygenation influences some important immune related molecules in ECV304 cells.2-hydroxymethyl-3,5,6-trimethylpyrazine could antagonize these influences to maintain the immune function of endothelial cells in a relative normal manner.

To reduce the serum clearance of interferon alpha2b, a chimeric gene encoding an human serum albumin(HSA)--human interferon alpha2b(IFNalpha2b) fusion protein was overexpressed in Pichia pastoris. After fermentation in a 5L bioreactor, the fusion protein, capable of cross-reacting with anti-IFN alpha and anti-HSA antibody, was purified from the culture of the recombinant yeast by ultrafiltration, blue Sepharose affinity, phenyl hydrophobic interaction and Q ion exchange chromatography. Its IFNa2b moiety exhibits antiviral activity similar to that of recombinant human IFNa2b. In Cynomolgus monkeys model, The fusion protein was detectable in plasma, even 336h after a single does of 90 microg/kg injection intravenously or subcutaneously. The elimination phase half-life of the fusion protein was 101h after intravenous injection and 68.2h after subcutaneous injection. Its Subcutaneous bioavailability was 67.9%. The enhanced pharmacokinetics of interferon a2b fused to human serum albumin suggest its promissing application in clinic medicine.
Animals ; Bioreactors ; microbiology ; Fermentation ; Humans ; Interferon-alpha ; biosynthesis ; genetics ; Macaca fascicularis ; Pichia ; genetics ; metabolism ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; pharmacokinetics ; Recombinant Proteins ; Serum Albumin ; biosynthesis ; genetics

Objective: To study the effect of arsenic trioxide (As2O3) on human pancreatic cancer cell proliferation and apoptosis (mainly early stage) in vitro. Methods: SW1990 cells line were trea ted with As2O3 at different concentration. Cell proliferation was evaluated by MTT and apoptosis by Annexin-Ⅴ-fluostaining, electron-microscopy, flow cytometry and immunocytochemical staining of Bcl-2 and Bax. Results: As2O3 and cisplatin had the same cytotoxity on SW1990. The cytotoxic effe ct on tumor cell was produced by induction of apoptosis. Twelve hours after cult ure with 10 μg/ml As2O3, much more SW1990 cells went into apoptosis than t he control. The apoptosis rate reached 24% after 48 h with the similar concentra tion of As2O3. Immunohistochemical study revealed that the expression of Bcl -2 was decreased after treated with As2O3. Conclusion: As 2O3 can depress the proliferation of SW1990 in vitro, mainly through the i nduction of apoptosis, and it is a potential agent for pancreatic cancer chemoth erapy.

Receptors play a crucial role in determining the host specificity and tissue tropism of virus. Foot-and-mouth disease virus(FMDV)has been showed to use four integrins, ?v?1, ?v?3, ?v?6 and ?v?8 as receptors to initiate infection and ?v?6 functions as the major receptor.The cDNA encoding bactrian camel integrin ?6 from the lung tissue was cloned and sequenced. The 2367bp cDNA of bactrian camel integrin ?6 encodes a polypeptide of 788 amino acids consisting of a 26-residue putative signal peptide, a 681-residue ectodomain with 8 potential N-linked glycosylation sites and 58 cysteine residues, a 29-residue transmembrane domain, and a 52-residue cytoplasmic domain with a NPLY motif and 1 potential N-linked glycosylation site. The nucleotide sequence similarity of integrin ?6 between bactrian camel and cattle, pig, sheep, human, mouse, Norway rat is 91.1%、91.8%、90.6%、90.5%、83.7%、84.1%, and the amino acid sequence similarity is 94.3%、93.4%、93.4%、93.7%、88.7%、88.6%, respectively. The bactrian camel ?6 gene exhibited the higher sequence homology with the ?6 gene of cattle, pig and sheep, indicating their close genetic relationships. It is possible that host tropism of FMDV may related to divergence in ?6 receptors among different species.

Animals ; Antineoplastic Agents ; administration & dosage ; Electroporation ; methods ; Interleukin-2 ; blood ; Interleukin-6 ; blood ; Liver Neoplasms, Experimental ; immunology ; mortality ; pathology ; therapy ; Male ; Neoplasm Transplantation ; Rats ; Rats, Sprague-Dawley

Stem cell factor is an important hematopoietic growth factor. In this study, the human stem cell factor was produced by recombinant E. coli, and the structure and biological activity of the recombinant stem cell factor(rhSCF) was studied. It was indicated that the rhSCF was a uncovalent dimer in phosphate buffer,and had the correct mass spectra, mass peptides spectra, composition of amino acid, N-terminal sequernce, C-terminal sequence and intrachain disulfide linkages, rhSCF alone or synergy with rhG-CSF could mobilze hematopoietic progenitors to blood in monkey.
Animals ; Cell Differentiation ; drug effects ; Cells, Cultured ; Chromatography, High Pressure Liquid ; Haplorhini ; Hematopoietic Stem Cells ; cytology ; drug effects ; Humans ; Recombinant Proteins ; chemistry ; genetics ; metabolism ; pharmacology ; Sequence Analysis, Protein ; Spectrometry, Mass, Fast Atom Bombardment ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Stem Cell Factor ; chemistry ; genetics ; metabolism ; pharmacology

BACKGROUND:Kawasaki disease (KD) is a common cause of acquired heart disease in children. Recent studies have focused on the biochemical markers of the myocardium, their high sensitivity and specificity and significance in the diagnosis of KD. This study aimed to determine the serum level of brain natriuretic peptide (BNP) and its relation with the heart function of children with KD and to explore its clinical value in diagnosis of KD. METHODS:Forty-three KD children, aged from 5 months to 8 years (mean 2.3±0.6 years ), were admitted to Qingdao Children's Hospital from February 2007 to April 2009. Among them 27 were male, and 16 female. The 43 patients served as a KD group. Patients with myocarditis, cardiomyopathy, congenital heart disease and other primary heart diseases were excluded. Thirty healthy children, aged from 3 months to 15 years (mean 2.5±0.8 years) or 17 males and 13 females served as a control group. There were no significant differences in age and gender between the two groups (P>0.05). In the KD group, ELISA was used to measure the levels of serum BNP in acute and convalescent stages;and in the control group, the levels of serum BNP were measured once randomly. Left ventricular ejection fraction (LVEF), left ventricular shorten fraction (LVSF), cardiac index (CI) and left ventricular inflow velocity through the mitral annulus (including E-velocity and A-velocity) were measured by two-dimensional echocardiography in the acute and convalescent stages in the KD group. All data were expressed as mean±SD. The methods of analysis included Student's t test and the linear regression analysis test. P<0.05 was considered statistically significant. RESULTS:The level of serum BNP in the acute stage (517.26±213.40) ng/ml was significantly higher than that in the convalescent stage (91.56±47.97) ng/ml in the control group (37.55±7.56) ng/ml (P<0.01). The levels of LVEF, LVSF and CI in the acute stage were significantly lower than those in the convalescent stage (P<0.05), but the E/A level was not significantly different between the acute and convalescent stages (P>0.05). Linear regression analysis showed that the BNP level was negatively correlated with the levels of LVEF, LVSF and CI(r=-0.63, -0.52, -0.53, P<0.05) , but not significantly correlated with the E/A level (r=-0.18, P>0.05). CONCLUSION:The levels of serum BNP are significantly increased in KD patients, and are negatively correlated with the levels of LVEF, LVSF, and CI. The detection of serum BNP level is of clinical significance in the diagnosis of KD.

Objective To discuss the treatment of ureteropelvic junction obstruction by laparoscopic pyeloplasty. Methods A retrospective review of consecutive laparoscopic pyeloplasty in 102 patients between September 2001 and December 2007 was performed. The ureterpelvic junction was dissected and the obstruction portion was excised. Anastomosis was then performed through the ureter and the renal pelvis walls with a stent. Results The mean operating time was 120 min and the average blood loss was 80ml. No major complication occurred intraoperative. The drainage was removed in 3-10 days. The average hospital stay was 8.5 days. The stent was kept for 30-60 days. IVU and B ultrasound examination revealed that the hydronephrosis alleviated during the follow-up and no anastomosis stricture occurred. Conclusions Laparoscopic dismembered pyeloplasty could provide lower morbidity, shorter hospital stay, and faster convalescence. It could be an effective treatment for ureteropelvic junction obstruction.