Objective To express,purify and identify recombinant hepatitis E virus(HEV) pb166-GST fusion protein using GST gene fusion system and investigate its potential role in researching Hepatitis E diagnostic antigen field.Methods The recombinant E.coli BL21 performed by our own laboratory was used to induce the HEV pb166-GST expression with IPTG.The products were purified by BD Biosience GST purifying system.The specific expression was identified by SDS-PAGE and Western blot.The experiment conditions and results were described and analysed.Results The resolved HEV pb166-GST fusion protein on SDS-PAGE showed a major band at position of 43 kD.The expressed proteins had a single expected band after purify and the protein was recognized by anti-GST antibody on PVDF membrane.Conclusion The recombinant HEV pb166-GST fusion protein is expressed in recombinant E.coli BL21 efficiently in this way,and might be used as a candidate for diagnostic antigen of HEV.

Objective To investigate the curative effect of acute severe brain injury complicated ARDS, Methods 31 patients who had acute severe brain injury complicated ARDS were divided into two groups:A group was early discovery of ARDS and given treatment.B group was late discovery of ARDS and treated late.Then the curative effects were compared.Results A group was significantly higher than B group in blood gas analysis(P

BACKGROUNDTuberculosis remains the leading cause of human death. Currently, Bacillus Calmette-Guérin (BCG) is the only available vaccine against tuberculosis but its efficacy is highly variable. Thus, developing new tuberculosis vaccines becomes an urgent task. In this study, we evaluated in BALB/c mice the humoral and cellular immune responses of recombinant BCG expressing the antigen ESAT-6 from Mycobacterium tuberculosis.

METHODSEscherichia coli-BCG shuttle plasmid named pDE22-esat-6 was constructed by inserting the BamHI/EcoRI digested esat-6 gene PCR product into the similarly digested parental plasmid pDE22. BCG cells were transformed with pDE22-esat-6, which was named recombinant BCG (rBCG). BALB/c mice were immunized subcutaneously on the back with 100 microl normal saline containing 10(6) CFU of BCG or rBCG. They were sacrificed after 4 weeks to detect their humoral and cellular responses.

RESULTSThere was no any significant differences in the growth characteristics between the conventional BCG and rBCG. In immunized mice, the IgG antibody titres of rBCG group were as high as 1:8000, which was significantly higher than that in BCG group (1:1400, P < 0.05). The elicited IFN-gamma level of rBCG group was (1993 +/- 106) pg/ml, which was also significantly higher than that in BCG group ((1463 +/- 105) pg/ml, P < 0.05). The splenocyte proliferation index of rBCG group reached 4.34 +/- 0.31, which was higher than that of BCG group (3.79 +/- 0.24, P < 0.05).

CONCLUSIONrBCG secreted expressing antigen ESAT-6 stimulated stronger humoral and cellular immune responses than BCG did, and, therefore may be the better vaccine against mycobacterium tuberculosis.

Animals ; Antigens, Bacterial ; genetics ; immunology ; BCG Vaccine ; immunology ; Bacterial Proteins ; genetics ; immunology ; Interferon-gamma ; biosynthesis ; Lymphocyte Activation ; Male ; Mice ; Mice, Inbred BALB C ; Mycobacterium tuberculosis ; immunology ; Recombinant Proteins ; immunology ; Vaccines, Synthetic ; immunology

To study the constituents from the chloroform extract of the roots of Lasianthus acuminatissimus Merr., various chromatographic techniques were used to separate and purify the constituents. The structure was established on the basis of ID, 2D NMR and HRMS spectroscopic analysis. A new compound was isolated and identified, which was 3, 8-dihydroxy-1-methoxy-2-methoxymethyl-9,10-anthraquinone (I). Compound I is a new anthraquinone, namely lasianthurin.
Anthraquinones ; chemistry ; isolation & purification ; Chromatography, Thin Layer ; Magnetic Resonance Spectroscopy ; Molecular Conformation ; Molecular Structure ; Plant Roots ; chemistry ; Plants, Medicinal ; chemistry ; Rubiaceae ; chemistry ; Spectrometry, Mass, Electrospray Ionization

OBJECTIVETo investigate the inhibition of COX-2 gene expression and its effects on malignant proliferation of human lung adenocarcinoma A549 cells after interfering at different target sites in vitro.

METHODSThe 3rd, 7th and 10th exon of COX-2 were selected as the targets and three COX-2 siRNA expression vectors with human U6 promoter were constructed. Three siRNA expression vectors and two vacant vectors were transfected into A549 cells expressing COX-2 with lipofectamine, respectively. The transfected cell strains were constructed and the change of COX-2 expression levels was examined by Western blot and RT-PCR. The effects on the proliferation of A549 cells after interfering at different target sites were studied by cell growth curve and colony formation assay in vitro.

RESULTSThe three siRNAs and U6 promoter were validated by PCR, restriction endonuclease digestion, DNA sequencing and BLAST alignment, and cloned into the pEGFP vector. The cell strains transfected were named as A549-3, A549-7, A549-10, A549-p and A549-pU6, respectively. A549-p cells showed expression of GFP and A549-3, A549-7, A549-10, A549-p and A549-pU6 cells did not show at 24, 48 and 72 hours after transfection. The results of RT-PCR and Western blot showed an inhibition of COX-2 expression after interfering at three target sites (3rd, 7th and 10th exons). In contrast to A549 cells, the levels of COX-2 mRNA of A549-3, A549-7 and A549-10 cells were reduced by 10.6%, 33.4% and 61.2%, respectively. The levels of COX-2 protein of A549-3, A549-7 and A549-10 cells were reduced by 26.7%, 44.7% and 56.2%, respectively. The results of cell growth curve and colony formation assay showed a slowing down of the growth of A549-10 cells and reduction of their colony formation rate. The other two targets had no apparent effect on the growth of A549 cells.

CONCLUSIONThere is a significant inhibiting effect of RNA interference on the malignant proliferation of A549 cells in vitro, and the most striking effect can be seen when the 10th exon of COX-2 is taken as the interference target.

Adenocarcinoma ; metabolism ; pathology ; Cell Line, Tumor ; Cell Proliferation ; Cyclooxygenase 2 ; genetics ; metabolism ; physiology ; Exons ; Genetic Vectors ; Humans ; Lung Neoplasms ; metabolism ; pathology ; Promoter Regions, Genetic ; RNA Interference ; RNA, Messenger ; metabolism ; RNA, Small Interfering ; genetics ; Transfection

In order to prove the feasibility of preparation of the drug-incorporated stent by immersing stent wires in the monoclonal antibody (mAb) solution, fluorescence stain and image analysis were used to evaluate the L-PLA-coated stent. Absorption was measured using a radioisotope technique after preparing the mAb-incorporated stent, and the absorption curve was determined from the absorption data. In an in vitro perfusion circuit, the antibody was eluted from the stent matrices, and the related influence factors were evaluated based on the release data.
Absorption ; Alloys ; chemistry ; Antibodies, Monoclonal ; chemistry ; immunology ; Drug-Eluting Stents ; Graft Occlusion, Vascular ; immunology ; prevention & control ; Humans ; Lactic Acid ; chemistry ; Platelet Aggregation Inhibitors ; chemistry ; immunology ; Platelet Glycoprotein GPIIb-IIIa Complex ; immunology ; Polymers ; analysis ; chemistry ; Time Factors

Researches on drug-eluting stents are now focusing on three main aspects: the stent materials, the coating matrix material and the selection, adhesion and controlled release of the biological agents. The current development progresses of the coating materials, their characteristics, and the coating method for metallic stents are reviewed in this paper.
Coated Materials, Biocompatible ; Coronary Restenosis ; prevention & control ; Drug Carriers ; Drug Delivery Systems ; Humans ; Polymers ; chemistry ; Stents

Objective To describe the current situation of diarrhea in children under five years old in Nepal and to explore its influencing factors. Methods Data were collected from the open-access database, Nepal Demographic and Health Surveys in 2006, 2011 and 2016. Chi-square ( 2) and Wilcoxon rank sum test were used to compare difference of potential risk factors between groups with and without diarrhea. Multiple Logistic regression model was adopted to identify significant influencing factors on diarrhea in children under five years old in Nepal. Results In 2006, 2011 and 2016, the incidence of diarrhea children under five in Nepal was 12.3%, 13.3% and 6.8%, respectively. Univariate analysis of the potential influencing factors showed that there were significant differences in the gender, water source, toilet facilities and fuel type, age of children, age of mother when she gave birth to the child and education years of mother and children with and without diarrhea (all P＜0.05). Multiple analysis revealed that improved toilet facilities (OR=0.874, 95% CI: 0.769-0.994, P=0.041) and the age of children(OR=0.613, 95% CI: 0.580-0.645, P＜0.001) were protective factors of childhood diarrhea, and the risk of boys was higher than that of girls(OR=1.277, 95% CI: 1.147-1.423, P＜0.001). Conclusions From 2006 to 2016, the incidence of diarrhea in children under five years old in Nepal was decreasing. Toilet facilities, age of children and gender of children were identified as the influencing factors of childhood diarrhea.

OBJECTIVETo explore the clinical and laboratory characteristics of two myelodysplastic syndromes (MDS) patients with double isochromosome 20q- anomaly.

METHODSBone marrow cell chromosome preparations were made with both direct method and short-term culture. Karyotype analysis was performed by R-banding technique, and dual-color FISH (fluorescence in situ hybridization) by using a 20q telomeric probe and a sequence-specific probe for 20q12.

RESULTSThe clinical and hematological findings were comparable with diagnosis of MDS. Karyotype analysis showed that both patients had double isochromosome 20q- anomaly: case 1 is 46, XX, der(20)? i(20q-) [6]/46, idem, der (6) i (6p) [1]/47, idem, +der (20)? i (20q-) [3]/47, idem, der(6)i (6p), +der(20)? i (20q-) [20]; case 2 is 45, XY, -7, der (20)? i (20q-) [17]/46, idem, +der(20) ? i(20q-) [3]. Two derivative chromosomes 20 were proved 20q isochromosomes with interstitial deletions by dual-color FISH in one patient.

CONCLUSIONSDouble isochromosome 20q- anomaly is a rare recurrent karyotype abnormality in MDS, and signals a poor prognosis.

Chromosome Banding ; Chromosomes, Human, Pair 20 ; genetics ; Female ; Humans ; In Situ Hybridization, Fluorescence ; Isochromosomes ; Karyotyping ; Male ; Middle Aged ; Myelodysplastic Syndromes ; genetics

OBJECTIVES(1) To evaluate the prevalence, phenotypes and suppressive function of CD4+CD25+ regulatory T cells (Tregs) among the in peripheral blood mononuclear cells (PBMCs) and tumor-infiltration lymphocytes (TILs) from hepatocellular carcinoma (HCC) patients and patients with chronic hepatitis B. (2) To investigate the correlation between the frequency of CD4+CD25+ Tregs and clinical characteristics of HCC patients.

METHODSPBMCs and TILs in 18 HCC patients, 10 chronic hepatitis B (CHB) patients and 15 healthy donors were evaluated for the phenotypes of CD4+CD25+ Tregs and the proportion of CD4+CD25+ Tregs as a percentage of the total CD4+ cells, by flow cytometric analysis with three or four color staining. The relationship between the frequency of CD4+CD25+ Tregs and tumor TNM stages was analyzed. The CD4+CD25+ Tregs and CD4+CD25- T cells were isolated from PBMC of HCC patients and donors. The suppressive function of CD4+CD25+ Tregs was analyzed.

RESULTSThe percentages of CD4+CD25+ Tregs of the HCC patients (6.38% +/- 6.30%) and CHB patients (4.29% +/- 1.82%) were significantly higher than those of the healthy donors (1.58% +/- 0.55%, P less than 0.01). Among the TILs, the percentage of CD4+CD25+ Tregs was higher (t = 4.39, P < 0.01). There were significant differences in the prevalence of CD4+CD25+ Tregs in early and advanced stage HCCs (stage II vs. III, P less than 0.05; stage II vs. IV P < 0.01). The proliferative capacity of CD4+CD25- T cells was inhibited by the presence of CD4+CD25+ T cells in a dose-dependent manner where the level of suppression was correlated to the ratio of the two-cell populations.

CONCLUSIONThese results suggest that the increase in frequency of CD4+CD25+ Tregs might play a role in the suppression of the immune response against HCC, which may contribute to the HCC cells that escaped from immunological surveillance.

Adult ; Aged ; Carcinoma, Hepatocellular ; metabolism ; Female ; Humans ; Interleukin-2 Receptor alpha Subunit ; Liver Neoplasms ; metabolism ; Male ; Middle Aged ; T-Lymphocytes, Regulatory ; immunology ; Young Adult