Objective To construct and express the recombinant plasmid pET28α-Sj26GST-Sj32 of Schistosoma japonicum(Sj) in Escherichia coli BL21 (DE3). Methods Total RNA was extracted from Sj adult worms by ultrasound-breaking, Sj26GST and Sj32 antigen gene was respectively amplified by RT-PCR from the total RNA; Sj26GST-Sj32 fusion gene obtained with gene splicing by overlap extension(SOEing) was cloned into prokaryotic expression plasmid pET28α and transformed into Escherichia coli BL2 (DE3) to construct pET28α-Sj26GST-Sj32;BL21 (pET28α-Sj26GST-Sj32) was induced with isopropyl-β-D-thiogalactopyranosid (IPTG), and the expressed products were analyzed and identified by sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE)and Western blotting. Results The 1991 bp Sj26GST-Sj32 fusion gene was successfully amplified by gene SOEing and cloned into pET28α by restriction analysis and PCR identification, the recombinant plasmid pET28α-Sj26GST-Sj32 was successfully constructed; the relative molecular mass of the expressed recombinant protein was approximately 69 × 103 by SDS-PAGE, and the amount of the expressed protein was 25% of the total bacterial proteins; the fusion protein could be recognized by sera from rabbits infected with Sj by Western blotting.Conclusions The recombinant plasmid pET28α-Sj26GST-Sj32 is successfully constructed and highly expressed in Escherichia coli in fused form with His-tag, and the expressed fusion protein shows specific antigenicity.

Objective To construct and express the recombinant plasmid pET32α-Sj26GST of Schistosoma japonicum(sj)in Escherichia coli(E.coli)B121(DE3).Methods The total RNA was extracted from sj adult worms by ultrasound-breaking,Sj26GST antigen gene was amplified by RT-PCR from the total RNA,then cloned into prokaryotic expression plasmid pET32α(+) and transformed into E.coli B12(DE3)to construct pET32α-Sj26GST;BL21(pET32α-Sj26GST)WaS induced with isopropyl-β-D-thiogalactopyranosid(IPTG),and the expressed products were analyzed and identified by SDS-PAGE and Western blot.Results The 676 bp Sj26GST gene was successfully amplified by RT-PCR and cloned into pET32α(+)by restriction analysis and PCR identification,the recombinant plasmid pET32α-Sj26GST was successfully constructed;the relative molecular mass of the expressed recombinant protein was approximately 49×103 by SDS-PAGE,and the amount of the expressed protein was 24%of the total bacterial proteins;the fusion protein could be recognized by sera from rabbits infected with sj by Western blot.Conclusions The recombinant plasmid pET32α-Sj26GST is successfully constructed and highly expressed in E.coli in fused form with Trx-tag and His-tag,and the expressed fusion protein shows specific antigenicity.

This study is to investigate if madecassoside can protect against myocardial reperfusion injury in rabbit heart in vivo. The ischemia reperfusion model was established. Left ventricular function and ECG were monitored at the ischemia and reperfusion period. The infarct areas were expressed as percentage. The levels of LDH, CK, MDA and SOD were measured and C-reactive protein (CRP) in serum was measured by ELISA kit. Cardiomyocyte apoptosis were measured by TUNEL staining. A monoclonal rabbit anti-goat Bcl-2 proteins as primary antibody was used for Bcl-2 immunohistochemical staining. Treatment with madecassoside (3.2, 1.6 and 0.8 mg x kg(-1)) i.v. during ischemia reperfusion injury attenuated myocardial damage, that is, characteristic of decreasing infarct size, decreasing LDH and CK release. Activities of SOD were diminished and MDA level increased obviously in control group whereas pretreatment with madecassoside significantly blunted the decrease of SOD activity, markedly reduced the levels of MDA, CRP and cardiomyocyte apoptosis, and upregulated the expression of Bcl-2. Madecassoside has the protective effect against myocardial ischemia reperfusion injury, and effects of anti-lipid peroxidation, enhancement of SOD activity, anti-inflammation and anti-apoptosis.
Animals ; Apoptosis ; drug effects ; C-Reactive Protein ; metabolism ; Cardiotonic Agents ; isolation & purification ; pharmacology ; Centella ; chemistry ; Creatine Kinase ; blood ; Electrocardiography ; L-Lactate Dehydrogenase ; blood ; Lipid Peroxidation ; drug effects ; Male ; Malondialdehyde ; blood ; Myocardial Reperfusion Injury ; metabolism ; pathology ; Myocardium ; metabolism ; pathology ; Myocytes, Cardiac ; pathology ; Plants, Medicinal ; chemistry ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Rabbits ; Random Allocation ; Superoxide Dismutase ; blood ; Triterpenes ; isolation & purification ; pharmacology

Professor LUO Cai-gui's experience of acupuncture at acupoint "Baliao" with twisting manipulation for treatment of low back pain is introduced. This method has significant efficacy on improving low back pain and numbness of lower extremities, which is characterized with short-time manipulation, quick de-qi and long effective time. The acupuncture methods, manipulations, precautions, etc. are elaborated in details. A typical case is added.
Acupuncture Points ; Acupuncture Therapy ; instrumentation ; methods ; Humans ; Low Back Pain ; therapy ; Male ; Middle Aged

OBJECTIVETo investigate the growth status of children under 7 years in Wuzhong City, Ningxia Hui Autonomous Region, China and its influential factors, and to provide a basis for related intervention measures.

METHODSChildren under 7 years were selected from two county-level districts in Wuzhong by stratified cluster sampling, and their growth status were evaluated by the Z score method.

RESULTSThe prevalence rates of growth retardation, underweight, and wasting were 12.58%, 5.71%, and 5.55% respectively. The height-for-age Z score, weight-for-age Z score, and weight-for-height Z scores were -0.26±2.50, 0.29±4.54. and 0.65±3.02 respectively. There were significant differences in the prevalence rate of wasting among children of different ethnic groups (P<0.05); also, there were significant differences in the prevalence rates of growth retardation and underweight among children from different regions and with different age (P<0.05). The main influential factors for growth retardation were region (OR=0.369, P<0.001), ethnic groups (OR=1.694, P=0.027), and age (OR=1.143, P=0.002). The main influential factors for underweight were region (OR=0.453, P=0.001) and age (OR=1.204，P=0.002). The main influential factor for wasting was nation (OR=1.735, P=0.024).

CONCLUSIONSIn Wuzhong, children under 7 years have poor growth status, which are related to ethnic groups, region, and age.

Body Height ; Body Weight ; Child ; Child Development ; Child, Preschool ; China ; epidemiology ; Diet ; Female ; Growth Disorders ; epidemiology ; Humans ; Infant ; Infant, Newborn ; Logistic Models ; Male

Microarray analysis revealed that the expression of ferric reductase (FRP1) can be regulated by the Riml01 protein. In order to find new transcriptional regulatory element in the promoter of FRP1, we analyzed the 1000 bp sequence upstream of ATG to find 2 potential Riml01p binding sites. We generated site-specific mutations in each of the two sites and fused these mutated promoters to LacZ. Then the promoter-LacZ fusion construct was recombinant into wild type and riml01-/- strains for beta-galactosidase assay. The results revealed that the FRP1 was up-regulated in alkaline pH and this was caused by iron starvation. The -650 site, not the -160 site, had an important role in FRP1 Riml01p-dependent expression. We conclude that Riml01p may interact with the -650 binding site of the promoter to regulate the FRP1 expression.
Candida albicans ; enzymology ; genetics ; DNA-Binding Proteins ; genetics ; FMN Reductase ; genetics ; Fungal Proteins ; genetics ; Mutagenesis, Site-Directed ; Promoter Regions, Genetic ; genetics

Adult ; Blood Donors ; China ; epidemiology ; Female ; Genotype ; HIV Infections ; complications ; epidemiology ; virology ; Hepacivirus ; genetics ; Hepatitis C ; complications ; epidemiology ; virology ; Humans ; Male ; Middle Aged

OBJECTIVE: To determine the effect of tanshinone IIA on the growth and apoptosis in human hepatoma cell line HepG2. METHODS: The human hepatoma cell line HepG2 was treated with tanshinone IIA at various concentrations for 72 h. The inhibition of proliferation was measured by MTT assay and apoptosis-related alterations in morphology measured by cytochemical staining (HT33258). DNA fragmentation was evaluated by agarose gel electrophoresis. Apoptotic rate and cell arrest were quantified by flow cytometry (FCM). RESULTS: Tanshinone IIA inhibited the growth of HepG2 in a time- and dose- dependent manner. The semi-inhibitory concentration (IC50) value after the treatment with tanshinone IIA on HepG2 for 24, 48 and 72 h were 14.7, 7.4, and 3.9 microg/ mL, respectively. After the treatment with 0.5 - 10 microg/mL tanshinone IIA for 72 h, the formation of apoptotic bodies was observed. DNA ladder was shown in agarose gel electrophoresis, in addition to the cells treated by 1.0 microg/mL tanshinone IIA . The apoptotic rates at 0.5, 1.0, 2.0, 5.0, and 10.0 microg/mL for 72 h were 20.32%+/-2.16%, 28.0%+/-2.35%, 33.87%+/-3.43%, 46.73%+/-4.08% and 57.85%+/-3.74%, respectively, which were all significantly higher than those of the control group (P<0.05). CONCLUSION Tanshinone IIA can inhibit the proliferation of human hepatoma cell line HepG2 in a time- and dose- dependent manner, and the mechanism of growth inhibition of human hepatoma cells may be related to the induction of apoptosis.
Abietanes ; Antineoplastic Agents, Phytogenic ; pharmacology ; Apoptosis ; drug effects ; Carcinoma, Hepatocellular ; genetics ; pathology ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; DNA Fragmentation ; drug effects ; Dose-Response Relationship, Drug ; Drugs, Chinese Herbal ; pharmacology ; Flow Cytometry ; Humans ; Liver Neoplasms ; genetics ; pathology ; Microscopy, Fluorescence ; Phenanthrenes ; pharmacology ; Time Factors

OBJECTIVETo evaluate the expression of epidermal growth factor receptor (EGFR) gene copy number and the expression of ERCC1 and BRCA1 proteins in patients with non-small-cell lung cancer (NSCLC) and the correlation between them.

METHODSThe status of EGFR gene copy number was determined by in situ hybridization (FISH), and the expression of ERCC1 and BRCC1 proteins was examined by immunohistochemistry (IHC). The relationship of EGFR gene copy number with the expression of ERCC1 and BRCA1 and the clinical pathologic features were analyzed.

RESULTSFISH-positive EGFR expression was identified in 40 of 166 samples (24.1%). More FISH-positive EGFR in the female than male patients (31.9% vs. 18.6%, P = 0.048), and non-smoker than smoker (32.8% vs. 16.7%, P = 0.045). FISH-positive EGFR was not associated with age, pathological type, clinical stage and metestasis status (P > 0.05). The expression of ERCC1 protein was identified in 60 of 132 samples (45.5%). The expression of ERCC1 protein varied significantly in tumors of different pathological types (P = 0.046), but not associated with age, gender, clinical stage, metestatic status and smoking status (P > 0.05). The expression of BRCA1 protein was identified in 46 of 131 samples (35.1%). The expression of BRCA1 was not associated with age gender, pathological type, clinical stage, metestatic ststus and smoking status (P > 0.05). There was a moderate correlation between the expressions of ERCC1 and BRCA1 (r = 0.449, P < 0.001), but EGFR gene copy number was not correlated with the expression of ERCC1 or BRCA1 protein.

CONCLUSIONSFISH-positive EGFR expression is associated with gender and smoking status, but not correlated with the expression of ERCC1 and BRCA1 proteins. There is a moderate correlation between the expressions of ERCC1 and BRCA1.

Adult ; Aged ; Aged, 80 and over ; BRCA1 Protein ; metabolism ; Carcinoma, Non-Small-Cell Lung ; genetics ; metabolism ; pathology ; DNA-Binding Proteins ; metabolism ; Endonucleases ; metabolism ; Female ; Gene Dosage ; Gene Expression Regulation, Neoplastic ; Genes, erbB-1 ; Humans ; Immunohistochemistry ; In Situ Hybridization, Fluorescence ; Lung Neoplasms ; genetics ; metabolism ; pathology ; Male ; Middle Aged ; Retrospective Studies ; Sex Factors ; Smoking ; Young Adult

The aim of this study was to evaluate the effect of all-trans retinoic acid (ATRA) on cell adhesion molecule expression and adhesion capacity of bone marrow stromal cells (BMSC) in patients after conditioning treatment for peripheral blood stem cell transplantation (PBSCT). BMSC of 27 patients before and after conditioning treatment for PBSCT were cultured in vitro. After treated with ATRA at 0.01, 0.1, or 1 micromol/L, expression of intercellular adhesion molecule-1 (ICAM-1) protein and vascular adhesion molecule-1 (VCAM-1) protein were detected by flow cytometry, and soluble ICAM-1 (sICAM-1) protein was determined by using radioimmunoassay. Then BMSC was co-cultured with CD34+ cells, and adhesion rate of BMSC to CD34+ cells was measured. The results showed that after pretreatment with conditioning regimen for PBSCT, the expressions of ICAM-1 and VCAM-1 proteins in BMSC and the expression level of sICAM-1 protein in supernatant of BMSC culture were down-regulated, and the adhesion rate of BMSC to CD34+ cells was decreased, after administration of ATRA, the expression of ICAM-1 protein in BMSC, sICAM-1 protein in culture medium and adhesion rate of BMSC to CD34+ cells all increased significantly, but expression of VCAM-1 protein changed no significantly. It is concluded that the ATRA can partly restore adhesion function of BMSC injured by pretreatment for PBSCT and contribute to hematopoietic reconstitution.
Adolescent ; Adult ; Antigens, CD34 ; Antineoplastic Agents ; pharmacology ; Bone Marrow Cells ; metabolism ; pathology ; Cell Adhesion ; drug effects ; Cell Adhesion Molecules ; biosynthesis ; genetics ; Child ; Coculture Techniques ; Hematologic Neoplasms ; metabolism ; pathology ; therapy ; Humans ; Middle Aged ; Peripheral Blood Stem Cell Transplantation ; Stromal Cells ; metabolism ; pathology ; Tretinoin ; pharmacology ; Tumor Cells, Cultured ; Vascular Cell Adhesion Molecule-1 ; biosynthesis ; genetics