Objective: To investigate the effects of recombinant parvovirus H-1 vector expressing p21(rhH1 Δ p21) on human gastric cancer cell line HGC27, and to further reveal the biological function of p21wif1 to provide the basis for cancer gene therapy. Methods: The recombinant parvovirus H-1 vector expressing p21 (rhH1 Δ/p21) was constructed by reverse transcriptase-polymerase chain reaction (RT-PCR), and was transfected into HGC27 cell line. The morphological changes of HGC27 cell were observed. The transgene protein expressions in the gastric cancer cells were detected by Western blot. The inhibitory effects of rhH1 Δ/p21 on the growth of HGC27 cells were measured by MTT assay. The cell cycle distribution was determined by flowcytometry. Results: The rhH1 Δ/p21 was successfully constructed, with a titer of 3.5 × 107 PFU/mL. The transgene protein p21 was over-expressed in the HGC27 cells. The cell cycle distribution was changed. The proportion of cells in G1 phase significantly increased. The cell growth was significantly inhibited. Conclusion: rhH1 Δ/p21 induces G1 arrest and inhibits the proliferation of gastric cancer HGC27 cells. It indicates that rhH1 Δ/p21 gene therapy can effectively inhibit the growth of gastric cancer cells in vitro.

Objective To observe the influence of coal burning fluorosis on learning and memory ability in rats and reveal its possible mechanisms. Methods Healthy 48 SD rats were divided into control, low-fluoride and high-fluoride group. All rats in fluoride exposed groups were fed with the eom polluted by drying processes with burning coal containing high level of fluoride obtained from the endemic fluorosis area to produce the animal model of fluorosis. The experiment period were 3,6 mouths, respectively. The ability of leaning and memory was measured by Morris test and cholinesterase activity detected by photometric method at 3 or 6 month after experiment, respectively. Results Fluoride contents signifieantlly influenced the escape latency, the numbers of crossing the platforms and the time of staying the platforms(the value of F was 29.29,6.47,6.50, respectively, P<0.01).In addition, the numbers of crossing the platforms and the time of staying the platforms were influenced by the exposed time(the value of F was 16.11,45.59, P<0.01). Furthermore, the fluoride contents and the exposed time had an interaction between the numbers of crossing the platforms and the time of staying the platforms (the value of F was 4.67,5.68, P<0.05 or<0.01). Three months after the experiment, the mean values of escape latency [(14.71± 4.85)s] of rats in highly fluoride exposed group were significantly prolonged as compared with controls [(9.28±4.22)s]; 6 month after the experiment, the mean values of escape latency[(12.42±8.03)s, (17.48± 8.05)s] of rats in both groups exposed to fluoride were significantly prolonged as compared to controls [(7.04± 3.29)s, P<0.05]. The decreased numbers of crossing the platforms[(1.62±0.87)number] and the declined time of staying the platforms[(16.70±5.02)s] were found in the rats exposed to high fluoride as compared to controls [(3.53±1.67 )number, (23.33±5.35)s, P<0.05]. The fluoride contents obviously influenced the activities of acetylcholinesterase and butylcolinesterase (the value of F was 12.83,13.27, P<0.01). On the other hand, the times of breeding also influnced the activities of butylcolinesterase (the value of F was 16.26, P<0.01). In 3 months of the experiment, the activities of butylcolinesterase [(0.55±0.12)kU/g] in low fluoride exposed group were significantly decreased in comparison with controls[(0.73±0.10)kU/g, P<0.05]. The activities of acetylcholinesterase[(0.62±0.42)kU/g] and butylcolioesterase[(0.58±0.10)kU/g] in high fluoride group were significantly decreased as compared to eontrois[(1.41±0.52), (0.73±0.10)kU/g, P<0.05]. The correlation analysis showed that there was a negative correlation between the cholinesterase and the escape latency(r=-0.68, P< 0.01), and a positive correlation between the cholinesterase and the time of staying the platforms(r=0.57, P< 0.01). Conclusions The ability of learning and memory in rats with coal buring fluorosis was decreased, which might be connected to the decreased activity of cholinesterase in a dose-effect correlation.

Objective：This study was designed to express chimeric anti-VEGF (vascular endothelial growth factor) antibody in dihydrofolate reductase-deficient Chinese hamster ovary (CHO-dhfr-)cells at high-level, and explore an optimum method to obtain high-level expression cells clone. Methods：The light chain and heavy chain genes of chimeric anti-VEGF antibody were induced into CHO-dhfr-cells using a novel eukaryotic high-level expression vectors system for genetic engineering antibodies. High-level expression was achieved after subcloning and several rounds of co-amplification of methotrexate (MTX). Biological features and productive amount of chimeric antibody was charactered by ELISA. Result：The cells strain that secret anti-VEGF chimeric antibody at the highest level of 28 μ g/ml was established. The cells were subcloned following each round of co-amplification of MTX, while greatly different results were obtained using three methods. The chimeric antibody contained constant regions of human immunoglobin and had the specificity against VEGF by ELISA. Conclusion：The anti-VEGF mouse-human chimeric antibody was expressed at high-level successfully in CHO cells. This may be an optimum method to obtain high-level expression cells clone for the eukaryotic high-level expression vectors system.

BACKGROUNDEpigallocatechin-3-gallate (EGCG) has been demonstrated to have anti-neoplastic activity, but the effective concentration of EGCG and its possible mechanisms are uncertain. The study on the killing effects of EGCG on different digestive tract cancer cell lines can find target sites of its anti-neoplastic effect and provide a theoretical basis for its clinical application in the treatment of cancers.

METHODSMethyl thiazolyl tetrazolium (MTT) analysis was made to detect the differential sensitivities of eight digestive tract cancer cell lines to EGCG. The effect of EGCG on cell cycle distribution of sensitive cancer cell line was measured by flow cytometry. By polymerase chain reaction (PCR)-enzyme linked immunosorbent assay (ELISA) protocol, the influence of EGCG on telomerase activity of sensitive cancer cell line was also investigated. RT-PCR method was employed to detect the influence of EGCG on the expressions of hTERT, c-myc, p53 and mad1 genes in sensitive cancer cell line.

RESULTSEGCG exhibited dose-dependent killing effects on all eight digestive tract cancer cell lines. The 50% inhibitory concentration (IC50) of SW1116, MKN45, BGC823, SGC7901, AGS, MKN28, HGC27 and LoVo cells were 51.7 micromol/L, 55.9 micromol/L, 68.5 micromol/L, 79.1 micromol/L, 83.8 micromol/L, 119.8 micromol/L, 183.2 micromol/L and 194.6 micromol/L, respectively. There were no apparent changes in cell cycle distribution of sensitive cancer cell line MKN45 48 hours after incubating with three different concentrations of EGCG compared with the controls. It was found that EGCG could suppress the telomerase activity of MKN45 cells, and the effects were dose- and time-dependent. After EGCG administration, the expression of hTERT and c-myc genes in MKN45 cells was decreased, that of the mad1 gene increased, and that of the p53 gene unchanged.

CONCLUSIONSEGCG has dose-dependent killing effects on different digestive tract cancer cell lines. Administration of EGCG has no obvious effect on cell cycle distribution of sensitive cancer cell line MKN45. The anti-neoplastic activity of EGCG might be due to the inhibition of telomerase activity by means of its influence on hTERT and the up-stream regulation genes.

Antineoplastic Agents ; pharmacology ; Catechin ; analogs & derivatives ; pharmacology ; Cell Cycle ; drug effects ; Cell Line, Tumor ; DNA-Binding Proteins ; genetics ; Digestive System Neoplasms ; drug therapy ; metabolism ; pathology ; Dose-Response Relationship, Drug ; Genes, myc ; Genes, p53 ; Humans ; Telomerase ; genetics ; metabolism

Objective To explore epithelial ovarian cancer(EOC)antigens that are potentially useful for cancer early detection and therapy.Methods A high quality cDNA library derived from ascites tumor cells of EOC patients(3 cases of serous EOC,1 case of mucinous EOC,and 1 case of endometrial carcinoma of ovary)was constructed,and the method of combining serological analysis of recombinant cDNA expression libraries(SEREX)and suppression subtractive hybridization(SSH)was used for screening cDNA library.All of the positive clones were sequenced and bioinformatics analysis with BLAST software in GenBank was performed.Serological mini-arrays of recombinant tumor antigens(SMARTA)was used to investigate the prevalence of autoantibodies to these antigens in both 96 ovarian cancer patients and 96 cancer-free controls.Results Fifty-five positive clones encoding different antigenic genes of EOC recognized by IgG and(or)IgM were obtained.It showed that these 55 clones derived from 45 distinct genes and these genes could be grouped into 6 classes as following according to homology with known expressed sequence tag(EST):(1)known ovarian carcinoma related genes:BARD1,et al;(2)homologous genes with other tumors:TM4SF1,et al;(3)homologous genes with special tissues:ILF3,FXR1,et al;(4) homologous genes with special function:TIZ,C1 D,et al;(5)embryo originating genes:PKHD1,et al; (6)novel genes:OV-189,et al.SMARTA results showed that the positive ratio of five EOC antigens TM4SF1(28% vs 9%),CID(21% vs 6%),BARD1(23% vs 5%),FXR1(23% vs 8%),OV-189 (31% vs 13%)which reacting with their IgG autoantibodies,three antigens TIZ(26% vs 8%),FXR1 (28% vs 11%),and OV-189(18% vs 7%)which reacting with their IgM autoantibodies in patients was higher than in controls(P

Objective To investigate the influences of Chinese herbal remedies (CHR) with the actions of replenishing qi and nourishing yin, and resolving mass and freeing the collateral vessels on renal nephrin-mRNA and ultrastructure in rats with early diabetes. Methods All SD rats were randomly divided into 6 groups including group 1 (with nephrectomy), group 2 (model), group 3 (with irbesatan), group 4 (with low-dose CHR), group 5 (with mid-dose CHR) and group 6 (with high-dose CHR). The relevant drugs or CHR were given intragastrically to different groups respectively. At the end of 6th week the quantity of 24-hour urine protein (UPro) and kidney function were detected. The pathological changes of renal cortex were observed by using transmission electron microscope (TEM) after HE staining. The expression of nephrin was determined by applying RT-PCR.Results There were pathological changes of diabetic nephropathy observed by the light microscope and TEM in group 2 and 1, which were alleviated in other 4 groups. The quantity of UPro decreased significantly in group 3, 4 and 5 compared with that in group 2 and 6 (P<0.05). The renal function was improved significantly in all treatment groups compared with that in group 2 (P<0.05). In group 3, 4, 5 and 6 the expression of nephrin-mRNA increased significantly compared with that in group 2 (P<0.05).Conclusion CHR with the actions of replenishing qi and nourishing yin, and resolving mass and freeing the collateral vessels have a protective effect on kidney in rats with diabetic nephropathy induced by STZ.

In this study, it is to compare the effectiveness of prevention against and treatment of doxorubicin (DOX) induced cardiotoxicity by dexrazoxane and schisandrin B (Sch B) in rats. Sprague-Dawley (SD) rats were randomly divided into the following 6 groups: normal saline group, DOX group, DOX+DEX group, DOX+Sch B (80 mg x kg(-1)) group, DOX+Sch B (40 mg x kg(-1)) group and DOX+Sch B (20 mg x kg(-1)) group. The results showed that Sch B could combat the increase of myocardial enzymes in peripheral blood, decrease of the enzyme activity of myocardial tissue antioxidant enzymes and disorders of systolic and diastolic function of heart in rats intravenously injected with doxorubicin (15 mg x kg(-1)). Sch B was better than DEX in protecting rat against DOX-induced the symptoms. Sch B could protect rat against DOX-induced acute cardiomyopathy and has clinical potential applications.
Animals ; Antibiotics, Antineoplastic ; adverse effects ; Antioxidants ; metabolism ; Cardiomyopathies ; chemically induced ; drug therapy ; Cardiotoxicity ; drug therapy ; Cyclooctanes ; therapeutic use ; Dexrazoxane ; therapeutic use ; Doxorubicin ; adverse effects ; Heart ; physiopathology ; Lignans ; therapeutic use ; Myocardium ; enzymology ; Polycyclic Compounds ; therapeutic use ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo further study the mechanism of the inhibitory effect of interferon beta-1a (IFN beta-1a) on the activation of human hepatic stellate cell (HSC) LX-2, and to analyze the differences on the protein expression in LX-2 induced by I IFN beta-1a.

METHODSCultured LX-2 cells were treated with 2000 U/ml IFN beta-1a for 48 h. Two-dimensional gel electrophoresis (2-DE) was performed to compare protein patterns of the control (untreated) and IFN beta-1a treated LX-2 and for quantitative and qualitative analyses of protein expression. A rat liver fibrosis model was established and the rats were sacrificed and their various tissues were obtained for the same analyses. Western blotting and RT-PCR were used to validate the expression of the changed proteins after treatment of IFN beta-1a in LX-2 cells and of various tissues of the rats.

RESULTS708 +/- 25 spots were detected in control LX-2 cells and 804 +/- 32 spots in IFN beta-1a-treated LX-2 cells. A match rate of 73%-82% was achieved. The results also showed that 31 protein spots displayed quantitative changes in expression after IFN beta-1a treatment. Of the 31 spots, 21 proteins were identified, of which, one was newly found, two were enhanced in abundance and 18 showed lower expressions. The newly found protein was glia maturation factor beta (GMF beta). The treatment of LX-2 with IFN beta-1a increased the production of GMF beta(GMF beta) protein in comparison with the untreated cells (t=1.81, P < 0.01). The expression of GMF beta protein (1.81 vs 0.10) and mRNA (0.85 vs 0.12) were more in the normal liver tissues than in the cirrhotic liver tissues (t=2.53, 2.13 respectively, P < 0.01). The expressions of GMF beta protein and mRNA were weak in rat heart and lung tissues, however, they were strong in rat liver, kidney, spleen and brain tissues (t=1.91, 1.94 respectively, P < 0.01).

CONCLUSIONThere is a significant difference of protein expression levels between IFN beta-1a untreated and treated LX-2 cells. These proteins, especially GMF beta, may be involved in an inhibition process of IFN beta-1a on activation and apoptosis of LX-2 cells. This proteome study may be useful in further studies of the relationship of IFN beta-1a treatment and human liver diseases.

Animals ; Cell Line ; Female ; Glia Maturation Factor ; metabolism ; Hepatic Stellate Cells ; metabolism ; Humans ; Interferon beta-1a ; Interferon-beta ; pharmacology ; Liver ; cytology ; Liver Cirrhosis ; metabolism ; Proteome ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo study the possible immunological mechanism of wheezing attack in children with cytomegalovirus (CMV) infection.

METHODSA total of 25 under-5-year-old children with wheezing following CMV infection were enrolled. The expression of serum regulatory T cells (Treg)/T helper 17 (Th17) cytokines interleukin (IL)-10, IL-6, and IL-17, and peripheral blood lymphocyte subsets were determined. Twenty age-matched healthy children were selected as the control group.

RESULTSThe wheezing group had a significantly reduced serum IL-10 level, significantly increased IL-6 and IL-17 levels, significantly reduced levels of natural killer cells, and significantly increased levels of CD8+ T cells and CD19+ B cells, as compared with the control group.

CONCLUSIONSWheezing children with CMV infection have Treg/Th17 imbalance and cellular immune dysfunction, which may be an important immunological mechanism of the development of wheezing in children after CMV infection.

Child, Preschool ; Cytokines ; blood ; Cytomegalovirus Infections ; immunology ; Female ; Humans ; Infant ; Male ; Respiratory Sounds ; etiology ; immunology ; T-Lymphocytes, Regulatory ; immunology ; Th17 Cells ; immunology

OBJECTIVETo evaluate the response rate to vaccination in different treatment groups (nonimmunosuppressants and immunosuppressants).

DATA SOURCESWe completed an online systematic search using PubMed to identify all articles published in English between January 1990 and December 2013 assessing the effect of the response rate to vaccination in different treatment groups (with and without immunomodulators). The following terms were used: "inflammatory bowel disease (IBD)" OR "Crohn's disease" OR "ulcerative colitis" AND ("vaccination" OR "vaccine") AND ("corticosteroids" OR "mercaptopurine" OR "azathioprine" OR "methotrexate [MTX]") AND "immunomodulators."

STUDY SELECTIONThe inclusion criteria of articles were that the studies: (1) Randomized controlled trials which included patients with a diagnosis of IBD (established by standard clinical, radiographic, endoscopic, and histologic criteria); (2) exposed patients received immunomodulators for maintenance (weight-appropriate doses of 6-mercaptopurine/azathioprine or within 3 months of stopping, 15 mg or more MTX per week or within 3 months of stopping; (3) exposed patients received nonimmunomodulators (no therapy, antibiotics only, mesalazine only, biological agent only such as infliximab, adalimumab, certolizumab or natalizumab or within 3 months of stopping one of these agents). The exclusion criteria of articles were that the studies: (1) History of hepatitis B virus (HBV), influenza or streptococcus pneumoniae infection; (2) patients who had previously been vaccinated against HBV, influenza or streptococcus pneumoniae; (3) any medical condition known to cause immunosuppression (e.g. chronic renal failure and human immunodeficiency virus infection); (4) individuals with positive hepatitis markers or liver cirrhosis; (5) patients with a known allergy to eggs or other components of the vaccines and (6) pregnancy.

RESULTSPatients treated with immunomodulators were associated with lower response rates to vaccination.

CONCLUSIONSImmunomodulators may impair the immune response to vaccination in patients with IBD. Vaccination should be made at the time of diagnosis or before starting immunosuppressed therapy.

Colitis, Ulcerative ; prevention & control ; Crohn Disease ; prevention & control ; Humans ; Immunosuppressive Agents ; therapeutic use ; Inflammatory Bowel Diseases ; prevention & control ; Vaccination