Objective To discover radioresistance-associated proteins by performing comparative proteomic analysis on nasopharyngeal carcinoma cell lines.Methods The total proteins were extracted from radioresistant human nasopharyngeal carcinoma cell line CNE-2R and its parental cell line CNE-2,respectively.These proteins were separated by high quality two-dimensional polyacrylamide gel electrophoresis (2-DE) and then the 2-DE profiles were screened for differentially expressed protein spots by the Image Master 5.0 software.Those spots were identified by a matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry.Results 32 significantly differentially expressed protein spots were screened in two different radiosensitivity cell lines and 11 proteins were identified by tandem mass spectrometry,among which 3 proteins were up-regulated in radioresistant human nasopharyngcal carcinoma cell line CNE-2R and the other 8 proteins were down-regulated.Conclusions The differentially expressed proteins of nasopharyegeal carcinoma cells with different radiosensitivity were mainly involved in apoptosis regulation,DNA damage and repair,cell cycle regulation,RNA transcription,cell signaling,cytoskeleton formation and radiation stress responses.

Objective To discover radioresistance associated molecular biomarkers and its mechanism in nasopharyngeal carcinoma by protein-protein interaction network analysis.Methods Whole genome expression microarray was applied to screen out differentially expressed genes in two cell lines CNE- 2R and CNE-2 with different radiosensitivity.Four differentially expressed genes were randomly selected for further verification by the semi-quantitative RT-PCR analysis with self-designed primers. The common differentially expressed genes from two experiments were analyzed with the SNOW online database in order to find out the central node related to the biomarkers of nasopharyngeal carcinoma radioresistance. The expression of STAT1 in CNE-2R and CNE-2 cells was measured by Western blot.Results Compared with CNE-2 cells,374 genes in CNE-2R cells were differentially expressed while 197 genes showed significant differences.Four randomly selected differentially expressed genes were verified by RT-PCR and had same change trend in consistent with the results of chip assay. Analysis with the SNOW database demonstrated that those 197 genes could form a complicated interaction network where STAT1 and JUN might be two key nodes.Indeed,the STAT1-α expression in CNE-2R was higher than that in CNE-2 (t =4.96,P ＜ 0.05).Conclusions The key nodes of STAT1 and JUN may be the molecular biomarkers leading to radioresistance in nasopharyngeal carcinoma,and STAT1-α might have close relationship with radioresistance.

Objective To investigate the role of autophagy in radiation-induced death response of human nasopharyngeal carcinoma cells.Methods MTT method was used to detect cell viability of CNE-2 cells in different time after irradiation.Clonogenic survival assay was used to evaluate the effect of autophagy inhibitor (chloroquine phosphate) and autophagy inductor (rapamycin) on radiosensitivity of nasopharyngeal carcinoma cells.Cell apoptosis was assessed by flow cytometry.The expressions of LC3 and P62 were measured with Western blot.Cell ultrastructural analysis was performed under an electron microscope.Results Irradiation with 10 Gy induced a massive accumulation of autophagosomes accompanied with up-regulation of LC3-Ⅱ expression in CNE-2 cells.Compared with radiation alone,chloroquine phosphate (CDP) enhanced radiosensitivity significantly by decreasing cell viability (F =25.88,P ＜ 0.05),autophagic ratio (F =105.15,P ＜ 0.05),and LC3-Ⅱ protein level(F =231.68,P ＜0.05),while up-regulating the expression of P62 (F =117.52,P ＜ 0.05).Inhibition of autophagy increased radiation-induced apoptosis (F =143.72,P ＜ 0.05).Rapamycin (RAPA) also significantly decreased cell viability,but increased autophagic ratio and LC3-Ⅱ protein level while down-regulated the expression of P62.Induction of autophagy increased radiation-induced apoptosis(F =167.32,P ＜ 0.05).Conclusions Blockage of autophagy with CDP could enhance radiosensitivity in human nasopharyngeal carcinoma cells,suggesting that inhibition of autophagy could be used as an adjuvant treatment to nasopharyngeal carcinoma.

Aged ; Breast Neoplasms ; metabolism ; pathology ; surgery ; Cadherins ; metabolism ; Carcinoma, Merkel Cell ; metabolism ; pathology ; Carcinoma, Small Cell ; metabolism ; pathology ; surgery ; Diagnosis, Differential ; Female ; Humans ; Lymphoma ; metabolism ; pathology ; Melanoma ; metabolism ; pathology ; Phosphopyruvate Hydratase ; metabolism ; Synaptophysin ; metabolism

Objective To study the difference of gene expression profile between the radioresistant human nasopharyngeal carcinoma cell line CNE-2R and CNE-2,and to screen the signaling pathway associated with radioresistance of nasopharyngeal carcinoma.Methods The radioresistant nasopharyngeal carcinoma cell line CNE-2R was constructed from the original cell line CNE-2.CNE-2R and CNE-2 cells were cultured and administered with 60Co γ-ray irradiation at the dose of 400 cGy for 15 times.Human-6v 3.0 whole genome expression profile was used to screen the differentially expressed genes.Bioinformatic analysis was used to identify the pathways related to radioresistance.Results The number of the differentially expressed genes that were found in these 2 experiments was 374.The Kegg pathway and Biocarta pathway analysis of the differentially expressed genes showed the biological importance of Toll-like receptor signaling pathway and IL-1 R-mediated signal transduction pathway to the radioresistance of the CNE-2R cells and the significant differences of 13 genes in these 2 pathways,including JUN,MYD88,CCL5,CXCL10,STAT1,LY96,FOS,CCL3,IL-6,IL-8,IL-1α,IL-1B,and IRAK2(t=13.47-66.57,P<0.05).Conclusions Toll-like receptor signaling pathway and IL-1R-mediated signal transduction pathway might be related to the occurrence of radioresistance.

OBJECTIVETo investigate the induction of antitumor immune responses and therapeutic effects of 10-hydroxycamptothecinc-treated (HCPT) DC-Hepa fusion vaccines by DC fused with hepal-6 cell from hepatoma.

METHODSThe fused cells were isolated by magnetic cell sorting and adherent culture. Cell apoptosis was detected by Rhodamine123/PI double-labeled assay, CTL activity by 4 h (51)Cr releasing assay. Protective and therapeutic effects of the fusion vaccine to the tumor-bearing mice was also observed.

RESULTSThe apoptosis rate was 29.7%+/-4.1% when DC-Hepa fusion vaccine was treated with 50 microg/ml HCPT for 24 h. After treatment with the HCPT-DC-Hepa fusion vaccine, the tumor grew obviously slowly, survival period of the mice was prolonged, induced more potent CTL cytotoxicity, and resisted against the rechallenge of Hepal-6 cells.

CONCLUSIONVaccination with HCPT-DC-Hepa fusion vaccine could elicit potent antitumor responses, which will provide a new approach to the DC-mediated therapeutic antitumor immunity.

Animals ; Antineoplastic Agents, Phytogenic ; pharmacology ; Apoptosis ; Camptothecin ; analogs & derivatives ; pharmacology ; Cancer Vaccines ; administration & dosage ; genetics ; immunology ; Cell Fusion ; Cytotoxicity, Immunologic ; Dendritic Cells ; immunology ; transplantation ; Female ; Liver Neoplasms ; immunology ; pathology ; Mice ; Mice, Inbred C57BL ; Neoplasm Transplantation ; Rats ; T-Lymphocytes, Cytotoxic ; immunology ; Tumor Cells, Cultured

Ten compounds, including seven sesquiterpenes, two phenols and one phenylpropanoid, were isolated from the roots of Illicium majus by means of silica gel, ODS, Sephadex LH-20, and preparative HPLC. On analysis of MS and NMR spectroscopic data , their structures were established as cycloparviflorolide (1), cycloparvifloralone (2), tashironin (3), tashironin A (4), anislactone A(5), anislactone B (6), pseudomajucin (7), syringaldehyde (8), methyl-4-hydroxy-3, 5-dimethoxybenzoate (9), and (E)-3-methoxy-4,5-methylenedioxycinnamic alchol (10), respectively. Compounds 1-4 and 8-10 were first isolated from this plant. In the in vitro assays, at a concentration of 1.0 x 10(-5) mol x L(-1), compounds 5 and 6 were active against LPS induced NO production in microglia with a inhibition rate of 75.31% and 53.7%, respectively.
Drugs, Chinese Herbal ; analysis ; chemistry ; Illicium ; chemistry ; Organic Chemicals ; analysis ; chemistry ; Plant Roots ; chemistry

OBJECTIVETo establish a set of procedure for recovery and species identification of Legionella from the surface environmental water.

METHODSForty-four water samples were collected in eight parks of Guangzhou city from August to November in 2006. The bacteriologic examination was performed by cultivation on BCYEalpha plate, and 108 presumptive Legionella colonies were picked and their homogeneous relationship was analyzed by using an amplified fragment length polymorphism (AFLP) method. Species identification was carried out by latex agglutination test, biochemical characterization, analysis of cellular fatty acids composition, 16 S rRNA gene and mip gene sequencing.

RESULTSLegionella was recovered among 27 (61.36%) samples of all eight parks, and 31 different strains were identified from those 108 presumptive Legionella isolates by AFLP method, including 20 strains of L. pneumophila, five strains of L. feeleii, four strains of L. longbeachae, one strain of L. oakridgensis and one strain of L. sainthelensi, and L. pneumophila could be easily differentiated by phenotypic and biochemical characteristics, latex agglutination test or analysis of the cellular fatty acids composition . However, uncertain factors were existing in those phenotypic identification methods as compared to the sequence analysis.

CONCLUSIONThe taxonomic analysis of the Legionellae family should be dependent on the 16 S rRNA gene or mip gene.

Bacteriological Techniques ; DNA, Bacterial ; genetics ; Environmental Monitoring ; methods ; Legionella ; genetics ; isolation & purification ; RNA, Bacterial ; RNA, Ribosomal, 16S ; Water Microbiology

OBJECTIVETo determine the antibiotics resistance, aminoglycoside-modifying enzymes and homology of high-level gentamycin resistant enterococcus in clinical specimens.

METHODSThe high-level gentamicin resistant (HLGR) isolates were screened by the agar method and the resistance of 14 antimicrobial agents was determined by K-B method. The aminoglycoside-modifying enzyme genes were detected by polymerase chain reaction (PCR). Pulsed-field gel electrophoresis (PFGE) was used to analyze the homology of HLGR isolates.

RESULTSThe ratio of HLGR was 64.2% (68/106). Among the HLGR,there were no isolates resistant to linezolid, vancomycin and tecoplanin, and Enterococcus faecium was more resistant to beta-lactam antibiotics and quinolone than Enterococcus faecalis. The positive rate of aac(6')-Ie-aph(2')-Ia was 92.6% and 3 isolates had the resistance gene mostly similar to aph(2')-Id. And among 51 HLGR isolates from the hospitalized patients, PFGE grouped 17 E. faecalis isolates into 4 clusters (A-D), and 33 E. faecium isolates into 8 clusters (A-H) with A cluster as predominant.

CONCLUSIONHLGR has become the important antibiotic resistance bacteria which results in nosocomial infection; and aac(6')-Ie-aph(2')-Ia is the main aminoglycoside-modifying enzyme gene which causes HLGR.

Drug Resistance, Bacterial ; genetics ; Electrophoresis, Gel, Pulsed-Field ; Enterococcus ; drug effects ; genetics ; Enterococcus faecalis ; drug effects ; genetics ; Enterococcus faecium ; drug effects ; genetics ; Gentamicins ; pharmacology ; Humans ; Kanamycin Kinase ; genetics ; Microbial Sensitivity Tests

Purpose: The aim of this study was to compare the efficacy of liver transplantation (LT) and liver resection (LR) for hepatocellular carcinoma (HCC) patients with portal vein tumor thrombus (PVTT) and to investigate risk factors affecting prognosis. Materials and Methods: A total of 94 HCC patients with PVTT type I (segmental PVTT) and PVTT type II (lobar PVTT) were involved and divided into LR (n=47) and LT groups (n=47). Recurrence-free survival (RFS) and overall survival (OS) were compared before and after inverse probability of treatment weighting (IPTW). Prognostic factors for RFS and OS were explored. Results: Two treatment groups were well-balanced using IPTW. In the entire cohort, LT provided a better prognosis than LR. Among patients with PVTT type I, RFS was better with LT (p=0.039); OS was not different significantly between LT and LR (p=0.093). In subgroup analysis of PVTT type I patients with α-fetoprotein (AFP) levels >200 ng/mL, LT elicited significantly longer median RFS (18.0 months vs. 2.1 months, p=0.022) and relatively longer median OS time (23.6 months vs. 9.8 months, p=0.065). Among patients with PVTT type II, no significant differences in RFS and OS were found between LT and LR (p=0.115 and 0.335, respectively). Multivariate analyses showed treatment allocation (LR), tumor size (>5 cm), AFP and aspartate aminotransferase (AST) levels to be risk factors of RFS and treatment allocation (LR), AFP and AST as risk factors for OS. Conclusion LT appeared to afford a better prognosis for HCC with PVTT type I than LR, especially in patients with AFP levels >200 ng/mL.