Background According to data from the China Hospital Invasive Fungal Surveillance Net (CHIF-NET) 2010,Candida tropicalis (C.tropicalis) is the third most common pathogen causing invasive candidiasis.Moreover,the majority of fluconazole-resistant C.tropicalis isolates were from a single hospital.Therefore,a molecular epidemiological survey is necessary to investigate the genetic relatedness of C.tropicalis isolates in China.Methods In this study,48 C.tropicalis isolates causing invasive fungal infections from four tertiary hospitals in China were studied.All the isolates were identified by sequencing the internal transcribed spacer region.Antifungal susceptibility to triazoles,amphotericin B,and caspofungin was determined by the Clinical and Laboratory Standards Institute standard broth microdilution method.Multilocus sequence typing (MLST) was performed,and phylogenetic analysis was further performed by the eBURST and maximum parsimony (MP) methods to characterize the genetic relatedness of isolates.Results MLST discriminated 40 diploid sequence types (DSTs) among 48 isolates,including 36 novel DSTs,and the XYR1 gene showed the highest discriminatory power.The DSTs obtained from this study were compared with those of previously reported C.tropicalis isolates,and there was poor type alignment with regional strains.Nine groups and 11 singletons were identified by eBURST,whereas two groups and 10 subgroups were clustered by MP analysis.Generally,there were no obvious correlations between clonal clusters generated and the specimen source or hospital origin.Seven fluconazole-resistant isolates were confirmed and assigned to three distinguishable branches.Conclusions The results suggested diverse origins of invasive C.tropicalis isolates in China.Although most invasive C.tropicalis strains in the mainland of China were clustered with previously characterized Asian isolates,major C.tropicalis clusters identified in this study were genetically distinct from those of other geographic regions.

Nasopharyngeal carcinoma (NPC) is among the most common malignancies in southern China. Deletion of genomic DNA, which occurs during the complex pathogenesis process for NPC, represents a pivotal mechanism in the inactivation of tumor suppressor genes (TSGs). In many circumstances, loss of TSGs can be detected as diagnostic and prognostic markers in cancer. The short arm of chromosome 3 (3p) is a frequently deleted chromosomal region in NPC, with 3p21.1-21.2 and 3p25.2-26.1 being the most frequently deleted minimal regions. In recent years, our research group and others have focused on the identification and characterization of novel target TSGs at 3p, such as RASSF1A, BLU, RBMS3, and CHL1, in the development and progression of NPC. In this review, we summarize recent findings of TSGs at 3p and discuss some of these genes in detail. A better understanding of TSGs at 3p will significantly improve our understanding of NPC pathogenesis, diagnosis, and treatment.
Cell Adhesion Molecules ; genetics ; Chromosome Deletion ; Chromosomes, Human, Pair 3 ; genetics ; Epigenesis, Genetic ; Gene Expression Regulation, Neoplastic ; Genes, Tumor Suppressor ; Heterotrimeric GTP-Binding Proteins ; genetics ; Humans ; Nasopharyngeal Neoplasms ; genetics ; RNA-Binding Proteins ; genetics ; Trans-Activators ; genetics ; Tumor Suppressor Proteins ; genetics

Adult ; Diagnosis, Differential ; Female ; Glycogen ; metabolism ; Glycogen Storage Disease Type II ; pathology ; Humans ; Liver ; metabolism ; pathology ; Muscle, Skeletal ; metabolism ; pathology ; Muscular Dystrophies, Limb-Girdle ; pathology ; Pyomyositis ; pathology ; Young Adult

OBJECTIVETo observe the effects of complement fragment C3f on expression and secretion of collagen I, III and transforming growth factor( TGF)-beta1 in human embryonic lung fibroblast (MRC-5) cells.

METHODSMRC-5 cells were cultured with C3f (the synthetic 17 peptides fragments of complement C3). The extracellular and intracellular expression levels of type I, III collagens and TGF-beta1 in MRC-5 cultures were detected by ELISA and immunohistochemistry, respectively.

RESULTSThe expression levels of type I, III collagen and TGF-beta1 in the supernatant of MRC-5 cultures decreased significantly with the concentrations of C3f as compared with controls (P < 0.05). Also the expression level of TGF-beta1 in MRC-5 cytoplasm reduced significantly as compared with controls (P < 0.05).

CONCLUSIONThe results of present in vitro study showed that the complement fragment C3f could reduce the formation of TGF-beta1 and type I, III collagens in MRC-5 cells, and inhibit the lung tissue fibrosis.

Cell Line ; Collagen Type I ; metabolism ; Collagen Type III ; metabolism ; Complement C3b ; pharmacology ; Fibroblasts ; drug effects ; metabolism ; Humans ; Lung ; cytology ; drug effects ; embryology ; Transforming Growth Factor beta1 ; metabolism

OBJECTIVETo study the inhibition of maxizyme (Mz) directed against the mutant-type p53 gene (mtp53) at codon 249 in exon 7 (AGG --> AGT) both in cell-free system and in MHCC97 cell lines.

METHODSMaxizyme and control mutant maxizyme (G5 --> A5) were designed by computer and cloned into the eukaryotic expression vector pBSKneoU6 (pU6Mz, pU6asMz). Mz was driven by T7 RNA polymerase promoter in vitro. In the cell lines, U6 promoter was driven by RNA PolIII. The mutant type p53 gene fragment was cloned into the pGEM-T vector under the T7 promoter control. The 32P-labeled mtp53 transcript was the target RNA. Cold maxizyme transcripts were incubated with 32P-labeled target RNA in vitro. pU6Mz was introduced into MHCC97 cells by Lipofectamine2000 and mtp53 expression was analyzed by RT-PCR and Western blot.

RESULTSIn vitro cleavage showed that pU6Mz was very active with cleavage efficiency of 42% while pU6asMz was not. The wild type p53 was not cleaved. Partial down-regulation of mtp53 mRNA and mtp53 protein were observed in MHCC97 cells transfected with pU6Mz but not those with pU6asMz. The proliferation of MHCC cells was inhibited by MTT analysis.

CONCLUSIONOur findings suggest that the chimeric U6 maxizyme against the mtp53 is a new promising gene therapeutic agent in treating hepatocellular carcinoma.

Carcinoma, Hepatocellular ; genetics ; Cell Line, Tumor ; Genetic Therapy ; methods ; Genetic Vectors ; Humans ; Liver Neoplasms ; genetics ; Nucleic Acid Conformation ; Point Mutation ; Protein Conformation ; RNA, Catalytic ; RNA, Messenger ; chemical synthesis ; metabolism ; Recombinant Fusion Proteins ; Ribonuclease T1 ; pharmacology ; Tumor Suppressor Protein p53 ; genetics

OBJECTIVEPersistent replication of hepatitis B virus (HBV) is one of the major obstacles in HBV infection treatment. Reduction or clearance of HBV propagation would be one of the aims of HBV therapy. The drugs approved in clinical used such as nucleotide analogs or interferon, were limited effects on HBV replication. The newly developing gene therapy method, dominant negative mutants, were be used as new promising HBV therapy strategy, and a dominant negative mutant of HBVX gene pRev X-GFP which we have reported in our previous study has some effects both on HBV replication and expression in transient expression, but the effects were interfered by persistent secretion of HBV in HepG2 2.2.15 cells without transfection pRev X-GFP in the experiment. To make sure the effects of dominant negative mutant of pRev X-GFP, we established a HBX DN stable express cell clone, and evaluated the effects of HBX dominant negative mutant on HBV replication.

METHODSThe X gene mutant, in which a specific point mutation of 3'-end ATG to AAG and fused with human green fluorescence protein (GFP) were cloned into pRev TRE vector, assigned to pRev HBX-GFP dominant mutant (pRev X-GFP). And the plasmid contains the wild type X gene or GFP gene was cloned into the same vector to construct pRev Xwt, pRev GFP constructs. All the constructs then transfected into HepG2 2.2.15 cells by liposome. After 7 days resistance selection of hygromycin (300 microg/ml), and cell clones which stable expression HBX-GFP, HBXwt, GFP were obtained. After reseeding of 106 cells of each clones in 12 wells with a 12 well cell plate and another 12 wells 2.2.15 cell were serve as blank control. The cells and media were harvested after cultured in DMEM with 10% FBS for 3 days. HBV-related DNA was assayed by dot blot and Southern blot.

RESULTSThe 100% expression of pRev HBX-GFP, GFP and wild type X constructs were obtained. The stable expressed HBX-GFP can significantly reduce HBV DNA level both in cell media and cells by dot blot and Southern blot analysis, but not for pRev Xwt and pRev GFP.

CONCLUSIONThe dominant negative mutant pRev HBX-GFP can significantly inhibit the HBV gene expression. It also suggested that X gene might be one of promising target for HBV gene therapy.

Carcinoma, Hepatocellular ; pathology ; Cloning, Molecular ; DNA Replication ; Genetic Vectors ; Green Fluorescent Proteins ; genetics ; Hepatitis B virus ; genetics ; physiology ; Humans ; Liver Neoplasms ; pathology ; Point Mutation ; Trans-Activators ; genetics ; Transfection ; Tumor Cells, Cultured ; Virus Replication

OBJECTIVETo explore the effect of total parenteral nutrition (TPN) supplemented with arginine on cellular immune function of patients with hepatocellular carcinoma (HCC) after radical tumor resection.

METHODSFifty-six HCC patients undergoing radical surgery received fat-free TPN support, routine TPN or TPN with arginine supplementation, and their clinical data were analyzed prospectively. The percentages of T lymphocyte subpopulation and national killer (NK) cells in peripheral blood are determined, and the levels of interleukin-2 (IL-2), IL-4 and interferon-gamma (IFN-gamma) were measured.

RESULTSNo marked changes were noted in peripheral blood CD4+, CD8+ T cells and NK cells, or in IL-2, IL-4 and IFN-gamma levels after fat-free TPN and routine TPN support. TPN supplemented with arginine resulted in significant increase in CD4+ T cells, NK cells and CD4+/CD8+ T cell ratio in the peripheral blood, as well as in IL-2 and IFN-gamma levels. Peripheral blood IL-4 level was decreased significantly.

CONCLUSIONTPN with arginine supplementation can augment the percentages of CD4+ T lymphocytes and NK cells, and increase IL-2 and IFN-gamma levels, suggesting that arginine can enhance cell-mediated immunity in postoperative patients with HCC.

Adult ; Aged ; Arginine ; pharmacology ; Carcinoma, Hepatocellular ; immunology ; metabolism ; surgery ; therapy ; Cytokines ; metabolism ; Dietary Supplements ; Female ; Humans ; Immunity, Cellular ; drug effects ; Liver Neoplasms ; immunology ; metabolism ; surgery ; therapy ; Male ; Middle Aged ; Parenteral Nutrition ; methods ; Postoperative Period ; T-Lymphocyte Subsets ; drug effects ; immunology

OBJECTIVETo evaluate the mutations of lamivudine-resistance using oligonucleotide microarray in hepatitis B virus (HBV) infected patients.

METHODSA randomized clinical trial was conducted on 20 lamivudine-treated patients for 18 months and 10 patients as controls. The serum HBV DNA was amplified by PCR and the lamivudine-resistance mutations in YMDD region were assayed by 4 sites microarray developed before.

RESULTSThis microarray could clearly differentiate the wide-type from mutated-type HBV with lamivudine-resistance mutations. The rate of mutations in YMDD region increased with the time of lamivudine treatment (chi2=6.69, P<0.01). The most common mutated type was M539V+L515M and next M539I. Continuous administration of lamivudine was no benefit for inhibiting the replication of HBV with YMDD mutation but helpful for wide-type HBV.

CONCLUSIONThe routine serum HBV DNA assay by PCR may introduce prejudice in monitoring HBV inhibitory effect by lamivudine, while the microarray technique can avoid this and is one of the best ways to monitor the lamivudine-resistance mutations in HBV. There is no effect of lamivudine on HBV with YMDD mutation in clinical practice.

Adult ; Antiviral Agents ; pharmacology ; therapeutic use ; Drug Resistance, Viral ; genetics ; Female ; Hepatitis B virus ; drug effects ; genetics ; Hepatitis B, Chronic ; drug therapy ; virology ; Humans ; Lamivudine ; pharmacology ; therapeutic use ; Male ; Middle Aged ; Oligonucleotide Array Sequence Analysis ; Point Mutation ; genetics ; Polymerase Chain Reaction

OBJECTIVETo explore the correlation between the exposure levels and serum protein fingerprints in population exposed to silica.

METHODSLiquid chip time-of-flight mass spectrometry technology was used to investigate the serum profiles in control group (30 cases), group exposed to silica (30 cases), silicosis group (I stage, 25 cases) and suspected silicosis group (30 cases), and screen the differential expression proteins. The correlation between the levels of the differential expression proteins and the exposure levels was performed.

RESULTSFive differential expression proteins were found among 4 groups, the expression of 5081 and 5066 proteins was upregulated, and the expression of 3954, 2021 and 1777 proteins was downregulated. There was no the correlation between the exposure levels and the peak with M/Z among those proteins.

CONCLUSIONthe results of present investigation indicated there was no correlation between the exposure levels and protein/peptide peak.

Adult ; Blood Proteins ; analysis ; Case-Control Studies ; Dust ; analysis ; Female ; Humans ; Male ; Mass Spectrometry ; Middle Aged ; Occupational Exposure ; analysis ; Peptide Mapping ; Proteomics ; Silicon Dioxide ; toxicity ; Silicosis ; blood

Objective To observe the effect of complement fragment C3f on synthesis of α-smooth muscle actin (α-SMA) in human embryonic lung fibroblasts (MRC-5).Methods MRC-5 was cultured with C3f of different concentrations (15.6,62.5,250 ng · mL-1) in low,moderate and high dosage experimental groups.And a positive control of 10 ng· mL-1 transforming growth factor-β1 (TGF-β1) and a negative control of 0 ng · mL-1 C3f were also set up for comparative study.Then α-SMA was detected by immunohistochemistry.Results The moderate dose experimental group and positive control group stimulates the synthesis of α-MRC-5 cell SMA with no statistical difference (7.46 ±0.66 vs 7.54 ±0.51,P ＞0.05).It demonstrated that C3f can stimulate the function of MRC-5 on synthesis of α-SMA.With the increase of C3f concentration,the expression of intracellular α-SMA in three doses experimental groups were increased for 4.95 ± 0.78,7.46 ±0.66 and 9.23 ±0.21,respectively,significant differences in three doses experimental groups (all P ＜ 0.05).Moreover,the concentration of C3f was positively correlated with the expression of α-SMA (P ＜ 0.05).C3f has the function of promoting the expression of α-SMA in MRC-5.Conclusion Complement fragment C3f can stimulate the synthesis of α-SMA in MRC-5 cells.