OBJECTIVE To analyze pathogenic bacterium distribution and antibiotic resistance of our hospital,and provide scientific basis for clinical rational using of antibiotics.METHODS The patients′ clean catch(midstream)(urine) was collected from Jan 2004 to Dec 2005 and cultivated.Antibiotic sensitivity test and adopted by Kirby-Bauer method.RESULTS The pathogenic bacteria mainly consisted of Gram-negatives,among which Escherichia coli was the most frequent,the others in turn were Pseudomonas aeruginosa,Enterobacter cloacae and(Proteus) mirabilis;Enterococcus were the most common among Gram-positives;fungal infection obviously(increased).The bacteria showed different antibiotic resistance rate and multi-drug resistance.CONCLUSIONS It′s very important for making the clinical use of antibiotic more reasonable and controlling drug resistant strains(transmission).

OBJECTIVE To establishan immuno-PCR assay with the carriers of gold-magnetic particles for detection of HIV-1 p24. METHODS The feasibility of using gold-magnetic particles as the carriers was verified. The gold-magnetic particles were coated with mouse anti-p24 monoclonal antibody as the capture antibody. The reporter DNA was initially generated by PCR amplification using a biotinylated primer, and was bound through streptavidin to biotinylated polyclonal antibody as the detection antibody. HIV-1 p24 sandwiched by two antibodies was detected by amplifying the reporter DNA using PCR. RESULTS The efficiency of gold-magnetic particles coated with mouse anti-p24 monoclonal antibody could reach up to 95%. Furthermore, the amount of antibodies immobilization was consistent among different batches of gold-magnetic particles and there was nearly without nonspecific adsorption. The detection limit of immuno-PCR assay was 0.1 ng/L, an approximately 1.5?104-fold higher compared with an enzyme-linked immunosorbent assay. The linear range of p24 concentration was 0.1-100 ng/L. CONCLUSIONS Gold-magnetic particle is one of the ideal immuno-PCR reaction carriers. The immuno-PCR for detection of HIV-1 p24 reported in this article is indicated to be a promising detection method.

Objective To describe a highly sensitive immuno-polymerase chain reaction (immuno-PCR) assay for the detection of human recombinant HIV-p24 antigen. Methods We used gold-magnetic particles as the carriers,mouse anti-p24 monoclonal antibody as the capture antibody and biotinylated goat anti-p24 polyclonal antibody as the detection antibody. The reporter DNA was initially generated by PCR amplification using a biotinylated primer,and was bound with streptavidin to biotinylated polyclonal antibody. Human recombinant p24 antigen sandwiched by antibodies was detected by amplifying the reporter DNA using PCR. The optimal concentration of sreptavidin and DNA label were determined using square titration. The electrophoresis gels were imaged and analyzed by Quantity One software. Results The optimal concentration of sreptavidin and DNA label were determined to be 0.1 mg/ L and 10 ng/ L,respectively. The detection limit of the immuno-PCR assay was 0.1 ng/L,higher than that of the conventional ELISA. Conclusion A highly sensitive immuno-PCR for human recombinant HIV-p24 antigen was indicated to be an available method for early screening of HIV infectors in blood donors.

OBJECTIVE To evaluate primarily the detection limit,specificity and reproducibility of immuno-PCR assay on HIV-1 p24 antigen.METHODS We p24 antigen were detected by established immuno-PCR system,and then the detection limit,specificity and reproducibility were discussed.We quantitatively analyzed the nonspecific amplified bands with fluorescence intensity(FI),and a preliminary determination of the lower specific amplification limit was made.RESULTS We taken(x-+3s)FI of nonspecific amplification as the lower limit of specific amplification signal.The detection limit of immuno-PCR assay was 0.1 ng/L.CONCLUSIONS The detection limit,specificity and reproducibility can meet the needs of HIV-1 p24 antigen detection.

OBJECTIVE To investigate the distribution and the drug resistance changes of the pathogenic bacteria isolated from the blood culture specimens collected during the period of 2006-2008.METHODS Blood culture of patients in our hospital was performed by BacT/Alert 240 and the isolated bacteria were identified by API and Microscan and tested for drug resistance against antimicrobial agent by K-B method.A retrospective analysis was made to the blood culture results during the period of 2006-2008 with WHONET 5.4 software.RESULTS Gram-negative rods were the predominant bacteria which caused sepsicemia.The isolated rates of Escherichia coli took the first place during the period of 2006-2008.Klebsiella pneumoniae,Pseudomonas aeruginosa and Staphylococcus aureus(SAU) were also the most important pathogens which caused blood infection.The infection rate of coagulase negative staphylococcus(CNS) and P.aeruginosa had increasing tendency.Imipenem and meropenem were the most effective antibiotics(100%) to E.coli and K.pneumoniae.Then amikacin and cefoxitin also had high susceptibility to E.coli and K.pneumoniae.Furthermore,drug susceptibility in 2008 was higher than that of two years before.All antibacterials had low drug susceptibility to P.aeruginosa,and its drug resistance rate rised obviously.The drug resistance situation of Acinetobacter baumannii was serious.Except imipenem and meropenem had relatively higher susceptibility(20-69.2%),the susceptibility to other antibacterials was lower than 41.7%.Vancomycin,teicoplanin and quinupristin/dalfopristin were the most effective antibiotics to(SAU).CONCLUSIONS The species and drug resistance of the bacteria isolated from blood specimens have changed.More attention should be paid to the detection and surveillance of bacterial resistance in blood culture to promote the rational use of antibiotics.

OBJECTIVE To investigate whether stromal cell-derived factor-1α (SDF-1α) might be able to prevent senescence of endothelial stem cell (ESC) and also study its effects on the telomerase activity. METHODS Total mononuclear cells (MNCs) were isolated from peripheral blood by density gradient centrifugation, and then the cells were plated on fibronectin-coated culture dishes. After cultured for 4 d, attached cells were divided into control and SDF-1α 1, 10, 50 and 100 μg·L~(-1) groups. ESC became senescent as determined by acidic β-galactosidase staining. The proliferation of ESC was assessed by MTT assay and colony-forming capacity. Telomerase activity was measured by telomerase-PCR ELISA and the phosphorylation of Akt was determined by using Western blotting. RESULTS Ex vivo prolonged cultivation of ESC led to rapid onset of ESC senescence. Compared with control group, SDF-1α concentration-dependently inhibited the onset of ESC senescence, maximum at 100 μg·L~(-1) (40.8±7.1 vs 17.5±3.0; P<0.01). Moreover, SDF-1α 100 μg·L~(-1) increased ESC proliferation (0.22±0.02 vs 0.39±0.04; P<0.01) and ESC colony-forming activity (7.8±2.2 vs 22.4±3.4). Compared with control group, SDF-1α 100 μg·L~(-1) also increased telomerase activity (0.34±0.05 vs 0.57±0.09; P<0.01). In addition, SDF-1α treatment of ESC stimulated a concentration- and time-dependent Akt phosphorylation. CONCLUSION SDF-1α-induced prevention of ESC senescence leads to the potentiation of proliferative activity, and clonal expansion, which may be related to the activation of telomerase and Akt phosphorylation.

BACKGROUND:The use of three-dimensional cel culture techniques can better simulate the cel ular microenvironment, providing new tools for tissue engineering research.
OBJECTIVE:To analyze the biomaterial selection and application characteristics in three-dimensional cel culture as wel as applications in tumor tissue engineering.
METHODS:We searched Wanfang database and PubMed database 1998-2015 years for relevant literature using keywords of“three-dimensional cultures;scaffold;cel growth;cel differentiation;tumor tissue engineering”in Chinese and English, respectively.
RESULTS AND CONCLUSION:The selection and application of three-dimensional scaffold materials is one of the keys. So far, scaffold materials, such as col agen gels, gelatin sponge, agarose, chitosan, demineralized bone matrix, cannot provide the extracel ular matrix similar to the micro-environment in which seed cel growth and proliferation are not affected, and the ability to secrete type II col agen and glycosaminoglycan is decreased, although they can provide three-dimensional space for seed cel s. Biomimetic scaffold characterized as little trauma and strong plasticity gradual y shows its unique advantages. Three-dimensional culture conditions raise pro-angiogenic growth factor secretion from tumor cel s, and this feature is positively correlated with the occurrence of in vivo tumor angiogenesis.

Objective To screen the non-nucleoside compounds against HIV-1 reverse transcriptase by molecular modeling and bioactivity assay. Methods Surflex-Dock module of Tripos SYBYL software was used to simulate the binding pattern of 22 000 compounds in SPECS database with the active pocket of HIV-1 reverse transcriptase. Based on the simulation results, the interaction mode between the above compounds and the crystal structure of HIV-1 reverse transcriptase was analyzed. The compounds with higher docking scores and better binding pattern were determined by anti-HIV-1 ac tivities test in vitro. Results The virtual screening results showed that the docking conformation of 1- (4-fluorophenyl) -3- [2- (1H-indol-3-yl) ethyl] thiourea was similar to the embedded ligand in Rilpivirine crystal structure. 1- ( 4-fluorophenyl) -3- [ 2- ( 1H-indol-3-yl) ethyl] thiourea was held together with the key residue Lys101 in docking pocket of HIV-1 reverse transcriptase by hydrogen bonds, and hadπ-πstacking action together with the conservative residue Trp229 and the aromatic residue Tyr181 respectively. The bioassay in vitro results showed that when the proliferation rate of C8166 lymphocyte syncytium infected by HIV-1ⅢB arrived 50% ( EC50) , the concentration of 1- ( 4-fluorophenyl) -3- [ 2- ( 1H-indol-3-yl) ethyl] thiourea was 5.45μg/mL. Conclusion Molecule docking technology is an effective approach to reducing the screening of candidate compounds with micromolecular activity, and can be used to predict the interaction mode between the compound and the target receptor. In the study, active compound 1- (4-fluorophenyl) -3- [2- (1H-indol-3-yl) ethyl] thiourea has been screened out by molecule docking technology.

Objective To analyze the local control rate and the desimetric patterns of local recurrence in nasopharyngeal carcinoma(NPC)patients having been treated with standardized conventional radiotherapy and to evaluate the delineation of rational target volume.Methods From Jan.2000 to Dec.2000,476 patients with untreated NPC were treated by standardized conventional radiotherapy alone at the Sun Yat-sen University Cancer Center.The radiation ports were designed on a X-ray simulator.The nasopharyngeal lesion demonstrated by CT scan and the subclinical spread regions adjacent to the nasopharynx were defined as the target volume.Kaplan- Meier method was used to calculate the cumulative local recurrence rate.For patients with locad recurrence,the primary and recurrent local tumor volumes(V_(nx),V_(recur))were delineated with three-dimensional treatment planning system(3DTPS),and the dataset of radiation ports and delivered prescription dose to the 3DTPS were transferred according to the first treatment.The dose of radiation received by V_(recur)was calculated and analyzed with dose- volume histogram(DVH).Local recurrence was classified as:1.“in-port”with 95% or mere of the recurrence volume((recur)_V_(95))was within the 95% isedase;2.“marginal”with 20% to95% of _(recur)V_(95)within the 95% isedese; 3.“outside”with only less than 20% of _(recur)_V_(95)within the 95% isodose curve.Results With the median follow- up of 42.5 months(range 8～54 months),52 patients developed local recurrence.The 1-,2-,3 and 4-year cumulative local failure rate was 0.6%,3.9%,8.7% and 11.5%,respectively.Among the 42 local recurrent patients who could be analyzed by 3DTPS,52% were in-port,40% were marginal and 7% were outside.For most of the marginal recurrence and all the outside recurrence patients,the main reason of recurrence were related to the unreasonable design of the radiation port and inaccuracy in the interpretation image findings.Conclusions The outcome of better local control rate and the dosimetric pattern of local recurrence show that the target volume is reasonable for NPC in Sun Yat-sen University Cancer Center.Enhancing the capability of correct interpretation of images,accurate design of the radiation pouts and making most useful molecular or functional imaging techniques to escalate the local radiation dose are promising ways to improve the local control further and better.

OBJECTIVETo examine the expression of membrane-type-1 matrix metalloproteinase (MT1-MMP) and reversion-inducing cysteine-rich protein with Kazal motifs (RECK) in gastric carcinoma, and investigate its clinical significance, at the same time analyze the correlation between MT1-MMP and RECK expression.

METHODSMT1-MMP and RECK expression in surgically resected tissue samples of gastric carcinoma was examined by immunohistochemical method (two-step method) , and its correlation with clinicopathological factors was analyzed.

RESULTSAmong the 44 gastric carcinoma samples, 37 (84.1%) were stained positive for MT1-MMP, and 31 (70.5%) for RECK. The expression of MT1-MMP was much higher in poorly differentiated gastric carcinoma samples than moderately and well-differentiated samples (P = 0.015). The expression level of MT1-MMP was associated with invasive depth of tumor cells (P = 0.007), but no difference between sex and lymph node metastasis. On the contrary, the well-differentiated samples showed higher expression of RECK than poorly and moderately differentiated gastric carcinoma samples (P = 0.006). The expression level of RECK did not correlate with sex, lymph node metastasis and invasive depth of tumor cells. RECK expression showed no relation to MT1-MMP expression in the gastric carcinoma.

CONCLUSIONOverexpression of MT1-MMP in gastric carcinoma may play an important role during tumor differentiation and metastasis, the RECK protein may have positive effects on the tumor differentiation.

Adult ; Aged ; Aged, 80 and over ; Carcinoma ; enzymology ; genetics ; metabolism ; pathology ; Female ; GPI-Linked Proteins ; genetics ; metabolism ; Gene Expression Regulation, Neoplastic ; Humans ; Male ; Matrix Metalloproteinase 14 ; genetics ; metabolism ; Middle Aged ; Stomach Neoplasms ; enzymology ; genetics ; metabolism ; pathology