OBJECTIVETo gain insights into the role of forkhead box protein 3 (Foxp3) in the pathogenesis of hepatocellular carcinoma (HCC) by performing a comparative analysis of Foxp3 mRNA expression and promoter methylation status in HCC and normal liver tissues.

METHODSThirty-nine HCC and 13 normal liver tissue specimens were evaluated by real-time quantitative PCR and pyrosequencing to measure the expression of Foxp3 mRNA and determine the methylation status of its promoter, respectively. Statistical analyses of the data were conducted by rank-sum test and Spearman's rank correlation coefficient test.

RESULTSThe HCC specimens showed significantly higher mRNA expression of Foxp3 (vs. normal liver tissues, Z =-2.770, P =0.0056). Moreover, the HCC specimens showed significant hypomethylation of the Foxp3 promoter site A (vs. normal liver tissues, Z =2.118, P =0.0339), and the Foxp3 mRNA level was negatively correlated with the methylation of site A (rs =-0.344, P =0.046). None of the other four sites in the Foxp3 promoter showed a significant difference in methylation, and the overall methylation was not significantly different between the HCC and normal liver tissues.

CONCLUSIONOverexpression and low methylation of Foxp3 may be involved in the oncogenic and progression processes of HCC.

Carcinoma, Hepatocellular ; metabolism ; pathology ; CpG Islands ; DNA Methylation ; Female ; Forkhead Transcription Factors ; genetics ; metabolism ; Humans ; Liver ; metabolism ; pathology ; Liver Neoplasms ; metabolism ; pathology ; Male ; Middle Aged ; Promoter Regions, Genetic ; RNA, Messenger ; genetics

OBJECTIVE: To evaluate the reliability and clinical practicability of cefoxitin disk diffusion test in the detection of methicillin-resistant staphylococcus (MRS) heterogenic drug-resistant strains. METHODS: Three hundred and ten strains of staphylococcus isolated from clinics were detected by the oxacillin disk diffusion test, the cefoxitin disk diffusion test as well as the oxacillin agar dilution test according to the standard operation procedures of NCCLS, and the detection of mecA gene of staphylococcus was used as a criterion. The sensitivities and specitivities of the 4 methods were compared. RESULTS: By the detection of mecA gene, the ratio for MRSA was 57.1%(113/198) and the ratio for MRCNS was 62.5%(70/112). Both the sensitivity and specificity of cefoxitin disk diffusion test in the detection of MRS were 100%, and those in the detection of MRCNS were 98.6% and 100%. CONCLUSION Cefoxitin disk diffusion test is reliable, simple and convenient, and it can be used as a conventional method for the detection of MRS in clinical laboratories.
Anti-Bacterial Agents ; pharmacology ; Bacterial Proteins ; genetics ; Cefoxitin ; pharmacology ; Humans ; Methicillin ; pharmacology ; Methicillin Resistance ; genetics ; Microbial Sensitivity Tests ; instrumentation ; methods ; Oxacillin ; pharmacology ; Penicillin-Binding Proteins ; Reproducibility of Results ; Staphylococcus aureus ; drug effects ; genetics ; isolation & purification

OBJECTIVE: To determine the distribution of pathogens and their characteristics of drug susceptibility originating from nosocomial infections in the intensive care units (ICU), and to provide evidence for clinical anti-infection treatments. METHODS: Retrospective analysis to the pathogens and their drug susceptibility characteristics was carried out. These pathogens were isolated from the samples that came from patients infected in the ICU from 2002 to 2004. RESULTS: The main nosocomial infective pathogens in the ICU were gram negative bacilli (48.2%), and the next ones were gram positive bacteria (43.3%) and fungus (8.5%). The most common gram negative bacilli were Pseudomonas aeruginosa and Escherichia coli; while for gram positive bacteria, the main bacterin were Staphylococcus aureus. The gram negative bacilli could resist 4 or more than 4 antibiotics, and the rate for resistance exceeded 40%. Similarly, oxacillin resistance staphylococcus could resist 7 antibiotics, and the rate was over 50%. The detective rates of ESBLs and AmpC enzymes produced by Escherichia coli and K. peumoniae were 34.0% & 30.7% and 13.2% & 23.1%, respectively. The rate for oxacillin resistance staphylococcus was 66.3%, and there was relative high resistance rate ( > 55%) for most antibiotics: There was statistical difference, compared with that of non-resistant strains. CONCLUSION Though gram positive coccus still play an important role, most infections are caused by gram negative bacilli of nosocomial infections in the ICU. The antibiotics resistant rate of all bacteria has been rising gradually. It shows strong resistance and multi-drug resistance. The most importment cause for resistance of gram negative bacilli is that the bacteria can produce ESBLs and AmpC enzymes. The antibiotic resistant rate for oxacillin resistance staphylococcus is really high.
Adolescent ; Adult ; Aged ; Child ; Child, Preschool ; Cross Infection ; microbiology ; Drug Resistance, Microbial ; Drug Resistance, Multiple ; Escherichia coli ; drug effects ; isolation & purification ; Female ; Gram-Negative Bacteria ; drug effects ; isolation & purification ; Humans ; Intensive Care Units ; Male ; Microbial Sensitivity Tests ; Middle Aged ; Oxacillin ; pharmacology ; Pseudomonas aeruginosa ; drug effects ; isolation & purification ; Retrospective Studies ; Staphylococcus aureus ; drug effects ; isolation & purification

OBJECTIVE: To examine the infection and bacteria resistance of Helicobacter pylori (H.pylori) to clarithromycin and furazolidone,to determine whether the antibiotic resistance is primary or secondary, and to decide if a new H.pylori infection plays a role in eradication failures. METHODS: Twenty one H.pylori had been isolated from human biopsy specimens, and antimicrobial susceptibility testing was performed. DNA fingerprints were generated using random amplification polymorphic DNA (RAPD) to determine the identity of H.pylori before and after the eradication therapy. RESULTS: Eight bacteria resisted against clarithromycin, and one against furazolidone, with the resistant rates 38.1% and 4.8% respectively. The number of primary antibiotic resistance, secondary resistance and new infection was 1 for each. CONCLUSION Resistance to clarithromycin is more common compared with that to furazolidone. Development of primary and secondary resistance to clarithromycin occurs as a rule in eradication failures. New H.pylori infection plays a role in eradication failures.
Anti-Bacterial Agents ; pharmacology ; therapeutic use ; Clarithromycin ; pharmacology ; therapeutic use ; DNA Fingerprinting ; DNA, Bacterial ; analysis ; genetics ; isolation & purification ; Drug Resistance, Bacterial ; Furazolidone ; pharmacology ; therapeutic use ; Helicobacter Infections ; drug therapy ; microbiology ; Helicobacter pylori ; drug effects ; genetics ; Humans ; Microbial Sensitivity Tests ; Random Amplified Polymorphic DNA Technique

OBJECTIVE: To determine the effect of CagA(+) Helicobacter pylori(H.pylori)strain and anti-H.pylori drugs on the expression of connexin 43(Cx43) and cell proliferation of BGC-823 cells in vitro,and to investigate the relation between the changes of Cx43 expression, cell proliferation of BGC-823 cells and CagA(+)H.pylori. METHODS: BGC-823 cells were co-cultured with CagA(+) H.pylori strain(NCTC J99) or CagA(-) H.pylori strain(NCTC 12908)at bacteria/cells ratio of 20:1,100:1 and 500:1 for 24 hours and 48 hours respectively. anti-H.pylori drugs was given in the group co-cultured at bacteria/cells ratio of 100:1 after 16 hours. In the control group, BGC-823 cells were cultured for 24 hours and 48 hours respectively,but without H.pylori or antij H.pylori drugs. Immunocytochemical SABC method and the image analysis of the computer were applied to detect the changes of Cx43 expression in BGC-823 cells. The cell proliferation was examined by methyl tetrazolium (MTT) method. RESULTS: (1)The expression of Cx43 in the control group after cultivation for 48 hours was higher than that for 24 hours (P< 0.05). The expression of Cx43 in the groups co-cultured with CagA(+) H.pylori strain after cultivation for 48 hours was lower than that co-cultured for only 24 hours, and that of the groups co-cultured with CagA(+) H.pylori strain was lower than that of the control group for both 24 hours and 48 hours (P< 0.05). The expression of Cx43 in the groups at bacteria/cells ratio of 500:1 was lower than that at bacteria/cells ratio of 20:1 and 100:1 for both 24 and 48 hours (P< 0.05),and that at bacteria/cells ratio of 100:1 was lower than that at bacteria/cells ratio of 20:1 for 48 hours (P< 0.05).However, there was no significant difference in Cx43 expression between 24 and 48 hours in the groups co-cultured with CagA(-) H.pylori strain (P>0.05). Cx43 expression in the groups co-cultured with CagA(-) H.pylori strain at the ratio of 100:1 and 500:1 was lower than that in the control group, and Cx43 expression at the ratio of 500:1 was lower than that at the ratio of 20:1 for 24 hours and 48 hours. Cx43 expression increased after the intervention with anti-H.pylori drugs for 48 hours. (2) In the groups co-cultured with CagA(+)H.pylori strain, the optical density value of MTT indicated that the cell proliferation at the bacteria/cells ratio of 100:1 was higher than that in the control group, but no significant difference was found in other two groups co-cultured for 24 hours. After co-culturing for 48 hours, the cell proliferation at the bacteria/cells ratio of 20:1 and 100:1 was significantly accelerated, while the cell proliferation at 500:1 was inhibited. In the groups co-cultured with CagA(-) H.pylori strain,there was no change in the cell proliferation. Intervention with anti-H.pylori drugs could suppress the cell proliferation. CONCLUSION CagA(+) H.pylori can down-regulate the expression of Cx43 in BGC-823 cells,which is related to the reaction time and the density of H.pylori. Low density of CagA(+)H.pylori suspensions can accelerate the proliferation of BGC-823 cells, while high density can suppress the cell proliferation. The CagA(-) H.pylori has no effect on the cell proliferation. Intervention with anti-H.pylori drugs can up-regulate the expression of Cx43,and suppress the cell proliferation of BGC-823 cells.
Anti-Bacterial Agents ; pharmacology ; Antigens, Bacterial ; genetics ; metabolism ; Bacterial Proteins ; genetics ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Connexin 43 ; biosynthesis ; Helicobacter pylori ; drug effects ; genetics ; metabolism ; Humans ; Immunohistochemistry ; Stomach Neoplasms ; metabolism ; microbiology ; pathology

OBJECTIVETo find sensitive and specific micro-metastic markers for prostate cancer.

METHODSUsing nested reverse transcription-PCR, we examined the expression of PSA, hK2 and PSMA mRNA in peripheral blood mononuclear cells of 51 patients with prostate cancer, 33 patients with benign prostate hyperplasia (BPH) and 32 normal young people.

RESULTSThe expression rates of PSA, hK2 and PSMA mRNA were 52.9%, 43.1% and 64.7%, respectively in prostate cancer group, and 6.2%, 7.7% and 4.6%, respectively in control group (BPH patients and normal young people) with statistical significance (P < 0.01). Although the expression rate of PSA and hK2 mRNA increased with cancer progression, there was no statistical significance among patients in different stages. The expression rate of PSMA mRNA was higher than that of PSA and hK2 mRNA in each clinical stage.

CONCLUSIONPSMA mRNA expression detected by nested RT-PCR is of greater value for the diagnosis, therapy choice and prognostic evaluation of prostate cancer patients.

Aged ; Antigens, Surface ; blood ; Biomarkers, Tumor ; blood ; Glutamate Carboxypeptidase II ; blood ; Humans ; Male ; Middle Aged ; Neoplasm Invasiveness ; Neoplasm Staging ; Prostate-Specific Antigen ; blood ; Prostatic Hyperplasia ; blood ; pathology ; Prostatic Neoplasms ; blood ; pathology ; Tissue Kallikreins ; blood

OBJECTIVETo investigate flap survival after transfection using gene encoding vascular endothelial growth factor (VEGF).

METHODSIn 30 Sprague-Dawley rats, the anterior abdominal skin flap supplied by the epigastric vessels was created. The animals were divided into three groups, with ten of each. The first group was treated with a mixture of liposomes and the cDNA encoding the 165-amino acid isoform of VEGF; the second group was treated with control blank plasmid DNA and liposome transfection medium; the third group was treated with physiological saline. Four days after injection, the epigastric artery and vein were ligated and the blood flow in the flap was evaluated by intraperitoneal injection of fluorescence solution. Seven days later, the survival area of the flap was measured by planimetry. After the animals were killed, specimens were harvested from the anterior abdomen skin flap for immunohistological evidence of VEGF expression and for hematoxylin and eosin staining of microvascular growth.

RESULTS30 minutes after pedicle ligation the average fluorescence staining planimetry of the three groups (PCD-VEGF165, PCD and physiological saline) was 60.64%, 30.15% and 29.89% respectively. Tissue survival planimetry of the three groups was 92.3%, 30.5%, 31.8%. There was significant difference between the first group and the latter two (P < 0.05). Immunohistochemical staining documented increased deposition of VEGF cDNA in the first group compared to the control groups (P < 0.05). Normal staining documented that the average vessel number of the three groups was 101.72, 91.35 and 89.85 (P < 0.05), the average vessel lumen diameter was 26 microns, 31.09 microns and 32.51 microns(P < 0.05).

CONCLUSIONThrough administration, PCD-VEGF165 can transfect the anterior abdominal skin flap and enhance its survival. There was express of VEGF protein in the treated flap.

Animals ; Biomarkers ; metabolism ; DNA, Complementary ; administration & dosage ; Epigastric Arteries ; Genetic Therapy ; Graft Survival ; physiology ; Liposomes ; Plasmids ; Rats ; Rats, Sprague-Dawley ; Surgical Flaps ; blood supply ; physiology ; Time Factors ; Transfection ; methods ; Vascular Endothelial Growth Factor A ; genetics ; metabolism

Recombineering, a new genetic engineering technology based on high efficiency in vivo homologous recombination, can be used in target DNA knock-in, knock-out and gene cloning. In the process of gene subcloning mediated by Recombineering technique, high-quality target DNA fragments were difficult to obtain using in vitro overlapping PCR,therefore the efficiency of in vivo homologous recombination was severely interrupted. To solve this problem, some technology improvements have been established based on the principle of Red recombinases. The PCR DNA fragments of egfp and kan genes with complementary sequences on the end of each fragment were co-introduced into a pcDNA3.1 vector and Red recombinases containing E. coli DY331 host cells by electroporation. A recombinant plasmid pcDNA3.1-egfp-kan was screened directly by antibiotic marker. The positive rates can reach to 45%. The EGFP gene expression of pcDNA3.1-egfp-kan can be observed by transient transfection of 293 eukaryotic cells.
Bacteriophage lambda ; enzymology ; genetics ; DNA, Recombinant ; genetics ; Electroporation ; Escherichia coli ; genetics ; Gene Fusion ; genetics ; Genetic Engineering ; methods ; Green Fluorescent Proteins ; genetics ; metabolism ; Plasmids ; genetics ; Recombinases ; genetics ; metabolism ; Recombination, Genetic

Objective To analyze the characteristics of clinical manifestations, risk factors, therapies and acute outcomes in patients with cerebral venous sinus thrombosis complicated by cerebral hemorrhage. Methods Seventy-five patients with cerebral venous sinus thrombosis were included in the study. According to the radiological findings on the brain image, patients were divided into two subgroups:cerebral hemorrhage group and non-hemorrhage group. The demo?graphic data, potential risk factors, clinical manifestations, radiological features, therapeutic strategies and acute out?comes were compared between two subgroups, and high risk factors were also analyzed. Results There were seventy-five patients with cerebral venous sinus thrombosis in the present study. Twenty-eight patients of them (37.2%) had cerebral hemorrhage whereas the remaining forty-seven patients (62.7%) did not have cerebral hemorrhage. Pregnancy/puerperi?um were significantly higher in patients with cerebral hemorrhage (with vs without;28.6%vs. 6.4%, P=0.015), while in?fection was markedly higher in patients without cerebral hemorrhage (with vs without;7.1% vs. 29.8%, P=0.021). Head?ache (92.9% vs. 70.2%, P=0.021), unconsciousness (25.0% vs. 6.4%,P=0.034), seizures (53.6% vs. 19.1%, P=0.002) and motor deficits (35.7% vs. 12.8%, P=0.019) were more common in patients with cerebral hemorrhage. Moreover, mul?tiple sinus involvement (1.4% vs. 44.7%, P=0.024) was significantly higher and the acute outcomes(mRS≥3: 46.4%vs.17.0%, P=0.006)were poorer in patients with cerebral hemorrhage. Binary Logistic analysis showed that pregnancy/pu?erperium (P=0.004) and multiple sinus involvement were positively, whereas infection was negatively correlated with cere?bral venous sinus thrombosis and hemorrhage ( P=0.007;P=0.03). Conclusions Pregnancy/puerperium, headache, uncon?sciousness, seizures, motor deficits and multiple sinus involvement are more frequently in patients with cerebral venous sinus thrombosis and hemorrhage, and the acute outcomes are poorer in patients with cerebral venous sinus thrombosis complicated by cerebral hemorrhage.

OBJECTIVETo analyze the characteristic of bacterial infections, and the relationship between antibiotics treatment and bacterial infections after liver transplantation, and to prevent antibiotic-resistant bacterial infections.

METHODS86 liver transplant recipients were retrospected. Different indexes including limited daily dose, the frequency of medication, drug use index were used to evaluate the rationality of the use of antibiotics, three-dimensional test was used to explore extended-spectrum beta-lactamase and AmpC enzyme of Gram-negative bacteria.

RESULTSThe major pathogens of infection after liver transplantation were Enterococcus faecalis, Enterobacter cloacae, fungi and E. coli. Pre-operative antibiotic utilization rate was 83.7%, it was mainly a single use of antibiotics; After- operative antibiotic usage was 100.0%, it was mainly joint use of two or three antibiotics; The top 3 antibiotics used were cephalosporins, the combined enzyme inhibitors and penicillin. Antibiotics with drug utilization index (DUI) more than 1.1 included ampicillin and Lalin proxy. 43.3% and 31.8% of Gram -Negative bacteria produced ESBLs and AmpC, respectively, while 21.3% Gram -Negative bacteria produced two enzymes.

CONCLUSIONThere is high incidence of bacterial infections after liver transplantation. The use of antibiotics is high dose, high-frequency and reasonable; High resistance of bacterial infections was prone to develop and the prevention of the high resistance of bacterial infections is very important.

Adolescent ; Adult ; Aged ; Anti-Bacterial Agents ; administration & dosage ; therapeutic use ; Bacterial Infections ; drug therapy ; etiology ; microbiology ; Child ; Child, Preschool ; Drug Resistance, Bacterial ; Female ; Gram-Negative Bacteria ; drug effects ; enzymology ; isolation & purification ; Gram-Positive Bacteria ; drug effects ; enzymology ; isolation & purification ; Humans ; Infant ; Liver Transplantation ; adverse effects ; methods ; Male ; Microbial Sensitivity Tests ; Middle Aged ; Postoperative Complications ; drug therapy ; epidemiology ; microbiology ; Retrospective Studies ; Young Adult ; beta-Lactamases ; biosynthesis