Objective To explore the effects of chlamydia and mycoplasma infections on the healing of wounds after surgical operation for haemorrhoids,fistula and anoschisis.Methods Sixty-two patients with haemorrhoids,fistula and anoschisis undergoing the surgical operation during 2000.1 to 2008.10 were collected,and wounds did not heal 40 days after operation and the wound's surface infection occurred after postoperative anti-inflammatory therapy and dressing change.All patients were positive at least for laboratory Ct or Uu,and those with infections caused by other fungi,bacteria,viruses and the other systemic diseases were excluded.The correlation between the infections in 62 cases and sex,surgical types was analyzed.Results In 62 patients,there were 30 cases (48.4%) positive for both Ct and Uu,22 cases for single Ct (35.5%),and 10 cases for single Uu (16.1%),respectively.The infection rate in females was higher than in males (P<0.01).The surgical types included:surgery for 8 cases of haemorrhoids (12.9%),for 18 cases of fistula(29.0%),and for 36 cases of anoschisis(58.1%).respectively.Conclusion There is the possibility of chlamydia and mycoplasma infections in the patients with delayed healing of wounds following the surgical treatment on the anal region.Early diagnosis,early treatment,avoidance of ineffective medicines and repeatedly management of the wounds can shorten the healing period.

Objective To determine the effect of cheA gene of Helicobacter pylori in the bacterial chemotaxis in vitro and colonization in vivo. Methods The entire cheA and cheY genes were amplified and cloned from genomic DNA of H. pylori NCTC11637 strain. Subsequently, the prokaryotic expression systems of cheA and cheY genes were generated and the target recombinant proteins rCheA and rCheY were extracted by Ni-NTA affinity chromatography. Rabbits were immunized with either rCheA or rCheY for obtaining antisera, and rCheA-IgG and rCheY-IgG in the antisera were prepared using ammonium sulfate precipitation plus DEAE-52 column chromatography. A suicide plasmid of cheA gene was constructed and then a cheA gene knock-out mutant ( cheA - ) was generated based on homologous recombinant exchange using the suicide plasmid. The cheA- mutant was identified using PCR and sequencing. The phosphorylation levels of CheA and CheY molecules of cheA - and wild-type strain were determined by using rCheA-IgG and rCheY-IgG anchoring the target proteins and protein phosphorylation detection kit. The differences of chemotaxis in vitro and colonization in vivo between cheA- mutant and wild-type strain were compared using chemotactic model and BALB/c infection model of H. pylori. Results The cheA gene knock-out in genome of cheA- mutant was confirmed by the results of PCR and sequencing. After treated with 0. 001-0. 1 mol/L HCI for 10 min, the phosphorylation levels of CheA and CheY molecules of wild-type strain were rapidly descended from ( 59.6 ±11.5) μmol and (55.5 ± 10.2) μmol to ( 10.8 ± 2.6) and (5. 5 ± 1.2) μmol (P ＜ 0.05 ), while the phosphorylation of CheY molecule of cheA - mutant was no markedly changed with a persistent lower level ( P ＞0.05). The diameters [(10-20) ± (2-3) mm] of chemotactic aggregative rings of cheA- mutant were significantly less than those [(16-24) ± (2-3)mm] of wild-type strain (P ＜0.05). The positive isolation rate (90%) of H. pylori in gastric biopsy specimens of mice that infected with wild-type strain was remarkably higher than that (40%) of mice that infected with cheA- mutant (P ＜0.05). The result of fluorescence quantitative was also showed that the numbers (6.3 × 103 ±2.1 × 103 copies/mg) of H. pylori in gastric biopsy specimens of wild-type strain infected mice were significantly larger than those (8.3 × 101 ±3. 1 × 101 copies/mg) in gastric biopsy specimens ofcheA- mutant infected mice (P＜0.05). Conclusion The cheA gene of H. pylori has an important role in the bacterial chemotaxis in vitro and colonization in vivo.

Objective To investigate the role of Stathmin in pancreatic cancer invasion and metastasis and its relationship with DNA methylation. Methods Immunohistochemical detection of MBDI and Stathmin protein expression in 40 cases of pancreatic cancer and 15 cases ot normal pancreatic tissue were performed,followed by analysis of their clinical and pathological relationship with pancreatic cancer; Human pancreatic cancer cell line BxPC-3 was treated with 5-Aza-2-dC (AZA).Both qRT-PCR and Western blot analysis of Stathmin expression were used before and after AZA treatment; Stathmin-siRNA transfected BxPC-3 cells were divided into the Stathmi-siRNA group and the empty vector control group.Transwell chamber invasion assay and animal experiment were performed to measure the changes in cell invasion and metastatic capability. Results lmmunohistochemistry showed positive MBDI and Stathmin expressions in 28 (70％) and 24 (60％) out of 40 cases of pancreatic cancer,respectively,which were significantly higher than that in the normal pancreatic tissue (P＜ 0.05); MBDI and Stathmin protein expressions were positively correlated (r =0.356,P =0.037),so were MBDI expression and lymph node metastasis (P=0.023).Stathmin expression was significantly correlated with clinical staging and lymph node metastasis (P =0.002,and P =0.001,respectively).After AZA treatment,both Stathmin mRNA and protein expression in BxPC-3 were significantly decreased.Transwell chamber invasion assay showed that compared with the control group,the cell invasion capability of the Stathmin-siRNA group was significantly decreased (P＜0.05).Animal experiment showed that the incidence of liver metastasis was significantly lower in the Stathmin-siRNA transfected group than the empty vector control group (P＜0.05).Conclusion Demethylation may contribute to the reduction of Stathmin expression in pancreatic cancer and further improve the prognosis of pancreatic cancer patients.

OBJECTIVE To study the distribution and antimicrobial resistance of Acinetobacter baumannii infection in our hospital.METHODS Five hundred and sixty-nine strains of A.baumannii isolated from patients with infection from Jan 2001 to Dec 2005 were collected and antimicrobial susceptibility tests were performed.RESULTS Two hundred and seventy-five strains(48.3%) of A.baumannii were from intensive care unit(ICU).Four hundred and five strains(71.2%) of A.baumannii were examined from sputum.A.baumannii had various drug resistances to 12 antibiotics,which were monitored and proved tending to strengthen.The resistance rate in the ICU was distinctly higher than the others with significant difference(P

OBJECTIVE To investigate the vicissitudes of resistance to imipenem in Gram-negative bacilli isolated from clinical specimens.METHODS Gram-negative bacilli were isolated from various clinical specimens in our hosptial from Jan 2004 to Jan 2008.The identification and antimicrobiol susceptiblity test was determined by VITEK-AMS full automated microbiology analyzer.RESULTS A total of 6983 strains of Gram-negative bacilli were isolated,including 92 species and 2986 isolates of Enterobacteriaceae and 3961 isolates of non-fermenters,accounted for 42.8% and 56.7%,respectively.The resistant rate of Enterobacteriaceae to imipenem was lower than 1.0%.The resistant rates of Stenotrophomonas maltophilia,Burkholderia cepacia,Chryseobacterium meningosepticum,Pseudomonas aeruginosaand Acinetobacter baumannii to imipenem were 98.4%,97.0%,98.2%,70.1% and 47.5%,respectively.CONCLUSIONS Non-fermenters are predominant pathogens.Imipenem has high antimicrobial activity to Enterobacteriaceae in invro,but not to non-fermenters.The resistant rates of P.aeruginosa and A.baumannii to imipenem are increasing gradually.

Aim To evaluate the efficacy of TFU as a precursor of 5-FU on the growth inhibition of human gastric carcinoma cell lines SGC-7901 and MKN-45.Methods In vitro experiments,cell growth inhibition was measured by MTT assay.The rates were compared in the presence and Absence of liver microsomal enzymes.The morphology of apoptotic cells was detected by observation with a fluorescence microscope after staining by using adridine orange/ethidium bromide solution.DNA fragmentation was analyzed by agarose gel electrophoresis and flow cytometry respectively.Western blot was employed to analyze the expression of Bcl-2 and Bax.The in vivo efficacy of TFU was assessed in nude mice bearing tumours.The specimens were re-moved and the in situ cell apoptosis detection kit was employed for TUNEL staining.Results Growth of SGC-7901 and MKN-45 cells was remarkably suppressed by treatment with TFU in the presence of liver microsomal enzymes in vitro,suggesting that TFU might be converted to 5-FU by the enzymes.Similar treatment of TFU induced apoptosis of the cells,which was deduced from typical apoptotic features such as morphology,the formation of characteristic ladder pattern of DNA migration and the accumulation of sub-G1 phase.Furthermore,a significant inhibition of Bcl-2 expression and the up-regulation of Bax were observed after treatment with TFU in the presence of liver microsomal enzymes.Growth of human gastric carcinoma cells was significantly delayed by oral administration of TFU with low side effects.Apoptosis in xenografts was also observed by means of TUNEL staining method.Conclusion Treatment of TFU in the presence of liver microsomal enzymes could promote the inhibition of gastric carcinoma cell proliferation.TFU might sustain release of 5-FU mediated by liver microsomal enzymes.Low dose of 5-FU might trigger the carcinoma cells apoptosis via regulation of Bax and Bcl-2.

Objective To study the microbial strains,risk factors and resistance profiles of lower respiratory tract fungal infection in hospital of Zhoushan archipelago area.Methods A total of 204 patients who were hospitalized for lower respiratory tract infection were retrospectively analyzed from May 2008 to April 2011 in Zhoushan archipelago area,and collected 204 fungal strains isolated from confirmed lower respiratory tract fungal infection cases.Chi-square test and Logistic regression analysis were performed.Results Among the 204 fungal strains isolated from lower respiratory tract specimens,110 (53.8％) strains of Candidaalbicans,32 (15.7％) strains of Candida tropicalis,24 (11.8％) strains of Candida glabrata,12 (5.9％) strains of Candida krusei,14 (6.9％) strains of other Candida,and 12 (5.9％) strains of Aspergillus were detected.Logistic regression analysis showed that chronic obstructive pulmonary disease,bacterial pneumonia,long-term use of broadspectrum antibiotics and corticosteroids,endotracheal intubation or incision,old age,exposure in intensive care unit (ICU),and hospitalization ≥7 days were major risk factors (P=0.000,0.001,0.000,0.000,0.012,0.000,0.000,0.000).The resistance rates of isolated Candida against amphotericin B,5-flucytosine,voriconazole,itraconazole and fluconazole were 0,2.1％,4.2％,14.8％ and 22.9％,respectively.Conclusions Candida albicans is the major pathogen of lower respiratory tract fungal infection in hospital of Zhoushan archipelago area,and Candida is sensitive to amphotericin B,5-flucytosine and voriconazole.

Objective To explore the relationship between methylenetetrahydrofolate reductase (MTHFR) 677C ＞ T and unexplained recurrent pregnancy loss (URPL).Methods All patients were recruited from the outpatient department of Obstetrics/Gynaecology & Genetics of Hangzhou First People's Hospital from January 2013 to May 2014.A case-control study was performed.According to the stochastic indicator method,there were 125 subjects with a history of ≥2 times URPL as the case group,and 905 healthy parous women with no history of URPL as the control group.Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used to detect the distributions of the polymorphisms of MTHFR 677C ＞ T,and the results were validated using oligo-chip and direct sequencing.Results The allele and genotype frequencies of MTHFR were 60.0％ for C,40.0％ for T,32.0％ for CC,56.0％ for CT,and 12.0％ for TT in the case group and 67.4％ for C,32.6％ for T,41.4％ for CC,52.0％ for CT,and 6.6％ for TT in the control group,respectively.The prevalence of allele T was significantly higher in the case group than in the control group (OR =1.379,95％ CI =1.051-1.808,P =0.020),the frequency of genotype TT was also significantly higher in the case group than in the control group (OR =2.344,95％ CI =1.220-4.503,P =0.009).Conclusion The fertile women with MTHFR 677T allele and 677TT genotype may be susceptibility to URPL in a Chinese Han population from the Hangzhou area.

Objective To investigate the effect of hydrogen-rich water on cerebral edema and aquaporin 1 (AQP1) expression in rats with traumatic brain injury (TBI).Methods Ninety male Sprague-Dawley (SD) rats were randomly divided into sham operation group,TBI model group,hydrogen-rich water treatment group (H group),with 30 rats in each group.TBI model was reproduced by weight dropping method.The skulls of rats in sham operation group underwent only craniotomy without direct hit and with bone wax sealed suture.5 mL/kg of hydrogen-rich water injection was given intraperitoneally after model reproduction in H group,and equal amount of normal saline was given in sham and TBI groups,once a day for both groups for 5 days.Six rats from each group were sacrificed at 6,12,24,48 hours and 5 days after evaluating neurological severity scores (NSS).The cerebral cortex was harvested,and the pathological changes in morphology of brain tissue were observed with light microscope.The positive expression of AQP1 in cerebral cortex was observed with immunohistochemistry by light microscopy,the AQP1 mRNA expression in cerebral cortex was determined by real-time fluorescent quantization reverse transcription-polymerase chain reaction (RT-PCR),and the AQP1 protein expression in cerebral cortex was determined by Western Blot.Results ① All rats in sham operation group had a NSS of zero at each time point.NSS of TBI group was obviously raised with time prolongation,and peaked at 24 hours followed by a lower tendency,while the score in H group was significantly lower than that of TBI group,and the difference was the most obvious at 24 hours as compared with TBI group (9.83 ± 2.78 vs.13.50± 2.42,P ＜ 0.05).② It was shown by light microscope that in the TBI group there were pathological changes in cerebral cortex,including obvious irregular arrangement of nerve cells,cerebral edema,obvious bleeding,especially at 24 hours,then the cerebral edema became vanished gradually;and the positive expression of AQP1 in the pia mater at all the time points in the TBI group was significantly increased,and it was most obvious at 24 hours.Compared with TBI group,the pathological changes at time points of 12 hours to 5 days in H group was significantly lessened,and the positive expression of AQP1 in the cerebral pia mater was reduced obviously.③ Compared with sham operation group,the mRNA and protein expressions of AQP1 in cerebral cortex in TBI group were significantly elevated,peaked at 24 hours [AQP1 mRNA (2-△△Ct):7.50±0.26 vs.1,AQP1 protein (gray value):1.986±0.110 vs.0.336±0.034,both P ＜ 0.05],then they gradually declined.The mRNA and protein expressions of AQP1 in cerebral cortex were significantly decreased after hydrogen-rich water treatment [24-hour AQP1 mRNA (2-△△Ct):5.40±0.21 vs.7.50±0.26,24-hour AQP1 protein (gray value):1.246±0.137 vs.1.986±0.110,both P ＜ 0.05].Conclusions The up-regulation of AQP1 mRNA and protein in ratst cerebral cortex after TBI perhaps participates in edema formation which might be involved in the pathophysiology of cerebral edema in TBI.Early treatment with an intraperitoneally injection of hydrogen-rich water is capable of attenuating the extent of TBI-induced up-regulation of AQP1 mRNA and protein,alleviating cerebral edema,and achieving its protective effects.

Objective To investigate the effects of hydrogen rich water on the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and oxidative stress in rats with traumatic brain injury (TBI).Methods Ninety healthy male Sprague-Dawley (SD) rats were randomly divided into sham operation group, TBI group and hydrogen rich water treatment group (HW group), with 30 rats in each group.TBI model was reproduced by the modified Feeney weight dropping method.The skulls of rats in sham operation group underwent only craniotomy without direct hit.The rats in HW group received brain injury by hitting after craniotomy, followed by injection of hydrogen rich water (5 mL/kg) intraperitoneally once a day for 5 days after successful reproduction of the model.The rats in sham operation group and TBI group were given an equal amount of normal saline in same manner.Six rats from each group were sacrificed at 6, 12, 24, 48 hours and 5 days after evaluating neurological severity scores (NSS).The brain tissue in injured ipsilateral cortex was harvested.The activity of catalase (CAT), glutathione peroxidase (GSH-Px), and content of malondialdehyde (MDA) were determined by spectrophotometry.The expressions of mRNA and nucleoprotein of Nrf2 were determined by quantitative real-time polymerase chain reaction (RT-qPCR) and Western Blot.The pathological changes were observed with microscopy after hematoxylin and eosin (HE) staining.Results ① NSS score:compared with TBI group, NSS in HW group at 12, 24, 48 hours and 5 days were significantly decreased (12 hours: 9.83±2.32 vs.13.17±2.71, 24 hours: 9.83±2.79 vs.13.50±2.43, 48 hours: 7.50±2.07 vs.11.83±2.14, 5 days:5.50 ± 1.87 vs.10.50 ± 2.43, all P ＜ 0.05).② Compared with sham operation group, the activity of GSH-Px and CAT in TBI group were markedly declined after operation, while the MDA content was elevated significantly, especially at 24 hours [CAT (kU/g): 1.080±0.312 vs.3.571 ±0.758, GSH-Px (kU/g): 9.195±3.173 vs.32.385± 10.619, MDA (μmol/g): 12.282±2.896 vs.4.349± 1.511, all P ＜ 0.01].Compared with TBI group, the parameters in HW group were improved, and they were similar as sham operation group.③ RT-qPCR: no significant difference was found in the expression of Nrf2 mRNA at each time point in three groups.④ Western Blot: the expression of Nrf2 nucleoprotein (gray value) in TBI group was apparently higher than that in sham operation group, and peaked at 24 hours (0.703 ± 0.262 vs.0.238 ± 0.120, P ＜ 0.05), and the expression in HW group was obviously higher than that in TBI group, especially at 24 hours (1.110 ± 0.372 vs.0.703 ± 0.262, P ＜ 0.05).⑤ HE staining: the brain structure in sham operation group was found to be intact.However, there were different degrees of pathological changes at each time in TBI group, especially at 24 hours.The pathological damage of brain tissue in HW group was significantly milder.Conclusions Hydrogen rich water can up-regulate the expression of Nrf2, and reduce oxidative damage of traumatic brain injury in rats.Nrf2 can up-regulate the expression of its downstream antioxidant enzymes, which may be the mechanism of the upregulation expression of Nrf2 in the study.