[Objective]To study the difference in expression of RFC, GST-?, DHFRmRNA between human osteosarcoma U2-OS cell line and the MTX-resisitant variants U2-OS/R1-R3, and to investigate the significance in MTX resistance for human osteosarcoma. [Methods]Three resistant MTX human osteosarcoma cell lines were established by pulse exposure parental cell line(U2-OS) in gradually increased dose of MTX . The expression of RFC,GST-?,DHFRmRNA were assayed by real-time fluorescence quantitative polymerase chain reaction(FQ-PCR).[Results]Three MTX-resistant variants(U2-OS/R1-R3) were successfully established , the results of the FQ-PCR revealed that the MTX resistance was associated with the decreased expression of the RFC mRNA and increased expression of DHFR mRNA and GST-? mRNA.[Conclusion]The author investigated the MTX resistant mechanism of human osteosarcoma cell line at a gene level. The decreased expression of RFC mRNA and the increased expression of DHFR mRNA and GST-? mRNA participate in the MTX resistance in human osteosarcoma cell lines U-2 OS. This provides the evidence for exploring the MTX resistance mechanism in clinical osteosarcoma patients ,and helps to screen the patients who are insensitive to MTX chemotherapy.

Objective To investigate the effects of different doses of dexmedetomidine on the minimum alveolar concentration (MAC) of sevoflurane for sedation in patients.Methods ASA physical status Ⅰ or Ⅱ patients of both sexes,aged 18-64 yr,undergoing elective lower abdominal surgery performed under general anesthesia,were randomly divided into 4 groups:control group (group C) and different doses of dexmedetomidine groups (D1,D2 and D3 groups).In D1,D2 and D3 groups,the loading dose of dexmedetomidine 0.4,0.6 and 0.8 μg/ kg was intravenously infused over 15 min,respectively,adverse cardiovascular events were then recorded,followed by infusion at 0.4,0.6 and 0.8 μg· kg-1 · h-1 via a pump,respectively,while in group C,the equal volume of 0.9 ％ normal saline was given instead of dexmedetomidine.Sevoflurane administration was begun after completion of infusion of the loading dose.Up-and-down sequential allocation was used to determine the MAC.The initial end-tidal concentration of sevoflurane was set at 0.8％,0.7％,0.6％ and 0.5％ in C,D1,D2 and D3 groups,respectively,and maintained at this level for 15 min.Each time the concentration of sevoflurane increased/decreased in the next patient depending on whether or not the patients correctly followed the verbal command to open his eyes.The ratio between the two consecutive concentrations was 0.9.The middle point between the positive response and negative response served as a crossover pair.After at least 7 independent crossover pairs were observed in each group,the experiment was stopped.The MAC and 95 ％ confidence interval of sevoflurane were calculated.Results The incidence of adverse cardiovascular events was significantly higher in D3 group than in D1 and D2 groups (P ＜ 0.05).In C,D1,D2 and D3 groups,the MAC (95％ confidence interval) of sevoflurane was 0.68％ (0.64％-0.74％),0.50％ (0.47％-0.52％),0.36％ (0.32％-0.41％) and 0.28％(0.26％-0.31％),respectively.The MAC-awake of sevoflurane was significantly lower in D1-3 groups than in group C,in D2 and D3 groups than in group D1,and in D3 group than in group D2 (P ＜ 0.05).Conclusion Dexmedetomidine 0.6μg/kg can significantly decrease the MAC of sevoflurane for sedation,induces no side effects and is the optimum dose in patients.

Objective To evaluate the effects of different doses of dexmedetomidine on the median effective concentration (EC50) of propofol given by target-controlled infusion (TCI) at loss of consciousness (LOC).Methods Eighty ASA Ⅰ or Ⅱ patients of both sexes,aged 18-64 yr,with body mass index ≤25 kg/m2,scheduled for operations under general anesthesia,were randomly allocated to one of four groups(n=20 each): control group (group C) and dexmedetomidine 0.4 μg/kg group (group D1),dexmedetomidine 0.5 μg/kg group (group D2) and dexmedetomidine 0.6 μg/kg group (group D3).Dexmedetomidine 0.4,0.5 and 0.6 μg/kg were infused intravenously over 10 min in groups D1-3,while the equal volume of normal saline was given instead of dexmedetomidine in group C.Propofol was then given by TCI and the EC50 was determined by up-and-down sequential trial.The target plasma concentration was set at 2.0μg/ml in the first patient in each group.The ratio of the target plasma concentration between the two consecutive patients was 1.1.Loss of response to eyelash stimulation and verbal command (2 times) was considered to be signs of LOC.The EC50 and 95％ confidence interval (CI) of propofol causing LOC were calculated.Complications such as bradycardia,hypotension and respiratory depression were recorded.Results The EC50 (95％ CI) of propofol causing LOC was 2.59 (2.51-2.67),2.09 (2.02-2.16),1.82 (1.70-1.95) and 1.60 (1.49-1.72) μg/ml in groups C and D1.3 respectively.The EC50 of propofol causing LOC was significantly lower in groups D1-3 than in group C.Dexmedetomidine significantly decreased the EC50 of propofol required for causing LOC in a dose-dependent manner in groups D1-3 (P ＜ 0.05).The incidences of bradycardia and hypotension were significantly lower in groups D1.3 than in group C (P ＜ 0.05).Compared with group D1,the incidence of bradycardia was increased in groups D2,3 and the incidence of hypotension was increased in group D3 (P ＜ 0.05),There was no significant difference in the incidences of bradycardia and hypotension between groups D2 and D3 (P ＞ 0.05).No patients developed respiratory depression.Conclusion The optimum dose for dexmedetomidine infused intravenously when combined with propofol given by TCI is 0.4 μg/kg and it can decrease the EC50 of propofol administered by TCI at LOC with no adverse reactions.

Objective To investigate the effect of dexmedetomidine on the expression of hypoxia-inducible factor-1α (HIF-1α) during hypoxia/reoxygenation (H/R) in human renal tubular epithelial cells.Methods Human renal tubular epithelial cells (HK-2 cells) cultured in vitro were randomly divided into 4 groups (n =24 each) using a random number table:control group (group C),dexmedetomidine group (group DEX),H/R group and H/R+ dexmedetomidine group (group H/R + DEX).In group C,the cells were incubated for 28 h in an incubator filled with normoxia at 37 ℃.In group DEX,dexmedetomidine 0.1 nmol/L (final concentration) was added to the culture medium and the cells were incubated for 2 h,and then incubated for 28 h in an incubator filled with normoxia at 37 ℃.In group H/R,the cells were incubated in an anaerobic chamber for 24 h at 37 ℃,and then incubated for 4 h in an incubator filled with normoxia at 37 ℃.In group H/R + DEX,the cells were incubated for 2 h in the culture medium containing dexmedetomidine 0.1 nmol/L (final concentration),incubated in an anaerobic chamber for 24 h at 37 ℃,and then incubated for 4 h in an incubator filled with normoxia at 37 ℃.After treatment in each group,the cell viability was measured by MTT assay,cell apoptosis was measured using flow cytometry,the expression of HIF-1α mRNA was detected using RT-PCR,the expression of HIF-1α and activated caspase-3 protein was detected by Western blot,and the cell growth was observed.The apoptosis rate was calculated.Results Compared with group C,the cell viability was significantly decreased,the apoptosis rate was increased,and the expression of HIF-1α mRNA and protein and activated caspase-3 protein was up-regulated in H/.R and H/R + DEX groups,and no significant change was found in group DEX.Compared with group H/R,the cell viability was significantly increased,the apoptosis rate was decreased,the expression of HIF-1α mRNA and protein was up-regulated,the expression of activated caspase-3 protein was down-regulated,and the cell status was significantly improved in group H/R + DEX.Conclusion The mechanism by which dexmedetomidine attenuates H/ R-induced damage to human renal tubular epithelial cells may be related to up-regulated expression of HIF-1 α and inhibited cell apoptosis.

Objective To investigate the effect of STH-2 cardioplegic solution containing levosimendan on ischemia-reperfusion (I/R) injury in isolated rat hearts. Methods Thirty-two male Wistar rats weighing 250-300 gwere anesthetized with intraperitoneal 3% pentobarbital 30 mg/kg. The hearts were rapidly excised and perfused with oxygenated (95% O2-5% CO2) K-H solution for 30 min in a Langendorff apparatus and then divided into 4groups (n = 8 each) according to the composition of cardioplegic solution: group Ⅰ control (group C) was perfused with STH-2 cardioplegic solution; group Ⅱ , Ⅲ and Ⅳ were peffused with STH-2 cardioplegic solution containing levosimendan 0.03 μmol/L (L1), 0.3 μmol/L (L2) and levosimendan 0.3 μmol/L + glibenclamide 10 μmol/L (L2+ G) respectively. The isolated hearts were first perfused with different cardioplegic solutions for 2 h and then with K-H solution for 30 min. The coronary effluent was collected before ischemia (baseline) and at 10, 20 and 30 min of reperfusion for measurement of creatine kinase (CK) and lactate dehydrogenase (LDH)activities. Myocardial specimens were obtained from apex at 30 min of reperfusion for determination of myocardial ATP and MDA contents and SOD activity. Results Perfusion with STH-2 cardioplegic solution significantly increased CK and LDH activities and MDA content, and significantly decreased SOD activity. Levosinendan 0.03or 0.3 μmol/L significantly attenuated the cardioplegia-induced increase in LDH,CK and SOD activities and MDA content. The protective effects of levosimendan on myocardium against I/R injury were reversed by glibenclamide to some extent. Conclusion Levosimendan can protect myocardium from I/R injury in a dose-dependent manner by opening KATP channel.

Objective:To evaluate the role of nuclear factor erythroid 2-related factor/ heme oxygenase-1 (Nrf2/HO-1) signaling pathway in dexmedetomidine-induced reduction of oxygen-glucose deprivation and restoration (OGD/R) injury to microglia.Methods:BV-2 microglia were cultured in high-glucose DMEM culture medium supplemented with 10% fetal bovine serum in an normal culture incubator at 37 ℃ (5%CO 2-21%O 2-74 %N 2). The cells were seeded in 96-well plates at a density of 1.5×10 4 cells/ml (200 μl/well) or 6-well plates at a density of 2×10 5 cells/ml (2 ml/well) and divided into 5 groups ( n=30 each) using a random number table method: control group (group C), dexmedetomidine group (group D), group OGD/R, OGD/R+ dexmedetomidine group (group OGD/R+ D) and OGD/R+ dexmedetomidine+ ML385 group (group OGD/R+ D+ ML). The cells in group C were continuously cultured in a normal culture incubator for 26 h. In group D, dexmedetomidine at the final concentration of 10 μmol/L was added, cells were incubated for 2 h, and then were continuously incubated in a normal culture incubator for 26 h. In OGD/R, OGD/R+ D and OGD/R+ D+ ML groups, the culture medium was replaced with glucose-free DMEM culture medium, cells were cultured for 2 h in an incubator at 37 ℃ (5%CO 2-1%O 2-94 %N 2), the culture medium was replaced with high-glucose DMEM culture medium containing 10% fetal bovine serum and then the cells were cultured for 24 h in a normal incubator.Dexmedetomidine at the final concentration of 10 μmol/L was added at 2 h before OGD in OGD/R+ D and OGD/R+ D+ ML groups.Nrf-2 inhibitor ML385 at the final concentration of 4 μmol/L was added at 30 min before dexmedetomidine was added in group OGD/R+ D+ ML.Cells in 6 wells in each group were selected randomly for assessment of cell viability (by methyl thiazolyl tetrazolium assay) and apoptosis (using flow cytometry), and for determination of the concentrations of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-10 in the supernatant (using enzyme-linked immunosorbent assay), the expression of Nrf2 in nucleus, Nrf2 and HO-1(by Western blot ) and the expression of HO-1 mRNA (by real-time polymerase chain reaction). Results:Compared with group C, the cell viability was significantly decreased, cell apoptosis rate and concentrations of TNF-α, IL-6 and IL-10 in the supernatant were increased, and the expression of Nrf2 in nucleus, Nrf2, HO-1 and its mRNA was up-regulated in OGD/R and OGD/R+ D groups ( P<0.05), and no significant change was found in each parameter mentioned above in group D ( P>0.05). Compared with group OGD/R, the cell viability and IL-10 in the supernatant concentration were significantly increased, cell apoptosis rate and concentrations of TNF-α and IL-6 in the supernatant were decreased and the expression of Nrf2 in nucleus, Nrf2, HO-1 and its mRNA was up-regulated in group OGD/R+ D ( P<0.05), and no significant changes were found in the parameters mentioned above in group OGD/R+ D+ ML ( P>0.05). Compared with group OGD/R+ D, the cell viability and concentration of IL-10 in the supernatant were significantly decreased, cell apoptosis rate and concentrations of TNF-α and IL-6 in the supernatant were increased and the expression of Nrf2 in nucleus, Nrf2, HO-1 and its mRNA was down-regulated in group OGD/R+ D+ ML ( P<0.05). Conclusion:The mechanism by which dexmedetomidine alleviates OGD/R injury to microglia may be related to promoting the activation of Nrf2/HO-1 signaling pathway and inhibition of inflammatory responses.

ObjectiveTo investigate the diagnostic value of single balloon enteroscopy (SBE) for obscure gastrointestinal bleeding.MethodsA total of 78 SBE procedures was conducted on 72 patients with obscure gastrointestinal bleeding,with 40 via oral route and 38 via anal route.The procedure time,insertion depth and rate of positive finding were recorded.ResultsFor 40 SBE procedures performed via oral route,the mean procedure time was 60 minutes ( 15-110 minutes),and the mean insertion depth was 195 cm at the distal end of Trentz ligament (30-240 cm).For 38 SBE procedures performed via anus,the mean procedure time was 75 minuets (30-120 minutes),and the mean insertion depth was 160 cm at the proximal end of ileocecal valve (50-200 cm ).The whole diagnostic yield of obscure gastrointestinal bleeding was 62.5％.ConclusionSBE is a safe and useful tool for the diagnosis of obscure gastrointestinal bleeding.

Objective To compare the difference of kidney function evaluated by using 3 different estimated glomerular filtration rate(GFR) equations in populations .Methods Retrospectively analyzed 65 856 patients who measured serum creatinine and Cysta‐tin C at the same time ,and come from the outpatients or inpatients of the hospital .The estimated GFR (eGFR) were calculated through 3 equations ,then compared the eGFR in the population and among different groups according to different kidney functions , and then grouped the people enrolled in the study again according to the eGFR calculated by using the 3 different equations and compared the differences among groups .Results Compared with the eGFR calculated by using Creatinine equation ,the correlation coefficients of the eGFRs calculated by using the other two equations were 0 .81 and 0 .90 ,respectively ,both P<0 .05 ;The differ‐ence between the means of eGFR were 6 .19 and 1 .79 mL/(min × 1 .73 m2 ) respectively with obvious significance (P<0 .01) ,in consistency analysis .There were obvious overestimation of kidney function when using Creatinine equation to calculate eGFR .Con‐clusion There is consistence and obvious difference by using the 3 CKD‐EPI′s eGFR equations .Physicians should choose suitable equations to evaluate kidney function in different populations .

Objective To survey on the prevalence of schistosomiasis among floating population in Zhejiang province. Methods A survey on prevalence of schistosomiasis among floating population was conducted from September to November 2008, and the stratified cluster sampling method was adopted in the survey. Totally 129 villages of 19 counties or districts were selected as survey sites, and 100 samples of migrants aged 6 to 65 from schistosomiasis-endemic areas were taken in each selected village. All selected individuals were surveyed by questionnaire and underwent serum indirect hemagglutination (IHA) test. For individuals with positive serum IHA testing the fecal examination was carried out to detect the eggs by nylon sedimentation method. SPSS 13.0 software was used for data processing. Results The number of migrants in survey sites was 3 357 420, among whom 303 219 were from schistosomiasis-endemic areas (9.03%).The positive rate in serum IHA test was 2.06% (286/13 898), 276 IHA-positives individuals received fecal examination, and 7 cases were positive (2.52%). Based on above data it was estimated that there would be potentially about 33 500 serum IHA-positive cases and 845 egg-positive cases among floating population in Zhejiang province. Conclusion The risk of schistosomiasis transmission still exists in Zhejiang province due to the infected migrants from endemic areas, and a surveillance system and quick response are required for prevention of re-emergence of the disease.

Objective To determine the feasibility and safety of transprostatic utricle seminal vesiculoscopy in the treatment of hemospermia. Methods Totally 11 patients with hemospermia, mean age of (46.6 ± 3.5) years, ranging from 38 to 68 years, for 3 months admitted from September 2012 to August 2015 were enrolled, Their main manifestations were hemospermia. Painful ejaculation was observed in 7 patients, and perineal and testicular pain occured in 4 patients. They all underwent transprostatic utricle seminal vesiculoscopy, and then were followed up for 3 to 6 months. Results Ten patients were operated successfully, but 1 patient failed. The operation revealed that the causes of hemospermia were seminal vesiculitis in 8 cases, seminal vesiculitis accompanied with seminal calculi in 2 cases, and ejaculatory duct cyst in 1 case. Operation time was (29.2 ± 3.2) min ( ranging from 25 to 37 min) , and hospital stay was 2 d ( from 2 to 4 d). Hemospermia disappeared in 10 patients within 1 month of surgery, and hemospermia recurrence was observed in 1 patient within 6 months. The patient was treated with transprostatic utricle Holmium laser incision, then hemospermia was dispeared,Two cases of postoperative epididymitis were cured after one week of antibiotic treatment. Conclusions Seminal vesiculoscopy is a safe and effective to treat hemaospermia.