Objective To investigate the effects and probable mechanism ofShenfu injection on the oxidaitve damage of H2O2-induced PC12 cells.Methods PC12 cells were cultured and exposed to 100μmol/L H2O2 for 1 h to establish the oxidative damage model. The protective effect ofShenfu injection was observed by the cell survival rate measured by colorimetric MTT assay, and the leakage rate of lactic dehydrogense (LDH). Western blot methods were used to detect the expression of NF-κB signaling pathway.Results Compared with the model group,Shenfu injection at 5, 10, 20 ml/L could improve the PC12 cells survival rate (83.11% ± 2.59 %, 87.99% ± 0.59%, 85.26% ± 1.07%vs. 73.82% ± 1.82%;P<0.01 orP<0.05), decrease the LDH leakage rate (32.75% ± 4.10%, 28.52% ± 1.14%, 35.79% ± 1.62%vs. 64.34% ± 3.18%;P<0.01 or P<0.05). Western blot results showed thatShenfu injection could protect the PC12 cells from oxidaitve damage by suppressing the p-p65/p65 (1.30 ± 0.10, 1.17 ± 0.06, 1.37 ± 0.15 vs. 1.70 ± 0.10;P<0.01 orP<0.05), p-IκBα/IκBα (1.07 ± 0.12, 1.00 ± 0.10, 1.03 ± 0.06 vs. 1.17 ± 0.06; P<0.01 orP<0.05).ConclusionShenfu injection has a obvious antioxidant effect on PC12 cells in vitro.

Objective To investigate the mechanisms underlying the effect of 5-aminolevulinic acid photodynamic therapy (ALA-PDT) on cutaneous squamous cell carcinoma (SCC) in mice.Methods A model of cutaneous SCC was established in 21 SKH-1 hairless mice,which were treated with topical ALA 8％ cream followed by single irradiation with He-Ne laser at a total dose of 30 J/cm2 (ALA-PDT).Three mice were sacrificed before and at 1,3,6,12,24 hours and 7 days after the irradiation,separately,and SCC tissue was taken from the mice.Transmission electron microscopy (TEM) and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) were performed to determine the pattern of tumor cell death(necrosis,apoptosis and autophagy) during 1-24 hours after ALA-PDT,and immunohistochemical techniques were used to estimate the expressions of LC3B and CD34 on SCC cells,as well as the quantity of CD1a+ cells,CD4+ T and CD8+ T lymphocytes in SCC tissue 7 days after the irradiation.Statistical analysis was done by two-sample t test using SPSS 17.0 software.Results TEM showed gradual necrosis and apoptosis (especially necrosis) of tumor cells and formation of autophagosomes in macrophages within 24 hours after ALA-PDT.The number of apoptotic cells per high power field (× 400) in SCC tissue significantly increased at 24 hours compared with that before ALA-PDT (7.30 ± 2.18 vs.2.00 ± 0.69,P ＜ 0.05).As immunohistochemistry revealed,there was a significant decrease in the number of CD34+ cells (1.33 ± 0.58 vs.19.00 ± 2.66,P＜ 0.01),but a marked increase in that of CD1a+ ce1ls (23.01 ± 2.04 vs.10.33 ± 1.88,P＜ 0.05),CD4+ T cells (28.67 ± 1.76 vs.12.40 ± 2.27,P＜ 0.05),CD8+ T cells (25.79 ± 2.37 vs.11.67 ± 1.45,P ＜ 0.05) and LC3B+ interstitial cells (30.6 ± 3.21 vs.21.44 ± 4.3,P ＜ 0.05) per high power field (× 400) in SCC tissue on day 7 compared with that before ALA-PDT.Conclusions ALA-PDT may directly kill SCC cells by inducing cell necrosis and apoptosis rather than autophagy.Additionally,ALA-PDT can injure microvascular endothelial cells and cause the aggregation of dendritic cells,CD4+ T cells and CD8+ T cells in SCC tissue.

Objective To investigate the influence of low-dose ketamine and dexmedetomidine on cardiovascular response during. sedative amnesia fiberoptic nasotracheal intubation. Methods Ninety ASA Ⅰ or Ⅱ patients scheduled to recerve general anesthesia were evenly random-ized to dexmedetomidine and ketamine (group DK),dexmedetomidine and propofol (group DP)and dexmedetomidine and remifentanil (group DR).Ten minutes before intubation,the patients in group DK received intravenously dexmedetomidine 1.0 μg/kg plus ketamine 0.5 mg·kg-1 ·h-1 ;those in group DP received intravenously dexmedetomidine 1.0 μg/kg plus propofol 2.0 mg · kg-1 · h-1 ;those in group DR received intravenously dexmedetomidine 1.0 μg/kg plus remifentanil 5.0μg·kg-1 ·h-1 .Nasotracheal intubation was performed with fiberoptic bronchoscopy after dexemeto-midine injection and complete topical anesthesia.HR,MAP,SpO 2 and Ramsay sedation score were re-corded before anesthesia (T0 ,baseline),before intubation (T1 ),immediately intubated (T2 )and five minutes after intubation (T3 ).Side effects such as restlessness,bucking,respiratory depression and cardiovascular event during intubation and awareness of intubation were also recorded.Results All pa-tients in three groups were performed successfully.HR and MAP were significantly decreased in groups DP and DR at T1 (P <0.05),SpO 2 was significantly decreased in group DP at T1 (P <0.05);MAP in group DR were higher than those in group DP,HR in groups DP and DR were significantly increased than those in group DK at T3 (P < 0.05 );Ramsay score were significantly decreased in groups DP and DR at T2 ,significantly lower in group DR at T3 than those in groups DK and DP (P<0.05).The incidences of bradycardia and respriatory depression were significantly higher in group DP than those in group DK,and bucking,restlessness,tachycardia incidence rate in group DR were significantly higher than those in groups DK and DP (P <0.05).Conclusion Dexmedetomidine com-bined with low dose ketamine together with topical anesthesia is an ideal method for sedative amnesia fiberoptic nasotracheal intubation with slighter cardiovascular response and less side effects.

Objective To explore the effects of aminolevulinic acid-based photodynamic therapy (ALA-PDT) on the proliferation and apoptosis of HaCaT cells stably expressing human papillomavirus type 16 E7 protein (HaCaT/HPVl6 E7 cells).Methods Cultured HaCaT/HPV16 E7 cells were divided into several groups: blank control group receiving no treatment, ALA group treated with ALA alone, irradiation group irradiated with 630-nm red laser (30 mW/cm2,12 J/cm2), ALA-PDT groups pretreated with ALA for 5 hours followed by 630-nm red laser radiation at 4, 8, 12 J/cm2 respectively.CCK8 assay was performed to determine the survival rate of cells at 24 hours after PDT, and flow cytometetry and confocal microscopy were conducted to detect cell apoptosis and observe cell morphology respectively at 3 hours.Results At 24 hours, the survival rate of cells was 68.98％ ± 1.03％, 46.03％ ± 2.96％ and 23.57％ ± 3.83％ in the 4-,8-and 12-J/cm2 ALA-PDT groups respectively, significantly lower than that in the blank control group, ALA group and irradiation group (99.15％ ± 0.64％, 98.13％ ± 0.83％ and 96.85％ ± 1.37％ respectively, all P ＜ 0.05).With the increase in radiation dose, cell apoptosis was accelerated with obvious morphological changes and shrinkage of cells in the ALA-PDT groups.Conclusion ALA-PDT can inhibit the proliferation, and promote the apoptosis of HPV-infected HaCaT cells in a dose-dependent manner within a certain range of radiation dose.

Aim To investigate the effect of indirubin derivative PHⅡ-7 and TRAIL on proliferation in breast cancer cell MCF-7 and its MDR counterpart MCF-7/ADR and the mechanism.Methods Growth inhibition rate was examined respectively by MTT assay under treatment with TRAIL or PHⅡ-7 or in combination. Cell apoptosis and ROS production were examined by flow cytometry.The change of TRAIL receptors(DR4/DR5 )in mRNA was analysed by realtime PCR.Re-sults IC50 of PHⅡ-7 on MCF-7 and MCF-7/ADR was (4.49 ±1.55 ),(3.44 ±0.90 )μmol · L-1 respec-tively;MDA-MB-231 was TRAIL sensitive cell line, and apparently TRAIL induced apoptosis in MDA-MB-23 1 .Low concentration of PHⅡ-7 in combination with TRAIL could augment TRAIL-induced cytotoxic effect including apoptosis while TRAIL or PHⅡ-7 treatment alone had limited cytotoxity to those cells.Besides, PHⅡ-7 at this concentration had little toxicity to hu-man peripheral blood mononuclear cells even if in com-bination with TRAIL.PHⅡ-7 generated ROS produc-tion inside MCF-7 and MCF-7/ADR cells and up-regu-lated DR4/DR5 expression concentration dependently. Once upon ROS scavenger NAC involved,the effect of TRAIL receptors up-regualtion by expression was abro-gated.Conclusions PHⅡ-7 at low concentration could improve the sensitivities of breast cancer cell MCF-7 and MCF-7/ADR to TRAIL,the mechanism of which may be the ability of ROS production by PHⅡ-7 help up-regulated TRAIL receptor DR4,DR5 .Our re-search set a solid foundation for PHⅡ-7 in combination with TRAIL in future clinical application.

Objective To investigate the molecular types of carbapenem-non-susceptible Esche-richia coli ( E.coli) isolates and their mechanism of carbapenem resistance .Methods Twenty-two carbap-enem-non-susceptible E.coli strains were isolated from 3 hospitals in Hangzhou from 2007 to 2011.The mini-mum inhibitory concentrations ( MICs) of antimicrobials to those isolates were determined by agar dilution method and E-test.The molecular mechanisms of carbapenem resistance of E.coli isolates were analyzed by conjugation experiment,PCR and DNA sequencing.Pulsed-field gel electrophoresis (PFGE),multilocus se-quence typing ( MLST ) , and phylogenetic typing were performed to analyze the molecular epidemiology of those isolates.Results The MICs of imipenem and meropenem to 22 E.coli isolates were ranged from 1 μg/ml to 16 μg/ml,and the MICs of ertapenem were 2 μg/ml to 64 μg/ml.All E.coli isolates produced the KPC-2 carbapenemase and various β-lactamases , and some of them also produced plasmid-mediated AmpC enzymes.Carbapenem resistance was transferred by conjugation and transformation from 22 E.coli iso-lates to E.coli EC600 strains.The E.coli transconjugants or transformants acquired the blaKPC-2 gene and showed similar antibiotic susceptibility patterns in comparison with donor strains .Only a few isolates were in-distinguishable or closely related as indicated by PFGE .Four sequence types including ST131 (9 isolates), ST648 (5 isolates),ST38 (2 isolates) and ST405 (2 isolates) were identified by MLST.Phylogenetic analy-sis indicated that 9 ST131 isolates belonged to phylogenetic group B 2 and the other isolates belonged to group D (11 isolates),group B1 (1 isolate) and group A (1 isolate),respectively.Conclusion The sequence type of prevalent E.coli isolates producing KPC-2 from Hangzhou was ST131,which is an international epi-demic,multidrug-resistant clone,followed by ST648.

OBJECTIVE:To provide reference for variety breeding,high yield and high quality cultivation of Momordica cochi-nchinensis. METHODS:Based on Chinese Pharmacopoeia (2015 edition,Vol Ⅰ),HPLC for determining the content of gyp-sogenin 3-O-β-D-glucuronic acid methyl ester in M. cochinchinensis was optimized,the contents of gypsogenin 3-O-β-D-glucuronic acid methyl ester in M. cochinchinensis from 14 production places of 8 provinces were compared,and cluster analysis was conduct-ed. Correlation of longitude,latitude and altitude with content of gypsogenin 3-O-β-D-glucuronic acid methyl ester in M. cochinchi-nensis was analyzed by SPSS17.0 software. RESULTS:Gypsogenin 3-O-β-D-glucuronic acid methyl ester in M. cochinchinensis showed good linear relationship in 0.05-0.5 mg/mL,regression equation was Y=4361.95X+67.3808(R2=0.9997);the limits of quantification and detection were 15.62 ng and 4.67 ng,respectively. Average recovery was 99.76%(RSD=1.36%,n=9). The content of gypsogenin 3-O-β-D-glucuronic acid methyl ester in M. cochinchinensis from 14 production places of 8 provinces had ex-tremely significant differences(P<0.01). Clustering analysis results indicated that M. cochinchinensis from 14 production places of 8 provinces were divided into Ⅰ,Ⅱ,Ⅲ,Ⅳ groups. Contents of gypsogenin 3-O-β-D-glucuronic acid methyl ester in M. cochi-nchinensis of Ⅲ,Ⅳ groups were relatively high,including Guangxi Pingnan,Guizhou Jiangkou,Guizhou Dejiang,Fujian Jian-yang and Hunan Huitong,in which,Guangxi Pingnan,Guizhou Jiangkou,Guizhou Dejiang and Hunan Huitong had high altitude and low latitude,Fujian Jianyang had high altitude. The content of gypsogenin 3-O-β-D-glucuronic acid methyl ester in M. cochi-nchinensis was positively correlated with altitude,while negatively correlated with longitude and latitude. CONCLUSIONS:HPLC is simple,accurate and reproducible for determining the content of gypsogenin 3-O-β-D-glucuronic acid methyl ester in M. cochi-nchinensis,and the determined contents of gypsogenin 3-O-β-D-glucuronic acid methyl ester in M. cochinchinensis from 14 produc-tion places of 8 provinces have reached the standard of Chinese Pharmacopoeia (2015 edition,Vol Ⅰ). Cultivation in Guangxi Pingnan,Guizhou Jiangkou,Guizhou Dejiang,Hunan Huitong and other areas with high altitude and low latitude helps to improve the content accumulation of gypsogenin 3-O-β-D-glucuronic acid methyl ester in M. cochinchinensis.

Objective To detect and explore the expression and clinical significance of dual specificity tyrosine phosphorylation regulated kinase1b (Dyrk1b) in the specimens and cells of cervical lesions. Methods (1)All the data were collected from 75 patients with cervical cancer and 52 cases with squamous intraepithelial lesion(SIL)admitted in the First Affiliated Hospital of Dalian Medical College during Jan. 2011 to Dec. 2013 and confirmed by pathological examination, included 60 cases of stageⅠand 15 cases of stageⅡ, 12 cases with low-grade squamous intraepithelial lesion(LSIL)and 40 cases with high-grade squamous intraepithelial lesion(HSIL). While, 28 cases with chronic cervicitis were chosen as the control group. The protein expression of Dyrk1b was detected by immunohistochemistry among the four groups.(2)The expression of Dyrk1b in HeLa and SiHa cells were detected by western blot method and the expression of Dyrk1b protein were also detected after treatment of AZ191 (5, 10 μmol/L) for 48 hours in HeLa and SiHa cells.(3)The cellular survival and proliferation of HeLa and SiHa cells treated by different concentrations of AZ191(2.5, 5, 10, 25, 50, 100 μmol/L)for 48 hours were detected by methyl thiazolyl tetrazolium (MTT) assay.(4)The rate of apoptosis of HeLa and SiHa cells was detected by flowcytometry after treatment of AZ191 (5, 10μmol/L) for 48 hours. Results (1)The positive rates of Dyrk1b protein in chronic cervicitis, LSIL, HSIL and cervical squamous cancer by immunohistochemistry were 11%(3/28), 1/12, 42%(17/40)and 71%(53/75), respectively. The expression of Dyrk1b in cervical squamous cancer and HISL were higher than those in LSIL and chronic cervicitis (P<0.01), there were significant difference between cervical squamous cancer and HSIL, or between HSIL and LSIL(all P<0.05), while there were not significant difference between LSIL and chronic cervicitis(P>0.05). Expression of Dyrk1b was correlated with stromal invasion depth of cervical cancer (P<0.05), but not with age, clinical stage, lymph node metastasis, and serum squamous cell carcinom antigen(SCC-Ag)levels (all P>0.05). (2) Dyrk1b protein was expressed in different levels in HeLa and SiHa cells, and the expression of Dyrk1b was decreased gradually as the increased of the concentration of AZ191 in both HeLa and SiHa cells by treatment of AZ191 for 48 hours. (3) Different concentration of AZ191 treated on cervical cancer cells could inhibit the cellular proliferation and induce cell apoptosis in a concentration-dependent manner(P<0.01), concomitant to the decreased cell survival rate. The apoptosis rate of HeLa and SiHa were increased significantly after 10μmol/L AZ191-treatment for 48 hours, but no any difference induced by 5 μmol/L AZ191-treatment compared to control group. Also,there was no any difference between Hela and SiHa cells in either inhibitory effect or apoptosis rate induced by AZ191. Conclusions Dyrk1b is over-expressed in either specimens or cells of cervical cancer. The expression of Dyrk1b protein in cervical lesions is increased as the progression of disease. Dyrk1b inhibitor AZ191 could inhibit cellular proliferation and induce apoptosis in a concentration-dependent manner in cervical cancer cells.

Objective To evaluate the cognitive level of proton pump inhibitors for the clinicians who prescribe these drugs often. Methods 10 cognitive latitudes for proton pump inhibitors were selected, which were receptor site, indication, indication of prophylaxis during perioperation, patients in contraindi-cated, high risk factors of stress ulcer, standard dose, daily dosing frequency, solvent selection for intrave-nous drip and main adverse reactions. Every cognitive latitude was assigned to points and one evaluation form was assigned 120 points. 175 clinicians participated the assessment voluntarily. Results Overall,the total average score is 58. 89. Each average score of 10 cognitive level is 10. 63, 1. 07, 6. 99, 5. 23, 3. 81, 11. 86, 11. 66, 3. 81, 7. 49 and 3. 81 respectively. The error rate is 10. 86％, 84. 57％, 45. 14％, 80. 00％, 59. 43％, 4. 00％, 2. 86％,80. 57％, 21. 71％ and 28. 00％ accordingly. Conclusions Over-all, the clinicians'cognitive level is low, while the senior doctors'are high in surgical prophylaxis;the physi-cians'are high in compatibility than surgeons ;the clinicians who used proton pump inhibitors ( PPI) more frequently are high in solvent selection for intravenous drip.

Objective:To explore the feasibility of dual-energy CT (DECT) in the evaluation of liver iodine concentration in patients on long-term oral amiodarone treatment (≥12 months).Methods:Eighteen subjects undergoing abdominal dual-energy CT who met the criterion in January 2017 were collected as a control group. Twenty-seven patients who received oral amiodarone treatment for more than 12 months from January 2017 to May 2019 were enrolled as an experimental group. The difference of CT value and iodine concentration of liver, pancreas and spleen in 140 kV, 100 kV and VNC images between experimental and control groups were measured and analyzed. The correlation between liver CT value and liver iodine concentration was analyzed by Spearman correlation.Results:The concentrations of iodine in the liver, pancreas and spleen of the experimental group were 0.2 (0.2, 0.4), 0.1 (0.1, 0.2) and 0.2 (0.1, 0.2) mg/ml, and those in the control group were 0.2 (0.1， 0.2), 0.1 (0.1, 0.2) and 0.1 (0.1, 0.2) mg/ml. The difference in the concentration of iodine of the liver was statistically significant ( Z=-3.354, P<0.05), however there was no significant difference in the concentration of pancreas and spleen between the two groups ( Z=-0.179 and -1.590, P>0.05). The CT values of 100 kV, 140 kV, VNC images in the experimental group [(74±18)， (70±10) and (71±5) HU] were higher than those in the control group [(60±6)， (59±6) and (62±6) HU], and the differences were statistically significant ( t=3.310, 4.205 and 5.241, P<0.05). There was a positive correlation between the CT value of 140 kV image and iodine concentration ( r=0.410, P<0.05). In the experimental group, the time difference of taking amiodarone was statistically significant ( P<0.05). Conclusions:DECT can be used to quantitatively measure the liver iodine concentration of patients with long-term oral amiodarone, and provides some biological indicators for the assessment of amiodarone induced liver injury.