Objective:To investigate the impact of recombinant flagellin targeting TLR5 and NLRC4 simultaneously or respectively on innate immune cells in mice. Methods: Induction,expression,purification and identification of recombiant FliC,which were FliC(activating both TLR5 and NLRC4);FliCΔ90-97(unable to activate TLR5),FliC-L3A(unable to activate NLRC4),FliCΔ90-97:L3A(unable to activate both TLR5 and NLRC4). The mice were divided into five groups,namely group FliC,FliC-L3A,FliCΔ90-97,FliCΔ90-97:L3A and PBS,which were injected with 100μl PBS or 10μg recombinant flagellin intraperitoneally,three mice in each group. 12 h later,the mice were executed using dislocation of cervical vertebra and the splenic and peritoneal cells were isolated. The spleen was grinded into single-cell suspension. The proportion of neutrophils,NK cells,DCs and the expression level of CD80 and CD86 on DCs were evaluated with flow cytometry. Results:Group FliC,group FliC-L3A and group FliCΔ90-97 shared the similar proportion of neutrophils in peritoneal cavity ( P>0. 05 ) , and all of which were significantly higher than group PBS and group FliCΔ90-97 ( P<0. 01),and NK cells also showed the similar trend. Compared with group FliCΔ90-97 and FliCΔ90-97:L3A,the mean fluorescence intensities(MFIs) of CD80 and CD86 in group FliC and FliC-L3A increased significantly(P<0. 01). The proportion of Treg in spleen was highest among all groups. Conclusion:Activation of TLR5 and NLRC4 had similar chemotaxis of neutrophils and NK cells. The ex-pression of CD80 and CD86 on DCs were upregulated after stimulation by flagellin and TLR5-dependent. Activation of TLR5,but not NLRC4,increased the proportion of Treg in spleen.

Objective To evaluate the effect and adverse drug reaction of Boulardii yeast combined with triple and quadruple therapy on eradication helicobacter pylori (H.pylori).Methods240 cases of peptic ulcer patients of H.pylori positive were selected in our hospital from January 2014 to December 2015, according to different treatment were divided into the triple threapy group(n=60), the triple therapy union group (n=60), the quadruple therapy group (n=60) and the quadruple therapy union group (n=60).The triple threapy group were given clarithromycin and amoxicillin and pantoraazole;on the basis of this, the triple therapy union group were given Boulardiiyeast.the quadruple therapy group were given clarithromycin and amoxicillin and pantoraazole and CBS capsule, on the basis of this, the quadruple therapy union group were given Boulardiiyeast.The four groups were treated continuously for 14 days.14C-UBT, H.pylori eradication rate and adverse drug reaction in the four groups were evaluated five weeks after treatment.ResultsCompared with the triple threapy group and the quadruple therapy group, H.pylori eradication rate in the triple therapy union group(91.2%) and the quadruple therapy union group(94.7%) were improved obviously, and the adverse drug reactions (31.6%、29.8%) decreased significantly, the cumulative recurrence rate of H.pylori were significantly decreased, and the differences were statistically significant (P< 0.05).ConclusionBoulardii yeast combined with triple and quadruple therapy can obviously increase the H.pylori eradication rate, reduce the incidence of adverse drug reactions and the risk of recurrence.The reasonable treatment plan should be selected according to the actual situation.

The problems in teaching infectious disease were discussed .The measures should be taken to improve the teaching quality from the varies of aspects, such as the aim and the model of the teaching, the arrangement of curricula, the cultivation of the clinical ability, and the method of examination and the application of multi-media materials.

Objective To develop neural stem cells(NSCs) which can stably express exogenous brain-derived neurotrophic factor(BDNF) in vitro. Methods NSCs from the subependymal zone of embry-onic day 14.5(E14.5) rat brain were purified by limiting dilution assay and then infected with supernatant of recombinant retrovirus pLXSN-BDNF and retrovirus pLXSN. The original copy numbers of exogenous gene templates from three groups NSCs(pLXSN-BDNF viral infection group, pLXSN viral infection group, control group) were detected by fluorescent quantitative PCR(FQ-PCR). ELISA assay was used for determining the protein contents of BDNF of supernatant from three groups NSCs for six days continually after seeded in 24-well plates in the same cell density. Results NSCs were purified successfully by limiting dilution assay.The original copy numbers of exogenous BDNF gene templates from pLXSN-BDNF viral infection group by FQ-PCR were (19.57±0.65) × 10~3 copies/μl, higher than those of another two groups(P < 0.05). The protein contents of BDNF of supernatant from NSCs of pLXSN-BDNF viral infection group was highest among three groups and compared with another two groups had statistical significance (P <0.05) . Conclusion The purified NSCs can be transduced exogenous BDNF successfully with supematant of recombinant retrovir-us pLXSN-BDNF which provide experimental evidences and laying foundations for further research of retinal transplantation and quantization investigation of gene therapy for optic nerve injury.

Animals ; Dementia, Vascular ; metabolism ; physiopathology ; Extracellular Signal-Regulated MAP Kinases ; metabolism ; Hippocampus ; cytology ; metabolism ; physiopathology ; Male ; Mice ; Neurons ; metabolism ; physiology ; Random Allocation

Objective To evaluate neurodevelopmental outcome of very preterm(gestational age ＜ 32 weeks) and very low birth weight infant (VLBWI) (weight ＜ 1 500 g) and to examine the effectiveness of an early intervention program till 12 months corrected age.Methods Seventy followed-up very preterms and VLB-WI in Jinan Maternity and Childcare Hospital were enrolled in this study from January 2012 to and were divided into two groups by birth weight.All infants received 20 items of behavioral neurological assessment at 1 to 12 months corrected age and tested mental and psychomotor development with the use of CDCC at 6,12 months corrected age.The preterms who were abnomal in the 20 items of behavioral neurological assessment would receive early intervention (including kinesitherapy, physiotherapy, cereal circulation therapeutic equipment) by physiotherapists and their parents who received an intervention program training and were strongly encouraged to participate in the intervention sessions.The intervention method was adjusted according to the neurological assessment.The SPSS statistical software package for Windows, version 15.0, was used to run Fisher's exact test and t-test on the data presented,and P value of less than 0.05 was regarded as statistically significant.Results The average gestational age of infants was (30.4 ± 1.8) weeks,and average birth weight (1 463.7 ± 307.5) g.The incidence of extrauterine growth restriction was 57.1％ at first follow-up.The incidence of neurodevelpmental impairment NDI) and cerebral palsy tendency at 6 corrected months were 14.3％ ,8.6％ respectively.At 12 months corrected age,the incidence of NDI decreased to 2.9％ and cerebral palsy to 2.9％.There was significant difference in the incidence of NDI between 6 and 12 corrected months.There was no significant difference in the incidence of psychomotor developmental index ＜ 70, mental developmental index ＜ 70, NDI and cerebral palsy between the two groups.Conclusion The early intervention program can improve VLBWI neurodevelopmental outcomes at 12 months' corrected age and reduce the incidence of cerebral palsy.

Based on target cycling amplification and 3 ,4 ,9 ,10-perylenetetracarboxylic dianhydride derivative functionalized singal probe, an ultrasensitive electrochemiluminescence ( ECL) sensor was designed for the detection of lead ions. The hairpin substrate DNA was immobilized on the electrode through molecular self-assembly. In the presence of Pb2+and DNAzyme, the substrate was cleaved with single strand DNA fragments left on the electrode surface. Meanwhile, the target and DNAzyme was released for another cleaving circularly. As a result, the single strand DNA fragments hybridized with the assist hairpin probe H1, which leaded to the fabrication of H2 labeled with the 3 , 4 , 9 , 10-perylenetetracarboxylic dianhydride derivative functionalized hollow gold nanoparticles. With the increasing concentration of Pb2+, much more signal probe was been captured and the ECL signal of the biosensor in peroxydisulfate ( S2 O2-8 ) solution would increase. An ECL assay demonstrates that the sensor has a good linear response to Pb2+ concentration in the range of 1í10-12 mol/L-1í10-6 mol/L, with a detection limit of 1í10-12 mol/L. The fabricated sensor shows good selectivity toward Pb2+against other common metal ions.

Objective To observe the clinical efficacy of Tanreqing Injection combined with levofloxacin injection in treatment of acute episode of chronic bronchitis and its effect on bacterial clearance rate. Methods Totally 66 patients were randomly divided into treatment group and control group (33 cases for each). Both groups were given expectorant and antispasmodic treatment. The treatment group was given Tanreqing Injection combined with levofloxacin injection, while the control group was given levofloxacin injection only. The treatment course was 12 days. The clinical symptoms, signs, and sputum culture before and after treatment were observed. The clinical efficacy and the bacterial clearance rate of the two groups were compared. Results The markedly effective cases and the effective cases in the treatment group were 15 and 29 respectively, and those in the control group were 7 and 21 respectively, with statistical significance (P<0.05). The bacterial clearance rate was 89.5% (17/19) in the treatment group, while 55.6%(10/18) in the control group, with significant difference (P<0.05). Conclusion Acute episode of chronic bronchitis treated by Tanreqing Injection combined with levofloxacin injectcion is more effective than levofloxacin injection only.

Objective To investigate the effect of different doses of ketamine on inflammatory cytokines in rabbits with severe burn at early stage and preliminarily approach its regulatory action on early stage of inflammatory reaction due to stress of trauma.Methods Forty healthy male New Zealand rabbits were randomly divided into four groups in accord with the random number table method: normal control group, scald model group, ketamine analgesia group and ketamine anesthesia group. Before scald, pentobarbital sodium was used for anesthesia, afterwards catheters were inserted into internal jugular vein and internal carotid artery respectively ready for use, and 24 hours later, Ⅲ degree scald at the animal back and buttocks occupying 30% total body surface area (TBSA) was performed as the scald model for all the rabbits except those in normal control group. In ketamine analgesia group, after scald for 0.5 hour, 0.5 mg/kg ketamine intravenous injection was given to the rabbits as the loading dosage and then persistent intravenous pump infusion of 9μg·kg-1·min-1 ketamine was applied for all together 24 hours. In ketamine anesthesia group, after scald for 0.5 hour, 1.5 mg/kg ketamine intravenous injection was given to the rabbits, and then persistent intravenous pump infusion of 45μg·kg-1·min-1 ketamine was applied for 4 hours to maintain systemic anesthesia. In normal control and scald model groups, only intravenous infusion of equal amount of normal saline was given to the rabbits. The amount of intravenous transfusion in each group and the total dosages of ketamine used in ketamine analgesia group and ketamine anesthesia group were recorded. Before scald and 0.5, 6, 12, 24 hours after scald, arterial blood gas analyses were made, and the levels of serum interleukins (IL-1, IL-6) and tumor necrosis factor-α (TNF-α) were determined.Results Although the indexes of blood gas analysis were changed in the four groups, they were all in the normal range, showing that the respiratory function was in the normal range and indirectly reflecting that the circulatory function was also in the normal range, thus the effects on cytokines by factors of respiratory and circulatory functions were ruled out. The levels of IL-1, IL-6 and TNF-α before scald showed no statistically significant differencesamong the four groups (allP > 0.05). From 0.5 hour after scald, the levels of IL-1, IL-6 and TNF-α were markedly higher in model group than those of normal control group [IL-1 (ng/L): 30.27±0.93 vs. 13.79±1.11, IL-6 (ng/L): 47.22±1.49 vs. 46.31±4.12, TNF-α (ng/L): 243.39±20.85 vs. 190.95±14.97, allP < 0.05], and the situation continued until 24 hours after scald; the levels of IL-1, IL-6 and TNF-α from 6 hours after scald were significantly decreased in ketamine analgesia and ketamine anesthesia groups compared with those in the model group, and from 12 hours after scald, the degrees of descent in levels of the above indexes in ketamine analgesia group were more obvious than those in ketamine anesthesia group [IL-1 (ng/L): 19.28±2.51 vs. 40.12±10.31, IL-6 (ng/L): 52.10±4.23 vs. 72.20±10.11, TNF-α (ng/L): 246.03±20.74 vs. 313.71±27.34, allP < 0.05].Conclusion The low-dose ketamine analgesia and ketamine anesthesia have certain degree of inhibitory effect on the expression and release of inflammatory cytokines at the early stage in rabbits with severe burn, the effect of long-term low-dose ketamine analgesia being more significant.

Objective:To explore the impact of TLR5 and NLRC4 activation on the proliferation of different breast cancer cell lines,MCF-7 and MDA-MB-23 i.Methods:Induction,expression,purification and identification of recombiant flagellin,including FliC (activating both TLR5 and NLRC4),FliC△90-97 (unable to activate TLR5),FliC-L3A (unable to activate NLRC4),FliC△90-97:L3A (unable to activate both TLR5 and NLRC4).Using different concentration of recombinant flagellin to stimulate MCF-7 and MDA-MB-231 cell lines,72 h later,the proliferation of tumor cells were detected with CCK8.We also used soft AGAR forming experiments to detect the inhibition ratio of recombinant flagellin on breast cancer cell lines.Briefly,1 000 cells were plated in the 6-well plate,then stimulated with 1 μg/ml recombinant flagellin,14 days later,the number of cloning were counted after crystal violet staining.Results:After stimulation with four recombinant flagellins at the concentration of 0.1 μ,g/ml,the inhibition ratio on MCF-7 reached 30％,and FliC△90-97 were dose-dependent on the inhibition of MCF-7 proliferation.At the concentration of 1 μg/ml,FliC-L3A which only activated TLR5 showed stronger inhibition ratio than FliC.FliC△90-97:L3A which did not activate both TLR5 and NLRC4 also inhibited the proliferation of MCF-7.After adding transfection reagent,four recombinant flagellins showed inhibition effect on MDA-MB-231.Conclusion:Flagellin can inhibit the proliferation of MCF-7 and MDA-MB-231,and the mechanism of inhibition on the proliferation were not TLR5 and NLRC4 pathway dependent.There might exist new mechanisms to explain this phenomenon.