Objective To evaluate the effects of propofol on the intracellular calcium ion concentration ([Ca2+ ]i) and nuclear factor kappa B (NF-κB) activity in hippocampal neurons of fetal rats in vitro .Methods Ten pregnant Sprague-Dawley rats at 16-18 days of gestation ,were sacrificed and the fetal rats were taken out from the abdominal cavity .The hippocampal neurons of the fetal rats were isolated and seeded in culture plates .After being cultured for 9 days ,the neurons were divided into 7 groups ( n=12 each ) using a random number table :control group (C group) ,intralipid group (I group) and propofol 0.1 ,1 ,10 ,100 ,1 000 μmol/L groups (P1-5 groups) .In group I ,10% intralipid was added to the culture media until the final concentration reached 100μmol/L .In P1-5 groups ,propofol was added to the culture media until the final concentration reached 0.1 ,1 ,10 , 100 and 1 000μmol/L ,respectively .The cells were then incubated for 3 h .The [Ca2+ ]i and cellular morphology of hippocampal neurons were examined by laser scanning confocal microscopy before incubation with propofol and within 10 min after the end of incubation with propofol .The expression of NF-κB protein in the nucleus was detected at 7 days after the end of incubation with propofol by Western blot analysis to reflect NF-κB activity . Results Propofol increased [Ca2+ ]i in P2-4 groups ,while decreased [Ca2+ ]i in group P5 ( P<0.05 ) .Compared with group C ,the activity of NF-κB was significantly decreased in P1-5 groups ( P<0.05 ) ,while no significant change was found in C and I groups ( P>0.05 ) .The structure of hippocampal neurons was normal in C ,I and P1 groups .The branchings of axons and dendrites in hippocampal neurons were significantly decreased in P 2-4 groups , while the structure of hippocampal neurons became fuzzy , the cell membrane was destroyed and the axons and dendrites were not seen in group P5 .Conclusion Propofol can produce neurotoxic effects on hippocampal neurons of fetal rats by changing the [Ca2+ ]i and promoting NF-κB activation in vitro .

Objective To evaluate the effects of dexmedetomidine on the expression of phosphor-cAMP response element binding protein (p-CREB) in isoloated hippocampal neurons of fetal rats.Methods SpragueDawley rats on 16-18 days of gestation were sacrificed and the fetal rats were obtained.The hippocampi of fetal rats were isolated and hippocampal neurons were seeded in culture medium for 8 days.The cells were then divided into 4 groups (n =12 each) using a random number table:control group (group C),dexmedetomidine 0.001 μmol/L group (group D1),dexmedetomidine 0.010 μmol/L group (group D2),and dexmedetomidine 0.100μmol/L group (group D3).In D1.3 groups,dexmedetomidine with the final concentrations of 0.001 μmol/L,0.010 μmol/L,and 0.100 μmol/L was added to the culture medium,respectively,and then the cells were incubated for 3.5 h.The apoptosis in hippocampal neurons was detected by flow cytometry.The expression of p-CREB in hippocampal neurons was determined by RT-PCR and Western blot.Results Compared with group C,apoptosis rate was significantly decreased and the expression of p-CREB was up-regulated in D1.3 groups.Conclusion Dexmedetomidine inhibits apoptosis in isolated hippocampal neurons of fetal rats by up-regulating the expression of p-CREB.

Objective To evaluate the effects of propofol on apoptosis in the hippocampal neurons of fetal rats in vitro.Methods The isolated hippocampal neurons were seeded into 96-well plates or 24-well plates at a density of 5 × 104 cells/ml.The cells were randomly divided into 5 groups (n =18 each) using a random number table:control group (group C),in tralipid group (group Ⅰ) and propofol 1,10,100 μmol/L groups (P1-3 groups).In group Ⅰ,10％ intralipid was added to the culture media until the final concentration reached 100 μmol/L.In groups P1-3,propofol was added to the culture media until the final concentration reached 1,10 and 100 μmol/L,respectively,and the cells were then incubated for 3 h.The cell apoptosis was assessed by flow cytometry.The expression of Bcl-2 mRNA and caspase-3 mRNA was detected by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR).The expression of Bcl-2 and actived-caspase-3 protein was determined by Western blot analysis.The apoptosis rate was calculated.Results Compared with group C,the apoptosis rate was significantly increased,the expression of Bcl-2 mRNA and protein was down-regulated,and the expression of caspase-3 mRNA and actived-caspase-3 protein was up-regulated in P1-3 groups (P ＜ 0.05).There was no significant difference in the parameters mentioned above between group Ⅰ and group C (P ＞ 0.05).Conclusion Propofol induces apoptosis in isolated hippocampal neurons by inhibiting Bcl-2 expression and enhancing caspase-3 activity in fetal rats.

Objective To evaluate the effect of dexmedetomidine on the expression of nerve growth factor (NGF) in isolated hippocampal neurons of fetal rats incubated with propofol.Methods Hippocampal neurons derived from the fetal rats of pregnant Sprague-Dawley rats at 5-13 days of gestation were primarily cultured for 7 days,and were inoculated in the culture plate at a density of 5×105 cells/ml.The neurons were randomly divided into 3 groups (n =15 each) using a random number table:control group (group C),propofol group (group P),and dexmedetomidine + propofol group (group DP).In group P,propofol with the final concentration of 100 μmol/L was added to the culture medium,and the cells were incubated for 3 h.In group DP,dexmedetomidine with the final concentration of 1 μmol/L was added to the culture medium,the cells were incubated for 30 min,and then propofol with the final concentration of 100 μmol/L was added to the culture medium,and the cells were incubated for 3 h.The viability of hippocampal neurons was assessed by CCK-8 assay.NGF mRNA expression was detected by real-time reverse transcriptase polymerase chain reaction.NGF protein expression was detected by Western blot.Results Compared with group C,the viability of hippocampal neurons was significantly decreased,and the expression of NGF protein and mRNA was down-regulated in group P (P＜0.05).Compared with group P,the viability of hippocampal neurons was significantly increased,and the expression of NGF protein and mRNA was up-regulated in group DP (P＜0.05).Conclusion Dexmedetomidine improve propofol-induced decrease in the viability of isolated hippocampal neurons of fetal rats through up-regulating the expression of NGF.

Objective To investigate the effect of insulin-like growth factor-1(IGF-1 ) on the osteogenic phenotype of fibroblasts. Methods Fibroblasts (Fbs) were derived from an adult New Zealand white rabbit through isolation,purification and cultivation.The experiment was conducted in 3 groups.In the control group,Fbs were cultured in conventional medium without any intervention factors.In the osteogenic induction group,the osteogenic induction medium was composed of conventional medium plus dexamethasone at concentration of 1 × 10-8 mol/L plus vitamin C at 50 mg/L plus sodium glycerophosphate-β at 10 mmol/L.In the experimental group,Fbs were cultured in osteogenic induction medium plus IGF-1 at the final concentration of50 ng/mL.Proliferation of Fbs in each group was detected by methyl thiazolyl tetrazolium (MTT)colorimetric assay.Alkaline phosphatase(ALP) activiiy was detected,osteocalcin 6 days after culture was determined,and the ability of osteogenic differentiation 2 weeks after culture was evaluated by calcium-cobalt staining. Results MTT showed that the cells grew significantly faster in the experimental group than in the other 2 groups( P ＜ 0.05),but there was no significant difference between the control group and the osteogenic induction group in this respect (P ＞ 0.05).The ALP expression in the experimental group was insignificantly higher than in the osteogenic induction group( P ＜ 0.05),but significantly higher than in the control group ( P ＜ 0.05).In osteocalcin secretory activity,the experimental group was significantly superior to the other 2 groups and the osteogenic induction group was significantly superior to the control group ( P ＜0.05).The experimental group had significantly more calcified nodules than the other 2 groups and there were few calcified nodules in the control group 2 weeks after culture. Conclusion IGF-1 can advance proliferation and osteogenic phenotype expression of the Fb stimulated.

OBJECTIVE:To systematically review the effect of ticagrelor versus prasugrel on platelet reactivity,and provide evi-dence-based reference for clinical treatment. METHODS:Retrieved from PubMed,CJFD and Wanfang Database,randomized con-trolled trials(RCT)about the effect of ticagrelor versus prasugrel on platelet reactivity were collected. Meta-analysis was performed by using Rev Man software after data extract and quality evaluation by Cochrane 5.1.0. RESULTS:Totally 17 RCTs were enrolled,involv-ing 2 757 patients. Results of Meta-analysis showed,regardless of Verity Now(VN)detection method [MD=15.43,95%CI(-0.39, 31.25),P=0.06] or vasodilator stimulus phosphoprotein(VASP)detection method [MD=-3.04,95%CI(-8.98,2.90),P=0.32], ticagrelor and prasugrel had the same effects on platelet reactivity under loading dose,the differences were not statistically significant;regardless of VN detection method [MD=-48.94,95%CI(-58.04,-39.84),P<0.001] or VASP detection method [MD=-14.32, 95%CI(-20.45,-8.20),P<0.001],the effects of ticagrelor were better than prasugrel on platelet reactivity under maintenance dose,the differences were statistically significant. CONCLUSIONS:At the loading dose,there was no difference between ticagrelor and prasugrel,but ticagrelor has more benefits than prasugrel under maintenance dose.

Objective To investigate the effects of inhibiting NgR on retinal ganglion cells density and synaptophysin expression of diabetic rat.Methods Thirty-two SD male rats were randomly divided into normal control group,diabetic group,siNgR group and scNgR group,8 rats in each group.Normal control group was given no any treatment.Diabetes model was induced by intraperitoneal injection of 50 mg · kg-1 streptozotocin in diabetic group,siNgR group and scNgR group,and the blood giucose more than 16.7 mmol · L-1 at 72 hours was set as the successfully model.The rats of siNgR group were intravitreally administrated with anti-NgR nucleotide and the rats of scNgR group intravitreally administrated with negative nucleotide.Eight weeks later,HE staining was conducted to detect density of retinal ganglion cell (RGC),immunofluorescence was used to evaluate the expression and distribution of synaptophysin (a marker of synaptic number).Relative expression of NgR and synaptophysin in retina were analyzed by Western blot.Results RGC density in normal control group,diabetes group,siNgR group and scNgR group were (624.33 ± 3.51) mm-2,(420.00 ± 2.65) mm-2,(621.67 ± 1.53) mm-2,(416.67 ± 2.52) mm-2,respectively.There was significant difference among four groups (F =5985.37,P ＜ 0.01).Compared with normal control group,RGC density in diabetes group and scNgR group were obviously decreased (all P ＜0.01),but siNgR group had no obviously change (P ＞ 0.05).The synaptophysin mainly expressed in the inner and outer network layer.Compared with normal control group,the positive expression of synaptophysin in diabetes group and scNgR group were decreased,but siNgR group had no obviously change.The relative expression of NgR in normal control group,diabetes group,siNgR group and scNgR group were (11.26 ±0.02) ％,(19.38 ± 0.10) ％,(11.17 ± 0.02) ％,(19.47 ± 0.31) ％,respectively.There was significant difference among four groups (F =2466.09,P ＜ 0.01).Compared with normal control group,the relative expression of NgR in diabetes group and scNgR group were obviously decreased (all P ＜ 0.01),but siNgR group had no obviously change (P ＞0.05).The relative expression of synaptophysin in normal control group,diabetes group,siNgR group and scNgR group were (35.76 ± 0.15) ％,(25.47 ± 0.36) ％,(35.28 ± 0.12) ％,(25.03 ± 0.75) ％,respectively.There was significant difference among four groups (F =583.70,P ＜ 0.01).Compared with the normal control group,the expression of synaptophysin in diabetic group and scNgR group were decreased increased (all P ＜ 0.01),while there was no significant difference in siNgR group (P ＞ 0.05).Conclusion Inhibiting the expression of NgR in the retina of diabetic rats can help to restore the number of synapses and protect the damaged RGC.

Currently,the toxicity study of traditional Chinese medicine is faced with the following problems. Firstly,the evaluation in vitro cannot fully reflect the true state of the body. Secondly,the traditional method is not sensitive enough to the early toxicity. Lastly,the toxicity evaluation indexes cannot determine whether the compatibility of traditional Chinese medicine produces toxicity or increases toxicity systematically. The paper proposed a synthesized early evaluation research method for target organ toxicity induced by traditional Chinese medicine:screening,validation,optimization and application. This method mainly inoolves early target organ toxicity biomarkers in screening,optimi?zation,validation,biological significance explanation,and application to the traditional Chinese medicine incompatibility based on the metabolic dynamic fingerprint spectrum in order to obtain biomarkers of target organ toxicity that are sensitive and precede conventional biochemical indices for early evaluation . We attempted to analyze the pattern of chang of the biomarkers for animals acted by″18 incompatible medicaments″compatibility combination. We found that Radix Aconiti Lateralis Preparata with cardiotoxicity were compatible with Rhizoma Pinelliae,and that Trichosanthes kirilowii Maxim,Fritillaria,Ampelopsis Radix and Bletilla striata without non-cardiotoxicity produced and increased cardiotoxicity systematically.

OBJECTIVE To study characteristics of changing T lymphocytes in epidemic hemorrhagic fever(EHF) patients during acute phase and find out the pathogenesis,in order to elevate the level of early diagnosis.METHODS The anticoagulant blood from 30 cases of EHF patients and 50 normal healthy blood donors was collected.T lymphocyte subsets were detected by flow cytometry.RESULTS Compared with those of normal persons,CD4+ T cell counts of EHF patients decreased,CD8+T cell and double CD4+CD8+ cell(double positive cells,DP cell) counts of EHF patients increased obviously,and 25 cases of EHF in recovery stage returned to normal.And in comparison with HIV,CMV and EBV patients,DP cell counts of EHF patients increased obviously.CONCLUSIONS T lymphocytes of EHF decrease obviously but could be resumed,detection of amounts of lymphocyte subsets and CD4+CD8+ cells can provide an early diagnosis method to EHF.

A15, a DNA aptamer with binding specificity for U87 glioma cells stably overexpressing the epidermal growth factor receptor variant III (U87-EGFRvIII), was generated by cell systematic evolution of ligands by exponential enrichment (cell-SELEX) using a random nucleotide library. Subsequently, we established a cell enzyme-linked assay (cell-ELA) to detect the affinity of A15 compared to an EGFR antibody. We used A15 as a detection probe and cultured U87-EGFRvIII cells as targets. Our data indicate that the equilibrium dissociation constants (K(d)) for A15 were below 100 nmol/L and had similar affinity compared to an EGFR antibody for U87-EGFRvIII. We demonstrated that the cell-ELA was a useful method to determine the equilibrium dissociation constants (K(d)) of aptamers generated by cell-SELEX.
Aptamers, Nucleotide ; metabolism ; Brain Neoplasms ; metabolism ; Cell Line, Tumor ; Glioma ; metabolism ; pathology ; Humans ; Mutation ; Protein Binding ; Receptor, Epidermal Growth Factor ; genetics ; metabolism ; SELEX Aptamer Technique ; methods