In central nervous system only a limited number of vesicles exist in the presynaptic terminals. The size and fusion modes of the vesicles were particularly important because of their potential impact on neuronal communications. Efficient methods were needed to analyze the recycling kinetics of synaptic vesicle and the size of readily releasable pool (RRP). In this study, fluorescent dyes with different affinity for membranes (FM1-43 with high affinity and FM2-10 with low affinity) were used to stain the functional synaptic vesicles of cultured hippocampal neurons and the kinetics of vesicle recycling was measured. The results showed that the destaining proportion was larger for FM2-10 than that for FM1-43 during the first trial, while it was greater for FM1-43 than FM2-10 during the second and third trials (first round, 93.0%+/-5.9% versus 57.9%+/-3.5% for FM2-10 and FM1-43, respectively, P<0.0001; second round, 1.4%+/-3.8% versus 24.0%+/-2.3%, P<0.0001; third round, 2.3%+/-1.6% versus 8.6%+/-1.5%, P=0.005). The results indicated that rapid endocytosis existed not only in the first round but also occurred when the vesicles were reused. Moreover, Both high-frequency stimuli and hypertonic sucrose stimuli were used to estimate the RRP sizes in the mix cultured hippocampal inhibitory neurons at 13-14 days in vitro (DIV). We found that the RRP size estimated by hypertonic sucrose stimuli [(200+/-23.0) pC] was much larger than that estimated by high-frequency stimuli [(51.1+/-10.5) pC]. One possible reason for the discrepancies in RRP estimates is that in mix cultured conditions, one neuron may receive inputs from several neurons and hypertonic sucrose stimuli will cause RRP of all those neurons release, while using dual patch recording, only the connection between two neurons was analyzed. Thus, to exclude out the impacts of inputs numbers on RRP sizes, it is more reasonable to use high-frequency stimuli to estimate the RRP size in mix cultured neurons.
Cells, Cultured ; Endocytosis ; Hippocampus ; cytology ; Neurons ; physiology ; Synaptic Vesicles ; physiology

OBJECTIVETo discuss the development module of ontology-based computerized clinical practical guideline, and to supply the technology support for implement of alerm/reminding, data sharing, evidence-based medicine and clinical decision making in the medical information system.

METHODSAnalyse the structure and field ontology of the Guideline Interchange Format (GLIF); expand GLIF based on the Guideline of Cerebrovascular Disease Prevention and Treatment in China and Neurology Disease Ontology.

RESULTSA Chinese computerized guideline of intravenous thrombolytic therapy of acute cerebral infarction is constructed, which included metadata of edition description, list of drug data item, explanation of evidence strength, steps of indications decision, contraindication decision and drug selection.

CONCLUSIONThe computerized clinical practical guideline combined with clinical information system and Electronic Medical Records plays an important role in clinical pathways optimizing and decision making.

Decision Making, Computer-Assisted ; Decision Support Systems, Clinical ; Medical Records Systems, Computerized ; Practice Guidelines as Topic ; Research Design

Objective To analyze the influence of age on the diagnosis of acute heart failure (AHF) with amino terminal brain natriuretic peptide (NT‐proBNP) ,and to explore the best cut‐off value of NT‐proBNP in the diagnosis of AHF .Methods Totally 457 inpatients suspected with AHF obtained a blood NT‐proBNP measurement within 24 hours when in charged ,including 194 patients with AHF .Patients were divided into three groups according to their age :≤50 years ,>50-70 years and >70 years . The relationship between NT‐proBNPand AHF was analyzed .The diagnostic value of NT‐proBNP was evaluated for AHF in differ‐ent age groups .Results The median NT‐proBNP levels of AHF in all age groups were significantly higher than those without (P<0 .01) .AUC of NT‐proBNP for AHF in ≤50 years ,>50-70 years and >70 years groups were 0 .95 ,0 .91 ,0 .88 ,the best diagnos‐tic cut‐off points were 1 100 ,1 800 ,3 000 pg/mL ,respectively .The diagnostic specificity was 88% ,76% and 73% ,respectively .O‐verall ,the specificity of NT‐proBNP cut‐off values for age group was higher than non‐age groups .Conclusion NT‐proBNP has a good value in the diagnosis of AHF in patients with different ages .According to the age group ,the different NT‐proBNP cut‐off values can be used to improve the specificity of diagnosis .

AIM: To investigate the affected proteins by tumor necrosis factor (TNF)-? in endothelial cells, and further explore the potential molecular mechanism of TNF-? on endothelial cells. METHODS: Nitric oxide (NO) production in the cultured human umbilical vein endothelial cells (HUVECs) was measured by a NO assay kit. Proteomic alterations were analyzed using two-dimensional electrophoresis, and peptide mass fingerprinting with matrix-assisted laser desorption/ionization-time of flight mass spectrometry. RESULTS: NO production in HUVECs decreased significantly after TNF-? treatement. Proteomics analysis showed 21 protein spots were changed including 9 spots that were increased and 11 spots that were decreased after TNF-? stimulation, and 1 spot was only detected in TNF-? activated cell gels. CONCLUSIONS: The decreased expression of ecNOS by TNF-? might result in decrease in NO production. Up-regulated MAP/ERK kinase 3 expression might imply that TNF-? activates the expression of adhesion molecules. Cytoskeletal protein actin is also involved in TNF-? injuried HUVECs. Proteomic analysis can find some clues for identifying new potential target of TNF-?. [

Objective To study the effects of different glucocorticasteroid(GCS) inhalation regimens for remissive children with asthma. Methods Three hundred and twenty - three patients with moderate asthma were enrolled on a 12 - week randomized parallel group remissive treatment after a 4 - week baseline treatment. During the baseline treatment terbutaline sulfate 250 ?g tid a day and bud esonide 200 ?g twice a day were given, and oral bronchodilators were used if necessary. The remissive treatment were composed of budesomde inhabit ion 100 ?g once a day (group A), 100 ?g twice a day(group B) and 200 ?g once a day(group 0). Patients subsequently returned to the clinic for 3 additional clinic visits (4,8 and 12 weeks) or telephone visits . On every clinic visit, the daytime and nocturnal time seventy score were recorded and spirometry was conducted in patients who were capable of performing the maneuver. Results Ultimately, 323 children were enrolled on the baseline treatment and 281 (87%) children achieved clinical remission. The rate of compliance decreased gradually during the remissive treatment, but in group B(P

OBJECTIVE To investigate the HIV infections states in tumor patients for clinical diagnosis,treatment and to prevent HIV infection in the tumor hospital.METHODS The result of HIV detection in tumor patients from Dec 2000 to Aug 2006 was analyzed by the review statistics analysis.RESULTS Totally 48 101 tumor paients were detected,and the number of tumor patients with positive HIV antibody result was 51(0.106%).Among the positive patients there were 21 cases with blood transfusion history,14 cases with blood donating experience,2 cases with both these two kinds of experiences and 14 cases without the two kinds of experiences.Their rate was separately 41.0%,27.5%,4.0% and 27.5%.Most of the HIV positive patients had no clinical synptoms.CONCLUSIONS The HIV positive rate of patients with blood transfusion or blood donating is significantly higher than the patients without these experiences.The routine detection for the HIV before the operation,blood transfusion or other traumatic detection is very necessary.

Objective To establish a simple and economic method for isolating rat hepatic lipocytes. Method Perfusion of the rat liver with pronase E and collagenase in succession was performed in cycles, and the hepatic lipocytes were isolated and purified by digesting the rat liver with DNase in the presence of the above 3 enzymes, followed by 11% Necodenz density gradient centrifugation. The resulted lipocytes were subjected to routine identification procedures and viability assessment. Results The yield of the lipocytes from each rat was 2×107 to 3×107, with viability over 95% and purity above 90%. Conclusion The method adopted in this study is simple, economic and highly productive.

Objective To establish a simple and economic method for isolating rat hepatic lipocytes. Method Perfusion of the rat liver with pronase E and collagenase in succession was performed in cycles, and the hepatic lipocytes were isolated and purified by digesting the rat liver with DNase in the presence of the above 3 enzymes, followed by 11% Necodenz density gradient centrifugation. The resulted lipocytes were subjected to routine identification procedures and viability assessment. Results The yield of the lipocytes from each rat was 2×107 to 3×107, with viability over 95% and purity above 90%. Conclusion The method adopted in this study is simple, economic and highly productive.

BACKGROUNDTo investigate the biological functions of a novel hepatitis B virus e antigen (HbeAg) interacting protein AK026018, and to use cDNA microarray technique to screen genes regulated by the protein.

METHODSThe AK026018 coding DNA fragment was amplified by reverse transcription polymerase chain reaction (RT-PCR) technique from HepG2 cell. The expressive vector of pcDNA3.1-AK was constructed by routine molecular biological methods. The HepG2 cells were transfected with pcDNA3.1 and pcDNA3.1-AK, respectively by using lipofectamine. The total RNA was isolated and reverse transcribed. The cDNA of each sample was subjected to microarray screening with 8,464 cDNA probes and analyzed by bioinformatics.

RESULTSThe expressive vector was constructed and confirmed by DNA sequencing analysis and restriction enzyme digestion. High quality mRNA and cDNA of transfected HepG2 cells had been prepared and successful microarray screening conducted. From the scanning results, there were 122 differential expression genes, of which 36 genes were down-regulated, and 16 genes were up-regulated.

CONCLUSIONMicroarray technique was successfully used to screen the genes trans-regulated by AK026018. The expression of AK026018 protein affects the expression spectrum of HepG2 cells.

Carrier Proteins ; genetics ; metabolism ; physiology ; Cell Line, Tumor ; Computational Biology ; Gene Expression Regulation, Neoplastic ; Hepatitis B e Antigens ; metabolism ; Humans ; Oligonucleotide Array Sequence Analysis ; methods ; Protein Binding ; Reverse Transcriptase Polymerase Chain Reaction ; Transfection

Bocavirus ; isolation & purification ; Humans ; Infant ; Male ; Oligonucleotide Array Sequence Analysis ; methods ; Parvoviridae Infections ; diagnosis ; Pneumonia ; diagnosis