Aim To evaluate the value of expressions of CD63 on basophils in diagnosing allergy to penicillins.Methods The expressions of CD63 on basophils activated with 9 kinds of antigens(PG:penicillin G,PV:phenoxomethylpenicillin,AMP:ampicillin,AX:amoxicilloyl,6-APA:6-aminopenicillanic acid,PHA:phenylacetic acid,PHOA:phenoxacetic acid,PHPG:N-(?-hydroxyphenyl)glycine,NPG:N-phenylglycine) were detected by FAST(Flow cytometric allergen stimulation test) in 43 patients with penicillins allergy.Eight types of specific IgE and IgG antibodies to penicillins were determinated by RAST and ELISA,respectively.Results The sensitivity and specificity of CD63 used as a marker in patients with penicillins allergy were 65.12% and 93.33%,respectively.Moreover,the sensitivities of CD63 and specific IgE were higher than those of specific IgG in patients with positive skin test(P

Objective To analyse the clinical features,imaging characteristics diversity of deep cerebral veins thrombosis (DCVT).Methods From 2004 to 2013,6 patients diagnosed as DCVT were recorded and a retrospective review of the cases were undertaken for the purpose of this analysis.Results Among the 6 patients with DCVT,4 were male and 2 were female,aged from 28 to 69 years old.The disease duration of 4 cases ranged from 2 to 7 days,remnants were 20 days and 3 months respectively.The first symptoms of 4 cases were headache,1 was feeblemindedness,and the other was hemiplegia.The secondary symptoms were disturbance of consciousness,apathy,diplopia and non-infectious fever.Non-contrast computed tomography showed low signal in the bilateral thalamus in four patients,high signal in the transverse sinus and straight sinus in one patient and high signal in torcular in one patient.Abnormal signal was found in bilateral thalamus on magnetic resonance imaging in all patients and some of them had abnormal signal in the mesencephalon or basal ganglia.The patients were definitely diagnosed as DCVT by magnetic resonance venography (MRV) or digital subtraction angiography (DSA).Among them,2 patients were confirmed by brain biopsy.Four patients were followed up with good outcome and 2 were lost to follow-up.Conclusions The clinical manifestations of DCVT are not specific.For acute-onset DCVT patients,the first symptoms are always headache and vomiting,while the main symptoms are declined cognition and slow reaction for chronic-onset ones.Along with the progress,the main symptoms of DCVT are disturbance of consciousness,psychiatric symptoms and intracranial hypertension.Changes in the bilateral thalamus and basal ganglia are especially main characteristics which are easily misdiagnosed as brain tumor according to the images.DCVT can be definitely diagnosed by no signal of deep cerebral veins on MRV or DSA.

Objective To observe the clinical efficacy and safety of ultrapulse CO 2 laser-assisted au-tologous epidermal grafting for the treatment of vitiligo.Methods1605patients with30660skin grafts were treated by CO 2 laser-assisted autologous epidermal grafting.Among them,782patients had localized vi-tiligo,319segmental,226general and278scattered.Epidermal grafts were harvested with negative pressure blistering method from normally pigmented skin,and recipient areas were prepared with a ultrapulse CO 2 laser.The curative effect was evaluated8weeks later.Results79.04%of the lesions were cured complete-ly,and total response rate was92.21%,with better effect in localized and scattered lesions than in other le-sions.The best results were shown in face,while the worst results shown in brow and hairline lesions.Con-clusions The advantage of combined therapy of CO 2 laser with autologous epidermal grafting in vitiligo is of easy performance,convenience,high response rate,which is worthy of large scale application.

Objective To prepare stable aqueous dispersions of chitosan/multi-walled carbon nanotubes (CS/MWCNTs) composites,and observe the effects of CS/MWCNTs on the growth of human umbilical vein endothelial cells (HUVEC).Methods CS/MWCNTs composites were prepared by electrostatic interactions between negatively charged MWCNTs and positively charged low-molecular-weight CS.The prepared CS/MWCNTs were characterized by transmission electron microscopy and Zetasizer nano-analyser.The cellular uptake of the fluorescently labeled CS/MWCNTs was observed by laser confocal microscopy after incubating with HUVEC for 24 h at different concentrations.In vitro cytotoxicity and cellular reactive oxygen were also detected.Results When the mass ratio of low-molecular-weight CS to MWCNTs was equal or greater than 10∶1,the CS/MWCNTs can be stabilized in solution.Cellular uptake experiments showed that the CS/MWCNTs could enter into the cells and locate mainly in the cytoplasm.Cytotoxicity study showed that the CS/MWCNTs composites was less toxic than MWCNTs alone at high concentration (10 and 20 μg/ml).However,there was no significant differencein the level of cellular reactive oxygen between the two groups (P＜0.05).Conclusions CS/MWCNTs composites showed low cytotoxicity and high stability,which would be a promising carrier for drug delivery.

Diabetes mellitus is a chronic disease, it is thus necessary for its patients to participate in its treatment and self-management. The emerging foreign and domestic methods for self-management of diabetes mellitus pa-tients were detected on different Websites and suggestions were proposed for its treatment and self-management in order to improve the quality of life in diabetes mellitus patients.

Objective To compare the imaging characteristics of multiple sclerosis (MS) and neuromyelitis optica (NMO) for better diagnosis and differential diagnosis.Methods The brain and spinal MRI images of 60 MS and 48 NMO cases were retrospectively reviewed.The imaging characteristics including the predilection site,morphological features,enhancement manifestations were summarized.All data was analyzed by using t test and Chi square test with SPSS 13.0.Results (1) The three top predilection sites of brain in head MRI of MS patients were periventricular white matter (34 cases in 60),subcortical white matter (27 cases in 60),brain stem (23 cases in 60).MS lesions also were found in basal ganglia,cerebellum,corpus callosum and thalamus,as well as cortex (9 cases in 60).By contrast,brain lesions were observed in 59.4％ (19/32) of NMO patients,and the three top predilection sites of NMO by turns were brain stem (13 cases in 19),periventricular white matter (12 cases in 19),subcortical white matter (7 cases in 19).Furthermore,the lesions surrounding third ventricle (6 cases in 19) and the tegmentum of brain stem near peri-aqueduct (8 cases in 19) in NMO were not found in patients of MS.The involvement of brain stem and thalamus was more frequent in NMO than in MS (x2 =5.267,6.004,P ＜0.05,respectively).(2) The lesions of spinal cord in MS patients were typically oval,peripheral,and asymmetric,but in NMO patients they were longitudinally extensive and centrally located.The mean number of involved vertebral segments in NMO patients was significantly more than that in MS patients (7.3 vs 2.2,t =-9.288,P ＜ 0.01).Furthermore,the number of spinal cord lesions in MS patients was remarkably more than that in NMO (2.0 vs 1.3,t =4.565,P ＜0.01).The ratios of occurrence of spinal cord swelling and distension of NMO patients was 58.3％ (28/48),which was significantly higher than 21.9％ in MS (7/32,x2 =10.370,P ＜0.01).(3)The enhancement pattern in MS was circular (7 cases in 42),oval (6 cases in 42) and irregular (4 cases in 42),while in NMO was mainly sheet-shaped with mild enhancement (5/11).The lesions of spinal cord showed in MS mainly manifested as oval enhancement (16 cases in 26) and linear enhancement (8 cases in 26),while in NMO lesions manifested as strand or mild linear enhancement (26 cases in 35).Conclusions NMO has several distinct imaging characteristics,which are helpful for differentiation from MS.

ObjectiveTo isolate and identify endothelial progenitor cells (EPCs) from human umbilical cord,and to study the cell proliferation and gene transfection of green fluorescent protein plasmid in vitro.MethodsEPCs were isolated from human umbilical cord in enzyme digestion method.The biological characteristics of EPCs were identified by flow cytometry and laser confocal microscope.The enhanced green fluorescent protein (EGFP) gene transfection mediated by EPCs was investigated using Lipofectamine 2000 as transfection reagent.ResultsEndothelial progenitor cells isolated from umbilical cord formed typical endothelial cell colony 9 days later.These cellsdisplayed an improved positive expression of CD133 and kinase insert domain receptor (KDR).The endotheliallineage characteristics of expanded cells were confirmed by fluorescein isothiocyanate (FITC)-UEA-1 binding and DiI-ac-LDL uptake assay with the aid of laser confocal microscope.The transfection results demonstrated high expression of EGFP taking EPCs as host cell.ConclusionEndothelial progenitor cells isolated from umbilical cord can be propagated and induced to differentiate into endothelial cells in the appropriate culture conditions.EPCs demonstrated to be an ideal carrier for gene and cell therapy.

ObjectiveConjugation of fluorescent dye onto plasmid DNA was investigated in order to monitor delivery process of plasmid DNA.MethodsPlasmid was activated with bromine,stored for different timeintervals at 4 ℃ or room temperature,and subsequently coupled with 1,10-diaminodecane to prepare aminemodified plasmid DNA.Amine-modified plasmid was then reacted with isothiocyanate (FITC) for fluorescent labeling,and the labeling ratio was calculated after purification.The effect of storage conditions (time/temperature) of bromine-actived plasmid (BP) on fluorescent labeling efficacy was estimated,and the cell transfection efficiency of fluorescent plasmid-lipofectamine complex was observed.The fluorescent plasmid delivered by lipofectamine 2000 in A10 cells was observed by laser scanning confocal microscope (LSCM) and flow cytometry.ResultsThe experimental data showed that prolonged storage time of bromine-activated DNA had a negative effect on the labeling ratio,and lower storage temperature had a positive effect on the labeling ratio.It also demonstrated that FITC modification had no effect on the transfection efficiency of plasmid-lipofectamine complex as compared with that of unlabeled plasmid-lipofectamine complex,and FITC modified plasmid had enough fluorescent intensity to monitor cell uptake with flow cytometer and sub-cellular distribution with LSCM.ConclusionA facile method for conjugating fluorescent dye onto plasmid was established in the study,and could be utilized to trace the plasmid delivery for investigating the transfection mechanism.

Objective To investigate the impact of chitosan and alkylated chitosan DNA nanoparticles on the function of human naive CD4+T cells.Methods The secretion of cytokines (IL-4 and TNF-γ) was observed after the co-incubation of human naive CD4+T cells with nanoparticles 12 h,24 h and 48 h,respectively.ResultsNone of the nanoparticles induced the production of cytokines ( IL-4 and TNF-γ ).Conclusion Chitosan and alkylated chitosan DNA nanoparticles will not induce the differentiation of human naive CD4+ T cells into T1 or T2 and may be considered as a safe gene carrier.

Objective To generate recombinant Pichia pastoris for high-copy expression of human tissue factor pathway inhibitor (TFPI). Methods The cDNA encoding human TFPI was inserted into the expression vector pPIC9K and the constructed expression vector rhTFPI-pPIC9K was confirmed by restriction endonuclease analysis and DNA sequencing. The recombinant plasmids were subsequently transformed into Pichia pastoris GS115 cells, and the transformants were confirmed by PCR amplification of the genomic DNA.The recombinant Pichia pastoris with high copies of TFPI cDNA was screened by G418 selection. Western blot and TFPI ELISA Kit were employed to analyzing. The temperature, time and concentration of methanol for the induction of recombinant protein were optimized. Results PCR analysis and DNA sequencing confirmed the successful construction of the expression vector rhTFPI-pPIC9K. Real time quantitative PCR and Western blot analysis demonstrated the positive correlation between TFPI expression level and the copy number of TFPI cDNA in Pichia pastoris cells. Optimization of the induction condition significantly elevated the expression level and activity of TFPI (9.95±0.78 mg/L and 3.91±1.37 U/mL). Conclusion The Pichia pastoris strain with high copy of TFPI expression was successfully constructed, which lays a solid foundation for the further investigation on the function of TFPI and its application in the prevention and therapy of diseases.