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To establish an assessment index system to objectively evaluate the implementation of health promotion in enterprises.Multi-methods, which include the reviewing references, the summarizing results of preliminary studies, the interview with experts and employers, were used in developing attentative index framework and working out the consultative questionnaires. Then the improved Delphi Method was adopted in collecting ideas from 20 experts in China, and they scored the importance of every single index by two rounds of consultation. On the basis of these data, the indicators were added , deleted or modified according to concentration and distraction levels of expert ideas. The method of the percentile weighted coefficient was conducted to decide the weighing of assessment index.The responding rates to the two rounds of questionnaires were 90.91% and 100.00% respectively. The overall specialist authoritative coefficient was 0.69, the coordination coefficient of all indicators was 0.623 (P0.01),and the coordination coefficient of the four kinds of first-ranking indicators was 0.313, 0.625, 0.390, 0.563, respectively (P0.01).Eventually, an assessment index system consisting of 4 first-ranking indicators and 10 sub-indicators which were further divided into 52 grade-three indicators was set up.The assessment index system possesses good content validity, and the liability and typicality were recognized by experts. However, it should be applied and validated in practice.

Preventive maintenance program was one of the basic requirements to the nuclear power operation units set by national nuclear safety authorities. This paper describes the specific processes with respect to the compilation of preventive maintenance for equipment at nuclear power plants (NPPs), with a combination of domestic and foreign maintenance optimization methods, and RCM analysis results, AP913 method, experience feedback and other methods to optimize preventive maintenance program, which has created considerable economic benefits and achieved good results in operating and new NPPs. This paper presents the strategies and methods of compilation and optimization of preventive maintenance program, providing an important reference for compilation and optimization of maintenance program for NPPs at home and abroad.

To study the chemical constituents of Rhododendron seniavinii.Compounds were isolated from the aqueous extract of the leaves of R. seniavinii by using Sephadex LH-20, ODS open column chromatography and other means. Their structures were elucidated according to spectral data and physiochemical properties.Thirteen compounds were isolated from R. seniavinii and identified as 5-methoxydehydroconiferyl alcohol (1), dehydroconiferyl alcohol (2), (-)-syringaresinol (3), (-)-lyoniresinol (4), (+)-lyoniresinol 3alpha-O-beta-D-glucopyranoside (5), (-)-lyoniresinol 3alpha-O-beta-D-glucopyranoside (6), 3,4,5-trimethoxyphenyl-1-O-beta-D-glucopyranoside (7), nikoenoside (8), 3,5,7-trihydroxychromone-3-0-alpha-L-rhamnopyranoside (9), 3,4,5-trimethoxyphenol (10), scopoletin (11), scopolin (12) and quercitrin (13).Compounds 1-12 were obtained from this plant for the first time.

High-altitude pulmonary edema (HAPE) is a non-cardiogenic type of pulmonary edema developing altitudes > 2,500 m. Angiotensin converting enzyme (ACE) and nitric oxide synthase 3 (NOS3) play important roles in regulating pulmonary vascular tone. To assess associations between genetic variants in the ACE and NOS3 genes and HAPE risk, 27 HAPE patients and 108 matched controls were genotyped and analyzed. The indicated HAPE association of the NOS3 G894T (Glu298Asp) single nucleotide polymorphism (SNP), which may change NO production, was further evaluated by a meta-analysis of six studies involving 399 HAPE patients and 495 controls. Odds ratios (ORs) and 95% confidence intervals (CIs) were determined with fixed-effects models. Stratification analyses of ethnicity and geographic location were performed. Significant associations were observed for the dominant model in two ACE tag SNPs influencing serum ACE concentrations (rs8066114 polymorphism: GG+CG vs. CC; rs4461142 polymorphism: TT+CT vs. CC). Furthermore, Single-locus analysis indicated significantly different distributions of G allele frequency between the cases (29.63%) and controls (17.13%) for the ACE rs8066114 polymorphism. The case-control distributions of genotype frequencies and T allele frequency of NOS3 G894T (Glu298Asp) polymorphism were significantly higher in the cases than controls, and the NOS3 G894T (Glu298Asp) SNP showed elevated HAPE risk under the dominant model (TT+GT vs. GG). Meta-analysis showed overall association of NOS3 G894T SNP with HAPE risk under the allele contrast and dominant genetic models, which remained significant for Asians. In conclusion, ACE rs8066114 and rs4461142 and NOS3 rs1799983 (G894T) polymorphisms may be associated with increased HAPE risk in Asians.

Density functional theory (DFT) calculations are performed to investigate the reactivity of Th atom toward ethane C–C bond activation. A comprehensive description of the reaction mechanisms leading to two different reaction products is presented. We report a complete exploration of the potential energy surfaces by taking into consideration different spin states. In addition, the intermediate and transition states along the reaction paths are characterized. Total, partial, and overlap population density of state diagrams and analyses are also presented. Furthermore, the natures of the chemical bonding of intermediate and transition states are studied by using topological method combined with electron localization function (ELF) and Mayer bond order. Infrared spectrum (IR) is obtained and further discussed based on the optimized geometries.

The CBL-CIPK signaling pathway represents a central and critical signaling system involved in plant response to abiotic stress and hormone signaling. In this study, we focused on AtCIPK6 gene, which have been shown to be required for development and salt tolerance in Arabidopsis, but the transcriptional regulatory mechanism of AtCIPK6 gene and whether it plays a role in ABA signaling is still unknown. Here, we showed that in addition to its expression induced by salt and drought stress, the transcripts of AtCIPK6 gene were largely accumulated in abscisic acid treated seedling, compared to basal level expression. Moreover, promoter sequence analysis revealed that there are some elements involved in stress and hormone response detected in the promoter region of AtCIPK6. AtCIPK6p:GUS transgenic assays unraveled that AtCIPK6 promoter is salt/osmotic stress- and ABA-inducible. Overexpression of AtCIPK6 gene in Arabidopsis increased plant tolerance to salt stress, but sensitivity to ABA. Our results contribute to the understanding of transcriptional regulatory mechanism of AtCIPK6 gene and may also provide a good stress-inducible promoter candidate for transgenic engineering.

RNA interference (RNAi), as a new technology of gene therapy, has been used in the studies of many diseases in vitro, however, targeting delivery of small interference RNA (siRNA) is still a bottleneck for clinical therapy of siRNA agents. Aptamer is a group of oligonucleotides with high affinity and targeting, and is becoming another important means of delivery for siRNA. In this review, we summarized siRNA delivery obstacles in vivo and recent attractive developments increatively using cell-internalizing aptamers to deliver siRNAs to target cells.

Neural tube defects (NTDs) are severe birth malformations that affect one in 1,000 live births. Recently, mutations in the planar cell polarity (PCP) pathway genes had been implicated in the pathogenesis of NTDs in both the mouse model and in human cohorts. Mouse models indicate that the homozygous disruption of Sec24b, which mediates the ER-to-Golgi transportation of the core PCP gene Vangl2 as a component of the COPII vesicle, will result in craniorachischisis. In this study, we found four rare missense heterozygous SEC24B mutations (p.Phe227Ser, p.Phe682Leu, p.Arg1248Gln, and p.Ala1251Gly) in NTDs cases that were absent in all controls. Among them, p.Phe227Ser and p.Phe682Leu affected its protein stability and physical interaction with VANGL2. Three variants (p.Phe227Ser, p.Arg1248Gln, and p.Ala1251Gly) were demonstrated to affect VANGL2 subcellular localization in cultured cells. Further functional analysis in the zebrafish including overexpression and dosage-dependent rescue study suggested that these four mutations all displayed loss-of-function effects compared with wild-type SEC24B. Our study demonstrated that functional mutations in SEC24B might contribute to the etiology of a subset of human NTDs and further expanded our knowledge of the role of PCP pathway-related genes in the pathogenesis of human NTDs.

To study the pharmacokinetics of cantide, an antisense oligonucleotide, and its metabolites after iv gtt administration in rhesus monkeys, a dual solid phase extraction pretreatment method coupling with non-gel sieving capillary electrophoresis analysis method was used for determination of cantide and its metabolites in plasma and their pharmacokinetic parameters were calculated. The pharmacokinetic behavior of cantide and its metabolites (M1 and M2) after iv gtt administration (8, 16 and 24 mg kg(-1)) in rhesus monkeys were investigated. After iv gtt administration of cantide to rhesus monkeys, cantide in plasma was eliminated rapidly and the terminal elimination half-life (t1/2) was 57.91-77.97 min, the correlation coefficients (r) to the dose of Cmax AUC(o-inf) and AUC(0-t) of the prototype was 0.9918, 0.9568 and 0.9773, respectively. The metabolites of cantide reached the Cmax following cantide immediately and the Cmax of metabolites were lower than that of the prototype. The CL(S) of cantide and its metabolites (M1 and M2) were 1.60-2.19, 5.92-8.58 and 6.07-8.78 mL min(-1) kg(-1), respectively. So, it is concluded that the Cmax of cantide and its metabolites increased with the dose, which is the same as their AUC(0-inf) and AUC(0-t). The CL(S) of metabolites were higher than that of the prototype. The MRT and t1/2 of metabolites in the high dose group increased obviously.

The distribution characteristics of polycyclic aromatic hydrocarbons in runoff from the middle line source area of south-to-north water diversion project were studied. Five groups of artificial runoff fields were established to collect runoff based on the different types of land-use, the contents of 16 USEPA priority PAHs in the runoff were determined using GC/MS method. The results showed that the average concentrations of PAHs of the aqueous phase in the collected runoff samples of different land-use types decreased in the order:cultivated land (26.53 ng x L(-1))oak forest (20.91 ng x L(-1))orchard (17.59 ng x L(-1)), and the average concentrations of PAHs of the particle phase were cultivated land (1 073.72 ng x g(-1))orchard (652.29 ng x g(-1))oak forest (385.46 ng x g(-1)). The high carcinogenic components Bap were detected in both run off of cultivated land and orchard with a detected rate of 30%. According to National Recommended Water Quality Standards of priority toxic pollutants (2006 USEPA), it was found that Chr exceed standard 40%, with a detected rate of 100%. It was also found that the runoff volume and the total PAHs content in runoff increase with the slope, and PAHs loss and slope were closely related in same land-use types. Based on the Molecular Markers Indicative Law, it can be concluded that the dominant source of PAHs in runoff of study area was combustion of coal, and a small amount came from vehicle exhaust emissions. There is a certain degree of ecological risk about runoff PAHs pollution in the study area, which is worth further attention.

Polyethylenimine-cyclodextrin-tegafur (PEI-CyD-tegafur) conjugate was synthesized as a novel multifunctional prodrug of tegafur for co-delivery of chemotherapeutic agent tegafur and enhanced green fluorescent protein (EGFP) reporter plasmid DNA. Conjugation of tegafur to PEI-CyD via chemical linkage was characterized by (1)H NMR spectrometry and ultraviolet (UV) spectrometry. PEI-CyD-tegafur was able to condense plasmid DNA into complexes of around 150 nm with positive charge at the N/P ratio of 25, in accordance with electron microscopy observation of compact and monodisperse nanoparticles. The results of in vitro experiments showed enhanced cytotoxicity and considerable transfection efficiency in B16F10 cell line. Therefore, PEI-CyD-tegafur may have great potential as a co-delivery system with anti-cancer activity and potential for gene delivery.

To examine the influence of chemotherapy with FOLFOX protocol (CT-F) on the immunologic function in patients with advanced colorectal cancer.A total of 43 patients with advanced colorectal cancer were included. Patients who received FOLFOX chemotherapy (Group A, n=22) were compared to those who did not(Group B, n=21). Blood was obtained from peripheral vein before chemotherapy (T0), at the time of completion of chemotherapy (T1), and 3 months after completion of chemotherapy (T2) to detect the percentage of regulatory T lymphocytes (CD4+CD25+Foxp3+T cells) in total T lymphocytes using fluorescence activated cell sorter (FACS). The level of Th1/Th2 cytokines in the serum including interleukin-2 (IL-2), interleukin-4 (IL-4), interleukin-10 (IL-10) and interferon-γ (IFN-γ) were detected by enzyme-linked immunoabsorbent assay(ELISA).The percentage of regulative T lymphocyte was significantly lower at T1, which increased after chemotherapy but was still lower than that at T0 and that in Group B [(4.15±0.56)%, (5.60±0.88)%, and(5.38±0.92)%, all P0.01]. The levels of IL-4, IL-10 decreased at T1, and increased to the normal level at T2 compared with those at T0 or those in the control group (all P0.01). In contrast, the levels of IL-2 and IFN-γ increased significantly at T1 and decreased to the normal level at T2 during the entire observation period.For patients with advanced colorectal cancer, FOLFOX chemotherapy can decrease the proportion of regulatory T lymphocytes and results in Th1/Th2 cytokine drift to Th1 type. Therefore, FOLFOX may help the release from the anticancer immune inhibition.

In wireless sensor networks, the energy of nodes is limited. so designing efficient routing for reducing energy consumption is important. In this paper we proposed A Low Power Grid-based Cluster Routing Algorithm of Wireless Sensor Networks (LPGCRA). The characteristic of this algorithm is that the WSN is divided into different grids according to information of the node location, and then the nodes are organized within the grid by the clustering way. The clustering head is chose dynamiclly according to energy dissipation of the cluster nodes, then communicates with BS node through a relaying node. This algorithm is useful to reduce node energy consumption and prolonged life of the system, also enhanced the balance of network load. The simulation results shows that this algorithm balanced the energy consumption of the nodes effectively and improved the network lifetime.

microRNAs function as effective molecules in regulation of many biological functions of organisms; in most case they regulate gene expression moderately. Emerging evidence suggests that microRNAs play a key role in the regulation of immunological functions including innate and adaptive immune responses. The research on microRNAs would be helpful in elucidation of the mechanisms of human immune system and in development of potential therapies based on microRNAs.

Aims Innate and adaptive immune responses are associated with the development of hypertension-induced myocardial hypertrophy and fibrosis. As a result, we investigated whether heat shock protein (HSP) 70, which is a molecule of damage-associated molecular patterns, could induce inflammation in the myocardium and promote the development of hypertension-induced cardiac hypertrophy and fibrosis. Methods and results We found that HSP70 serum levels, as well as the amount of HSP70 translocation to the cardiomyocyte membranes and the interstitial space, were elevated in the hypertensive mice caused by abdominal aortic constriction (AAC). Transcriptional inhibition of HSP70 expression by a specific heat shock transcript factor inhibitor, KNK437, reduced the serum level, and the re-distribution of HSP70. It promoted myocardial hypertrophy and cardiac dysfunctions although it protected animals from AAC-induced cardiac fibrosis. On the other hand, the functional antagonism of HSP70 by an anti-HSP70 antibody attenuated AAC-induced cardiac hypertrophy and fibrosis without adverse haemodynamic effects. The cardioprotective effect of the anti-HSP70 antibody was largely attributed to its ability to block AAC-activated immune response in the heart, as was indicated by suppressing the hypertension-enhanced conjugation of HSP70 with toll-like receptor 4, reducing heart-infiltrating macrophages, decreasing the expression of pro-inflammatory factor monocyte chemoattractant protein-1 and profibrotic factor transforming growth factor beta 1, and attenuating pro-hypertrophy signal MAPK P38 and ERK. Conclusion These results indicate that intracellular and extracellular HSP70 have different roles in the regulation of cardiac remodelling and function in response to hypertension. Extracellular HSP70 is a potential therapeutic target against cardiac hypertrophy and fibrosis.

To find out whether there was any change in early infant temperament after mothers had received group psychological therapy on depression and anxiety during pregnancy period.A total of 800 subjects meeting the inclusion criteria, without the exclusion criteria and willing to sign the informed consent were recruited randomly from Shanghai International Peace MaternityChild Health Hospital in their l6th-20th weeks of pregnancy. They were randomized into the therapy group and the control group by the doll randomization table. Women in the therapy group would have a group psychological therapy for 6 times, 1.5 hours each time, while the control group not. The group psychological therapy included therapist introduction and participatory discussion. The therapy concerned the antepartum and postpartum depression, the risk factors concerned with antepartum and postpartum depression, antepartum and postpartum anxiety, psychological defense theory, reflex training and spouse lesson. Mothers reported their babies' infant temperament by filling the early infant temperament questionnaire (EITQ) within 3 months post-delivery.The percent of easy infant temperament type was 87.9% in the psychological therapy group and 81.7% in the hard group. And there was significant difference between two groups (chi2 = 4.530, upsilon=1, P = 0.033). And there were significant differences in the dimensions of approach-withdrawal and threshold of responsiveness in infant temperament between two groups.The maternal antepartum psychological therapy can increase the ratio of easy infant temperament. And it has effects upon the temperament dimensions of approach-withdrawal and threshold of responsiveness.

The syndrome of inappropriate secretion of antidiuretic hormone (SIADH) as a dilutional hyponatremia is due to a pathological increase of antidiuretic hormone (ADH). It is characterized by hyponatremia and decreased serum osmolarity as well as an increase in urinary osmolarity. The most common etiological factors of this syndrome include diseases or trauma of the central nervous system and malignant tumor or inflammation of the lung. SIADH following abdominal surgery is rare.We report the case of a 68-year-old woman who developed, 24 hours after common bile duct exploration and stone removal, continuous hyponatremia for 20 days and clinical manifestations of nausea, vomiting, and lethargy without focal neurological signs.Laboratory examinations supported the diagnosis of SIADH. After therapy with fluid restriction, the patient recovered.There are diverse causes for SIADH. It is important to have kept this clinical possibility in mind in the differential diagnosis of refractory hyponatremia under any circumstances.

To prepare the anticolon carcinoma vaccine with rich chaperone peptide and to examine its anticancer immunological efficacy.CT26 colon carcinoma cells were cultured in 1 mg/L Trichosanthin 1640 medium at different temperatures to induce the chaperone expression and promote the synthesis of antigen peptides. Groups of these cells treated under the different condition were lysed by the sonic disintegration, and the lysates were centrifuged. The rawpurified proteins were obtained from the supernatants by precipitating with saturated ammonium sulfate and removing the molecules below 50,000 and above 300,000 in molecular weight via dialysis. Furthermore, the proteins with the molecular weights in 70,000, 90,000, 95,000, 110,000 and 170,000 were collected through gel filtration and SDS-PAGE. The purified proteins were analysed by Western blotting, and inspected on the anticancer immunological effects including lymphocyte proliferation and the activities of NK and CTL.Major of the chaperone peptides of anticancer effects in CT26 cells, including antigen peptides joining with HSP70, HSP90, gp96, HSP110 and HSP170, was satisfactorily extracted and condensed, and rich chaperone peptide composites were successfully obtained. The composites prepared under various condition could all enhance lymphocyte proliferation and the activities of CTL and NK(P0.01).The rich chaperone peptide composites are successfully prepared via dialysis, salt fractionation and gel filtration combined with SDS-PAGE. Both the heat stress and Trichosanthin can increase the composites, which treated by 42 centi-degree heat stress and Trichosanthin are found to possess the strongest anticancer efficacy.

To develop a novel non-viral gene delivery vector CY11-PEI-beta-CyD and to test its gene transfection efficiency.CY11 (CGMQLPLATWY) was conjugated to polyethylenimine-beta-cyclodextrin to form CY11-PEI-beta-CyD with a cross-linker [N-succinimidy-3-(2-pyridyldithio) propionate, SPDP]. (1)H-NMR and TGA were used to confirm the structure of vector. The DNA condensing ability of CY11-PEI-beta-CyD was investigated by gel retardation assay. Cytotoxicity of CY11-PEI-beta-CyD was determined by MTT assay and transfection efficiency was investigated in COS-7, Hela and B16 cells.CY11 was conjugated onto PEI-beta-CyD successfully, confirmed by(1)H NMR and TGA. The novel vector effectively condensed DNA at N/P ratio of 4îIt showed low cytotoxicity up to the concentration was 160 Mgr;g/ml. The transfection efficiency was 17-fold higher than that of PEI 25 kDa at N/P ratio of 20.The novel vector CY11 -PEI-beta-CyD with low cytotoxic and high transfection efficiency may be used as a potential carrier for gene delivery.

To develop a novel vector for gene delivery with low molecular weight polyethylenimine grafted to the natural polysaccharide and conjugated to folic acid (LNT-PEI-FA).The properties of LNT-PEI-FA were characterized by (1)H-NMR, FT-IR and TGA, respectively. The particle size of LNT-PEI-FA/DNA complex was measured. The DNA binding ability of LNT-PEI-FA was detected by gel electrophoresis retardation assay.The particle size of LNT-PEI-FA/DNA complex was about 200 nm. Gel electrophoresis showed that at N/P ratio of 1.8 (W/W) the polymer was able to completely condense DNA. In vitro experiments showed a high efficiency of gene transfection in A293 and B16 cell lines.A novel non-viral vector LNT-PEI-FA was successfully synthesized and characterized, which may be applied in gene transfection research in the future.

To develop a novel gene delivery vector with poly-aspartamide-glutamic acid and polyethylenimine as the backbone.alpha, beta-poly-(N-2-hydroxypropyl)-D, L-aspartamide-glutamic acid (PHPAG) was synthesized and low molecular weight polyethylenimine (PEI 1.8 kDa) was grafted to form PHPAG-PEI 1800. Chemical and biological characterization of the polymer was identified.The polymer was confirmed by (1)H-NMR, and the molecular weight was about 1.2 x 10(4). The ability of DNA binding was showed by gel retardation assay at N/P ratio of 3. 5. MTT assay showed that the polymer was non toxic in COS-7 and A293 cell lines. In vitro test demonstrated that it had high transfection efficiency in B16 and Hela cell lines.PHPAG-PEI 1800 was successfully synthesized,which might be a potential vector for gene delivery.

To develop a new prodrug of 5-fluorouracil-polyethylenimine-beta-cyclodextrin-floxuridine (PEI-beta-CyD-Fd) and to test its antitumor activity.Floxuridine was conjugated to polyethylenimine-beta-cyclodextrin to form prodrug PEI-beta-CyD-Fd. The structure of synthesized PEI-beta-CyD-Fd was confirmed by (1)H-NMR, FT-IR and UV. MTT assay and cell wound healing assay were performed on human hepatic carcinoma cell line HepG2.The drug loading was 2 %. The MTT assay and cell wound healing assay indicated that PEI-beta-CyD-Fd significantly inhibited proliferation and migration of HepG2 cells.The synthesized prodrug PEI-CyD-Fd has a significant antitumor activity on HepG2 cells.

To develop a novel non-viral gene delivery vector based on polyethylenimine and beta-cyclodextrin targeting to Her-2 receptor (MC10-PEI-beta-CyD).The PEI-beta-CyD was synthesized by low molecular weight polyethylenimine (PEI, Mw 600) cross-linked beta-cyclodextrin (beta-CyD) via N, N-carbonyldiimidazole (CDI). The chemical linker[N-succinimidy-3-(2-pyridyldithio) propionate, SPDP] was used to bind peptide MC10 (MARAKEGGGC) to PEI-beta-CyD to form the vector MC10-PEI-beta-CyD. The (1)H-NMR was used to confirm the structure of vector. The DNA condensing ability,and the particle size of MC10-PEI-beta-CyD/DNA complexes were demonstrated by gel retardation assay and electron microscope observation (TEM). Cell viability was tested by MTT assay. The transfection efficiency was determined on cultured SKOV-3, A549 and MCF-7 cells.MC10 was linked onto PEI-beta-CyD successfully. The vector was able to condense DNA at N/P ratio of 5 and particle size was about (170 +/-35)nm. The vector showed low cytotoxicity and high transfection efficiency in cultured SKOV-3, A549 and MCF-7 cells.A novel non-viral vector MC10-PEI-beta-CyD with low cytotoxicity and high transfection efficiency has been successfully synthesized.

To develop a novel gene delivery vector TAT-PEI-beta-CyD.beta-cyclodextrin (beta-CyD) was linked by low molecular weight (PEI 600) via 1, 1-carbonyldiimidazole (CDI), and TAT peptide (RRRQRRKKRC) was coupled to PEI 600 by [N-succinimidy-3-(2-pyridyldithio) propionate, SPDP]. The copolymer was characterized by (1)H-NMR and FT-IR. Physiochemical characteristics of TAT-PEI-beta-CyD/DNA complexes were tested by agarose gel electrophoresis and particle size measurements. Cell viability and transfection efficiency were evaluated in A293 and B16 cells using PEI 25 kDa as a control.TAT peptide was successfully coupled to PEI-beta-CyD. The result of gel electrophoresis showed that the TAT-PEI-beta-CyD was able to condense DNA efficiently at N/P ratio of 4. The particle size of TAT-PEI-beta-CyD/DNA complexes was around 100 nm. The cytotoxicity of TAT-PEI-beta-CyD was lower than that of PEI 25 kDa. The transfection efficiency of TAT-PEI-beta-CyD was higher than that of PEI 25 kDa in A293 and B16 cells at N/P ratio of 30.The novel vector TAT-PEI-beta-CyD has been developed successfully with low cytotoxicity and high transfection efficiency.

To construct a drug carrier and gene vector PEG-PEI-Pt.Polyethyleneglycol (PEG) was coupled to polyethylenimine (PEI 600) and platinum tetrachloride; PEG-PEI-Pt complex was formed in ethanol. The complex was characterized by XRD, UV-VIS and FT-IR and the DNA condensation was tested by electrophoretic mobility shift assay. The cell viability was evaluated by MTT assay in Hela, B16, A293 and COS-7 cells and in vitro transfection efficiency was measured in A293 and B16 cells.The structure of PEG-PEI-Pt was characterized by XRD, UV-VIS and FT-IR. PEG-PEI-Pt complex was able to bind DNA at N/P weight ratio of 0.4:1; the complex showed cytotoxicity on Hela and B16 cells. The complex had higher transfection efficiency in A293 and B16 cells than PEI 600.A novel drug carrier and gene vector PEG-PEI-Pt was constructed successfully.

Altered peptide ligand (APL), a short peptide with immune regulatory activity and substitutions of a single or multiple amino acids in an antigenic peptide, has shown potential therapeutic effect on autoimmune disease, tumor and virus infection. APL regulates immune responses by interfering the interaction between the major histocompatibility complex (MHC), antigenic peptide and T cell receptor (TCR), or by regulating the intracellular signaling of antigen presenting cells, bystander suppression and inducing heterogenous immune responses. High-specific and high-affinity APL screened from peptide laboratory by phage display, has a potential to be a new resource for drug with antigen specificity.

To construct a novel gene delivery vector using polyethylenimine (PEI) as backbone modified with the peptide CP9 containing Arginine-Glycine-Aspartic acid (RGD) sequence and to verify its physicochemical characters and the gene delivery function.The chemical linker [N-Succinimidyl-3- (2-pyridyldithio) ] propionate (SPDP) was employed to bind CP9 onto PEI to form a novel gene delivery vector CP9-PEI. The (1)H-NMR and FT-IR were used to verify the linkage of CP9. The plasmid DNA condensing ability of CP9-PEI,the shape and the particle size of the polyplexes formed with CP9-PEI-plasmid DNA were demonstrated by gel retardation assay, electron microscope observation and particle size assay,respectively. The enhanced transfection efficiency and the integrin targeting capacity were detected by the transfection experiments in HepG2 cells and free RGD peptide inhibition test.CP9 was linked onto PEI successfully. The new synthesized vector CP9-PEI could efficiently condense plasmid DNA at N/P ratio of 4 and when N/P ratio was equal to 10, the shape of polyplexes formed with CP9-PEI-plasmid DNA was round or round-alike with particle size of about 200 nm. The transfection efficiency of CP9-PEI was nearly 2 times of PEI in HepG2 cells and the free RGD peptide had the inhibition effect on the efficiency of CP9-PEI.The modification of CP9 on PEI can improve the transfection efficiency of PEI and has the integrin targeting ability.

To develop a novel dual-modified vaccine, the superantigen-linked intestine-carcinoma cells expressing membrane-bound heat shock protein 70 (HSP70), and further examine its anticancer therapeutic effect.The pre-established intestine carcinoma CT26 line expressing membrane-bound heat shock protein 70 (HSP70) was amplified and incubated with superantigen fusion protein, staphylococcal enterotoxin A (SEA) fused with transmembrane sequence (SEA-TM), thereby the dual-modified vaccine was prepared after inactivation. The anticancer efficacy of the vaccine was examined.The laser confocal microscopy and flow cytometry showed that there co-existed much HSP70 and SEA on the vaccine membrane surface. Both of the single-modified vaccines, the SEA-linked vaccine and membrane-bound-HSP70-expressing one, displayed marked tumor suppression, a prolonged survival period, augmented lymphocyte proliferation and higher NK and CTL activity in the vaccinated mice when compared with its counterpart. Furthermore, the dually modified vaccine induced lymphocyte proliferation most intensively, generated the highest NK and CTL activity as well as the strongest tumor rejection in the vaccinated mice. The survival period of the mice was further prolonged.A new vaccine, SEA-linked and membrane-bound-HSP70-expressing intestine-carcinoma cells can induce more potent anticancer immunity and produce better therapeutic efficacy.

To construct the recombinant plasmid of RCAS1, to express and purify its fusion protein GST-RCAS1, and to investigate its biological function.RCAS1 encoding gene was amplified by RT-PCR from total RNA extract of MCF-7 cells and was ligated with expression plasmid vector pGEX-2T by T4 DNA ligase after digested by the restricted endonucleases BamH I and EcoR I. Then the ligated products were inserted into competence JM109 E. Coli and the positive recombinants were identified by restriction endonuclease digestion assay and DNA sequencing. The GST-RCAS1 fusion protein expression was induced by IPTG in BL21 E. Coli and was purified with GST column and identified by SDS-PAGE and Western blotting with anti-GST monoclonal antibody, anti-RCAS1 (N-18) and anti-RCAS1 (C-20) polyclonal antibody. The apoptosis of activated T cells induced by GST-RCAS1 fusion protein was detected by flow cytometry with Annexin V and propidium iodide (PI) staining.A 642 bp product was cloned by RT-PCR and the recombinant plasmid was constructed successfully. The GST-RCAS1 fusion protein was recognized by GST monoclonal antibody and RCAS1(N-18 and C-20) polyclonal antibody. FACS analysis showed that GST-RCAS1 fusion protein induced apoptosis in activated T cells.The recombinant plasmid of RCAS1 has been successfully constructed and the GST-RCAS1 fusion protein expressed and purified. The apoptosis inducing effect of GST-RCAS1 fusion protein on activated T cells is demonstrated.

To explore the biological clock of pulp sensibility in young people so as to enrich the theory of pulp-chronobiology and conduce to clinical diagnosis and the treatment of pulposis.40 healthy young volunteers (20 males and 20 females) were examined. Pulp sensibility test was performed using the pulp sensibility tester produced in France. Pulp sensibility reading was obtained at each 4 hours from 8:00 a.m. till next 8:00 a.m., thus there were totally seven time-pints in 24 hours. And the readings were averaged.The pulp sensibility data of every volunteer were analyzed by methods for cosinor-rhythmometry, and significant difference (P0.05) was found. The acrophase and bathyphase appeared at 0:00 and 12:00 separately. We did not find significant difference of the mesor, amplitude and phase between young males and females (P0.05).Circadian rhythm is demonstrated in thepulp sensibility data of young people; the highest pulp sensibility is at 12:00 while the lowest is at 0:00.

To develop a new vaccine expressing membrane-bound heat shock protein 70 (mbHSP70) and further study its antitumor therapeutic effect.The pre- established vector expressing mbHSP70 was transfected into CT26 cells of colorectal cancer. After the CT26 cells were incubated with 900 microg/ml G418, the sub-clones resistant to G418 were harvested and the HSP70 positive clones were selected by limiting dilution. The clones were amplified and inactivated, thereby the vaccine expressing mbHSP70 was prepared. Lymphocyte proliferation stimulated by the vaccines, NK and CTL activity was observed. The antitumor efficacy of vaccine was observed in BALB/c mice model with colorectal cancer.The laser confocal microscopy and flow cytometry showed that there existed much HSP70 on the vaccine membrane surface. The HSP70 gene-modified vaccine displayed augmented lymphocyte proliferation and higher NK and CTL activity in vitro,and marked tumor suppression and prolonged survival time of the vaccinated micein vivo, when compared with its counterpart. Furthermore, mbHSP70-expression vaccine elicited lymphocyte proliferation most intensively, generated the highest NK and CTL activity as well as the strongest antitumor effect, and prolonged survival time of the vaccinated mice.A new vaccine expressing mbHSP70 has more potent antitumor immunity and better therapeutic efficacy than HSP70 gene-modified vaccine did.

To prepare the SEA-TM and mB7.1-GPI dual-anchored EL-4 cell vaccine and to investigate its antitumor effects.mB7.1-GPI-anchored EL-4 cell vaccine, SEA-TM-anchored EL-4 cell vaccine, SEA-TM and mB7.1-GPI dual-anchored EL-4 cell vaccine were prepared. In vitro the biological activities of these vaccines were measured using a lymphocyte proliferation assay and cytokine release assay on splenocytes derived from C57BL/6 mice. The splenocytes were co-cultured with EL-4 or EL-4/mB7.1-GPI or EL-4/SEA-TM or EL-4/SEA-TM + mB7.1-GPI (treated with Mitomycin C). Lymphocyte proliferation was determined with MTT assay, the concentrations of cytokines (IL-2 and IFN-gamma) were measured using a ELISA technique. Forty C57BL/6 mice were inoculated with EL-4 cells, after 3 days the mice were randomly divided into 5 groups with 8 in each and were treated with PBS, EL-4 cell vaccine, EL-4/mB7.1-GPI cell vaccine, EL-4/SEA-TM cell vaccine and EL-4/SEA-TM + mB7.1-GPI cell vaccine respectively, vaccines were injected three time with two-day interval. Animals were observed daily, tumor sizes were measured every third day. Twenty-five days after tumor challenge, 3 mice in each group were sacrificed and splenic lymphocytes were isolated to examine the activity of natural killer cells (NK) and cytolytic T lymphocytes (CTL). The survival of the remaining 5 mice in each group was observed till the 90th day.mB7.1-GPI or/and TM-SEA fusion protein was stably anchored onto the surface of EL-4 tumor cells. EL-4/mB7.1-GPI or EL-4/SEA-TM had a stronger ability to stimulate lymphocyte proliferation and IL-2 and IFN-gamma production than EL-4 (P0.05); while EL-4/SEA-TM + mB7.1-GPI showed a further increased ability than EL-4/mB7.1-GPI and EL-4/SEA-TM in stimulating lymphocyte proliferation and cytokine production in vitro (P0.05). Volume of tumor was smaller and survival time of mice was longer in EL-4/mB7.1-GPI vaccine group, EL-4/SEA-TM vaccine group and EL-4/SEA-TM + mB7.1-GPI vaccine group, comparing with PBS group and EL-4 cell vaccine group (P0.05). Tumor volume was much smaller and survival time of mice was much longer in EL-4/mB7.1-GPI + mB7.1-GPI vaccine group, comparing with EL-4/SEA-TM vaccine group and EL-4/mB7.1-GPI vaccine group (P0.05). Lymphocytes derived from the mice treated with EL-4/SEA-TM + mB7.1-GPI showed much higher NK activity and CTL activity than those derived from EL-4/mB7.1-GPI vaccine group and EL-4/SEA-TM vaccine group (P0.05), meanwhile the NK activity and CTL activity of EL-4/mB7.1-GPI vaccine group and EL-4/SEA-TM vaccine group was higher than EL-4 vaccine group (P0.05).mB7.1-GPI or/and SEA-TM fusion protein was stably anchored onto the surface of EL-4 tumor cells. The tumor cell vaccines prepared from these cells exhibited antitumor effect. The mB7.1-GPI and SEA-TM dual-anchored tumor cell vaccine had much stronger antitumor effect than the single-anchored tumor cell vaccine.

To investigate the influencing factors of polyethylenimine (PEI) in gene transfer in vitro.Cytotoxic effects of PEI on in vitro cultured NIH 3T3 cells were quantified by MTT assay. The interaction between PEI and DNA at different charge ratios was analyzed by agarose gel electrophoresis retardation assay. The expression of gene transfer was monitored in Cos-7 cells using pEGFP and pSV beta plasmids as the reporter gene systems. Influences of chloroquine, albumin, serum, salt ion strength, and Mg(2+) ion and other factors on PEI/DNA transfer efficiency were evaluated.The survival rate of NIH3T3 cells at 6 mg/L of PEI was 64.2% and at 7 mg/L of PEI was 54.4%. Gel electrophoresis retardation assays showed that PEI completely retarded DNA migration at 3.0 PEI nitrogen per DNA phosphate. Chloroquine enhanced the transfection efficiency of PEI. Albumin and serum in the culture medium decreased the transfection efficiency. HBS(HEPES buffered solution) or 150 mmol/L NaCl as the dilution solution of PEI/DNA was superior over 278 mmol/L glucose solution in the transfection efficiency. Mg(2+) in the dilution solution decreased the transfer efficiency of PEI/DNA.PEI is efficient gene transfer agent of eukaryotes in vitro, and can be possibly used in vivo.

To study the local hemorrhagic activity of a fibrinolytic enzyme (FII(a)) from Agkistrodon acutus venom and its mechanism.The local hemorrhagic activity was determined by subcutaneous injection on the back of mouse. The effects of FIIa on factor X, prothrombin, gelatin, and collagen were visualized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Platelet aggregation assays were performed in rat platelet-rich plasma (PRP). Human umbilical vein endothelial cells (HUVEC) were cultured and passaged in complete M199 medium. Cell viability and nuclear morphology change were determined by fluorescein diacetate (FDA) staining and Hoechst 33258 staining, respectively.The minimum hemorrhagic dose (MHD) of FIIa was 89 microg. In vitro, FII(a) (0.25 g/L) degraded factor X, prothrombin, collagen, and gelatin, and dose-dependently (0.25, 0.50, 0.75, and 1.00 g/L) inhibited the platelet aggregation induced by ADP in rat PRP. When HUVEC in culture treated with FII(a), HUVEC showed detachment in a dose-dependent manner, but no apoptosis sign was observed.FIIa had local hemorrhagic activity, and the mechanism was related to the degradation of factor X, prothrombin, gelatin, and collagen, the inhibition of ADP-induced platelet aggregation, and inducement of HUVEC detachment.

OBJECTIVE: To investigate the antiangiogenesis effect of linomide in treatment of transplanted human squamous cell carcinoma of tongue in BALB/C nude mice and to study its relations to regulation on cytokine secretion of macrophages. METHODS: An animal model was established by inoculating the human squamous cell carcinoma cell line Tca8113 into the BALB/c nu/nu nude mice. The mice were randomly divided into three groups and received linomide therapy. The microvessel density (MVD) in the tumor tissue was investigated by immunohistochemistry The productions of TNFalphaand GM-CSF of peritoneal macrophages derived from the tumor -bearing nude mice and cultured murine macrophage cell line RAW264.7 after linomide treatment were detected by ELISA assay. RESULTS: It showed that the tumor weight of mice injected intraperitoneally with 100 mg/kg d-(1), 50 mg/kg d-(1) linomide and mice of control group were 0.47+/-0.25g, 0.92+/-0.30g and 1.75+/-0.38g, respectively. The microvessel density (MVD) in tumor tissue from mice treated with linomide 100 mg/kg d-(1), 50 mg/kg d-(1) decreased 38.2%, 57.8% respectively when compared with that in mice of control group. After linomide treatment, the function of TNFalpha secretion of peritoneal macrophages from tumor-bearing nude mice was significantly inhibi ted when compared with macrophages from untreated mice. And linomide inhibited the release of TNFalpha of RAW264.7 cells in a dose dependent manner. CONCLUSION: It indicats that linomide can effectively inhibit the growth of human squamous carcinoma of tongue in nude mice and decrease the microvessel density in the tumor tissue. The affection of release of antiangiogenic factor TNFalpha of macrophage may be an important mechanism of antitumor activity of linomide.

CaTiO3 incipient ferroelectric ceramics have been modified by introducing BiFeO3. The single-phase (Ca1−yBiy)(Ti1−yFey)O3 solid solutions in space group Pbnm at room temperature are obtained for y ≤ 0.7. An obvious relaxor dielectric peak is induced in the present ceramics, and the peak temperature Tm increases with increasing y, where the dielectric constant as function of temperature can be well fitted with Barrett equation. The monotonic increase of T1 and T0 and decrease of (T1 − 2T0) in Barrett equation with increasing y suggest an approximation to the ferroelectric behavior, where the ferroelectric ordering becomes stronger compared with the quantum fluctuations. That is, the ferroelectric transition can be induced by introducing BiFeO3 in CaTiO3 incipient ferroelectric ceramics.