332 research outputs found
Expression of Multiple Artificial MicroRNAs from a Chicken miRNA126-Based Lentiviral Vector
Background: The use of RNAi in both basic and translational research often requires expression of multiple siRNAs from the
same vector.
Methods/Principal Findings: We have developed a novel chicken miR126-based artificial miRNA expression system that can
express one, two or three miRNAs from a single cassette in a lentiviral vector. We show that each of the miRNAs expressed
from the same lentiviral vector is capable of potent inhibition of reporter gene expression in transient transfection and
stable integration assays in chicken fibroblast DF-1 cells. Transduction of Vero cells with lentivirus expressing two or three
different anti-influenza miRNAs leads to inhibition of influenza virus production. In addition, the chicken miR126-based
expression system effectively inhibits reporter gene expression in human, monkey, dog and mouse cells. These results
demonstrate that the flanking regions of a single primary miRNA can support processing of three different stem-loops in a
single vector.
Conclusions/Significance: This novel design expands the means to express multiple miRNAs from the same vector for
potent and effective silencing of target genes and influenza virus.National Institutes of Health (U.S.) (Grant R01AI056267)Cobb-Vantress, inc
Hospital mortality among major trauma victims admitted on weekends and evenings: a cohort study
Article deposited according to publisher policy posted on SHERPA/RoMEO, 14/09/2010.YesFunding provided by the Open Access Authors Fund
Membrane diffusion- and capillary blood volume measurements are not useful as screening tools for pulmonary arterial hypertension in systemic sclerosis: a case control study
<p>Abstract</p> <p>Background</p> <p>There is no optimal screening tool for the assessment of pulmonary arterial hypertension (PAH) in patients with systemic sclerosis (SSc). A decreasing transfer factor of the lung for CO (TLCO) is associated with the development of PAH in SSc. TLCO can be partitioned into the diffusion of the alveolar capillary membrane (Dm) and the capillary blood volume (Vc). The use of the partitioned diffusion to detect PAH in SSc is not well established yet. This study evaluates whether Dm and Vc could be candidates for further study of the use for screening for PAH in SSc.</p> <p>Methods</p> <p>Eleven SSc patients with PAH (SScPAH+), 13 SSc patients without PAH (SScPAH-) and 10 healthy control subjects were included. Pulmonary function testing took place at diagnosis of PAH. TLCO was partitioned according to Roughton and Forster. As pulmonary fibrosis in SSc influences values of the (partitioned) TLCO, these were adjusted for fibrosis score as assessed on HRCT.</p> <p>Results</p> <p>TLCO as percentage of predicted (%) was lower in SScPAH+ than in SScPAH- (41 ± 7% <it>vs</it>. 63 ± 12%, p < 0.0001, respectively). Dm% in SScPAH+ was decreased as compared with SScPAH- (22 ± 6% <it>vs</it>. 39 ± 12%, p < 0.0001, respectively), also after adjustment for total fibrosis score (before adjustment: B = 17.5, 95% CI 9.0–25.9, p = < 0.0001; after adjustment: B = 14.3, 95% CI 6.0–21.7, p = 0.008). No difference was found in Vc%. There were no correlations between pulmonary hemodynamic parameters and Dm% in the PAH groups.</p> <p>Conclusion</p> <p>SScPAH+ patients have lower Dm% than SScPAH- patients. There are no correlations between Dm% and hemodynamic parameters of PAH in SScPAH+. These findings do not support further study of the role of partitioning TLCO in the diagnostic work- up for PAH in SSc.</p
Modulation of growth and angiogenic potential of oral squamous carcinoma cells in vitro using salvianolic acid B
<p>Abstract</p> <p>Background</p> <p>Our previous studies showed that Salvianolic acid B (Sal B) inhibited 7,12-dimethylbenz[a]anthracene (DMBA)-induced oral carcinogenesis in hamsters and such anti-cancer effects might be related to the inhibition of angiogenesis. This study was aimed to further investigate the anti-proliferative effect of Sal B on the most common type of oral cancer, oral squamous cell carcinoma (OSCC) and the possible mechanisms of action with respect to angiogenesis inhibition.</p> <p>Methods</p> <p>Two well-characterized oral squamous cell carcinoma cell lines, CAL27 and SCC4, and premalignant leukoplakia cells were treated with different concentrations of Sal B. Cytotoxicity was assessed by MTT assay. cDNA microarray was utilized to evaluate the expression of 96 genes known to be involved in modulating the biological processes of angiogenesis. Real-time reverse transcription-polymerase chain reaction analysis was conducted to confirm the cDNA microarray data.</p> <p>Results</p> <p>Sal B induced growth inhibition in OSCC cell lines but had limited effects on premalignant cells. A total of 17 genes showed a greater than 3-fold change when comparing Sal B treated OSCC cells to the control. Among these genes, HIF-1α, TNFα and MMP9 are specifically inhibited, expression of THBS2 was up-regulated.</p> <p>Conclusions</p> <p>Sal B has inhibitory effect on OSCC cell growth. The antitumor effect can be attributed to anti-angiogenic potential induced by a decreased expression of some key regulator genes of angiogenesis. Sal B may be a promising modality for treating oral squamous cell carcinoma.</p
Protein kinase Cepsilon is important for migration of neuroblastoma cells
<p>Abstract</p> <p>Background</p> <p>Migration is important for the metastatic capacity and thus for the malignancy of cancer cells. There is limited knowledge on regulatory factors that promote the migration of neuroblastoma cells. This study investigates the hypothesis that protein kinase C (PKC) isoforms regulate neuroblastoma cell motility.</p> <p>Methods</p> <p>PKC isoforms were downregulated with siRNA or modulated with activators and inhibitors. Migration was analyzed with scratch and transwell assays. Protein phosphorylation and expression levels were measured with Western blot.</p> <p>Results</p> <p>Stimulation with 12-<it>O</it>-tetradecanoylphorbol-13-acetate (TPA) induced migration of SK-N-BE(2)C neuroblastoma cells. Treatment with the general protein kinase C (PKC) inhibitor GF109203X and the inhibitor of classical isoforms Gö6976 inhibited migration while an inhibitor of PKCβ isoforms did not have an effect. Downregulation of PKCε, but not of PKCα or PKCδ, with siRNA led to a suppression of both basal and TPA-stimulated migration. Experiments using PD98059 and LY294002, inhibitors of the Erk and phosphatidylinositol 3-kinase (PI3K) pathways, respectively, showed that PI3K is not necessary for TPA-induced migration. The Erk pathway might be involved in TPA-induced migration but not in migration driven by PKCε. TPA induced phosphorylation of the PKC substrate myristoylated alanine-rich C kinase substrate (MARCKS) which was suppressed by the PKC inhibitors. Treatment with siRNA oligonucleotides against different PKC isoforms before stimulation with TPA did not influence the phosphorylation of MARCKS.</p> <p>Conclusion</p> <p>PKCε is important for migration of SK-N-BE(2)C neuroblastoma cells. Neither the Erk pathway nor MARCKS are critical downstream targets of PKCε but they may be involved in TPA-mediated migration.</p
Arabidopsis AtVPS15 is essential for pollen development and germination through modulating phosphatidylinositol 3-phosphate formation
Arabidopsis thaliana phosphatidylinositol 3-kinase (AtVPS34) functions in the development and germination of pollen by catalyzing the biosynthesis of phosphatidylinositol 3-phosphate (PI3P). In yeast, Vps15p is required for the membrane targeting and activity of Vps34. The expression of Arabidopsis thaliana VPS15 (AtVPS15), an ortholog of yeast Vps15, is mainly detected in pollen grains and pollen tubes. To determine its role in pollen development and pollen tube growth, we attempted to isolate the T-DNA insertion mutants of AtVPS15; however, homozygous lines of atvps15 were not obtained from the progeny of atvps15/+ heterozygotes. Genetic analysis revealed that the abnormal segregation is due to the failure of transmission of the atvps15 allele through pollen. Most pollen grains from the atvps15/+ genotype are viable, with normal exine structure and nuclei, but some mature pollen grains are characterized with unusual large vacuoles that are not observed in pollen grains from the wild AtVPS15 genotype. The germination ratio of pollen from the atvps15/+ genotype is about half when compared to that from the wild AtVPS15 genotype. When supplied with PI3P, in vitro pollen germination of the atvps15/+ genotype is greatly improved. Presumably, AtVPS15 functions in pollen development and germination by regulating PI3P biosynthesis in Arabidopsis
Implications of Recent Data on Neutrino Mixing and Lepton Flavour Violating Decays for the Zee Model
We study implications of recent data on neutrino mixing from T2K, MINOS,
Double Chooz and from MEG for the Zee model. The simplest
version of this model has been shown to be ruled out by experimental data some
time ago. The general Zee model is still consistent with recent data. We
demonstrate this with a constrained Zee model based on naturalness
consideration. In this constrained model, only inverted mass hierarchy for
neutrino masses is allowed, and must be non-zero in order to have
correct ratio for neutrino mass-squared differences and for mixing in solar and
atmospherical neutrino oscillations. The best fit value of our model for
is from T2K and MINOS data, very close to the central
value obtained by Double Chooz experiment. There are solutions with non-zero CP
violation with the Jarlskog parameter predicted in the range , and respectively for a 1, 2 and 3
ranges of other input parameters. However, without any constraint on the
-parameter above respective ranges become ,
and . We analyse different cases to obtain a branching ratio for
close to the recent MEG bound. We also discuss other
radiative as well as the charged trilepton flavour violating decay modes of the
-lepton.Comment: References added, one extra figure added, typos corrected, few more
related phenomenology discussion added/modified; 25 pages, 10 figure
Effective Rheology of Bubbles Moving in a Capillary Tube
We calculate the average volumetric flux versus pressure drop of bubbles
moving in a single capillary tube with varying diameter, finding a square-root
relation from mapping the flow equations onto that of a driven overdamped
pendulum. The calculation is based on a derivation of the equation of motion of
a bubble train from considering the capillary forces and the entropy production
associated with the viscous flow. We also calculate the configurational
probability of the positions of the bubbles.Comment: 4 pages, 1 figur
Sustainable Sources of Biomass for Bioremediation of Heavy Metals in Waste Water Derived from Coal-Fired Power Generation
Biosorption of heavy metals using dried algal biomass has been extensively described but rarely implemented. We contend this is because available algal biomass is a valuable product with a ready market. Therefore, we considered an alternative and practical approach to algal bioremediation in which algae were cultured directly in the waste water stream. We cultured three species of algae with and without nutrient addition in water that was contaminated with heavy metals from an Ash Dam associated with coal-fired power generation and tested metal uptake and bioremediation potential. All species achieved high concentrations of heavy metals (to 8% dry mass). Two key elements, V and As, reached concentrations in the biomass of 1543 mg.kg−1 DW and 137 mg.kg−1 DW. Growth rates were reduced by more than half in neat Ash Dam water than when nutrients were supplied in excess. Growth rate and bioconcentration were positively correlated for most elements, but some elements (e.g. Cd, Zn) were concentrated more when growth rates were lower, indicating the potential to tailor bioremediation depending on the pollutant. The cosmopolitan nature of the macroalgae studied, and their ability to grow and concentrate a suite of heavy metals from industrial wastes, highlights a clear benefit in the practical application of waste water bioremediation
Disinfection of Ocular Cells and Tissues by Atmospheric-Pressure Cold Plasma
Background: Low temperature plasmas have been proposed in medicine as agents for tissue disinfection and have received increasing attention due to the frequency of bacterial resistance to antibiotics. This study explored whether atmospheric-pressure cold plasma (APCP) generated by a new portable device that ionizes a flow of helium gas can inactivate ocular pathogens without causing significant tissue damage.
Methodology and Principal Findings: We tested the APCP effects on cultured Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus, Candida albicans, Aspergillus fumigatus and Herpes simplex virus-1, ocular cells (conjunctival fibroblasts and keratocytes) and ex-vivo corneas. Exposure to APCP for 0.5 to 5 minutes significantly reduced microbial viability (colony-forming units) but not human cell viability (MTT assay, FACS and Tunel analysis) or the number of HSV-1 plaque-forming units. Increased levels of intracellular reactive oxygen species (ROS) in exposed microorganisms and cells were found using a FACS-activated 2',7'-dichlorofluorescein diacetate probe. Immunoassays demonstrated no induction of thymine dimers in cell cultures and corneal tissues. A transient increased expression of 8-OHdG, genes and proteins related to oxidative stress (OGG1, GPX, NFE2L2) was determined in ocular cells and corneas by HPLC, qRT-PCR and Western blot analysis.
Conclusions: A short application of APCP appears to be an efficient and rapid ocular disinfectant for bacteria and fungi without significant damage on ocular cells and tissues, although the treatment of conjunctival fibroblasts and keratocytes caused a time-restricted generation of intracellular ROS and oxidative stress-related responses
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