301 research outputs found
Fabrication of Vertical Array CNTs/Polyaniline Composite Membranes by Microwave-Assisted In Situ Polymerization
Defects in the Outer Limiting Membrane Are Associated with Rosette Development in the Nrlβ/β Retina
The neural retinal leucine zipper (Nrl) knockout mouse is a widely used model to study cone photoreceptor development, physiology, and molecular biology in the absence of rods. In the Nrlβ/β retina, rods are converted into functional cone-like cells. The Nrlβ/β retina is characterized by large undulations of the outer nuclear layer (ONL) commonly known as rosettes. Here we explore the mechanism of rosette development in the Nrlβ/β retina. We report that rosettes first appear at postnatal day (P)8, and that the structure of nascent rosettes is morphologically distinct from what is seen in the adult retina. The lumen of these nascent rosettes contains a population of aberrant cells protruding into the subretinal space that induce infolding of the ONL. Morphologically adult rosettes do not contain any cell bodies and are first detected at P15. The cells found in nascent rosettes are photoreceptors in origin but lack inner and outer segments. We show that the adherens junctions between photoreceptors and MΓΌller glia which comprise the retinal outer limiting membrane (OLM) are not uniformly formed in the Nrlβ/β retina and thus allow protrusion of a population of developing photoreceptors into the subretinal space where their maturation becomes delayed. These data suggest that the rosettes of the Nrlβ/β retina arise due to defects in the OLM and delayed maturation of a subset of photoreceptors, and that rods may play an important role in the proper formation of the OLM
Front-like entire solutions for monostable reaction-diffusion systems
This paper is concerned with front-like entire solutions for monostable
reactiondiffusion systems with cooperative and non-cooperative nonlinearities.
In the cooperative case, the existence and asymptotic behavior of spatially
independent solutions (SIS) are first proved. Combining a SIS and traveling
fronts with different wave speeds and directions, the existence and various
qualitative properties of entire solutions are then established using
comparison principle. In the non-cooperative case, we introduce two auxiliary
cooperative systems and establish some comparison arguments for the three
systems. The existence of entire solutions is then proved via the traveling
fronts and SIS of the auxiliary systems. Our results are applied to some
biological and epidemiological models. To the best of our knowledge, it is the
first work to study the entire solutions of non-cooperative reaction-diffusion
systems
Reduced cAMP, Akt Activation and p65-c-Rel Dimerization: Mechanisms Involved in the Protective Effects of mGluR3 Agonists in Cultured Astrocytes
In recent decades, astrocytes have emerged as key pieces in the maintenance of normal functioning of the central nervous system. Any impairment in astroglial function can ultimately lead to generalized disturbance in the brain, thus pharmacological targets associated with prevention of astrocyte death are actually promising. Subtype 3 of metabotropic glutamate receptors (mGluR3) is present in astrocytes, its activation exerting neuroprotective roles. In fact, we have previously demonstrated that mGluR3 selective agonists prevent nitric oxide (NO)-induced astrocyte death. However, mechanisms responsible for that cytoprotective property are still subject to study. Although inhibition of adenylyl cyclase by mGluR3 activation was extensively reported, the involvement of reduced cAMP levels in the effects of mGluR3 agonists and the association between cAMP decrease and the downstream pathways activated by mGluR3 remain neglected. Thus, we studied intracellular signaling mediating anti-apoptotic actions of mGluR3 in cultured rat astrocytes exposed to NO. In the present work, we showed that the cytoprotective effect of mGluR3 agonists (LY379268 and LY404039) requires both the reduction of intracellular cAMP levels and activation of Akt, as assessed by MTT and TUNEL techniques. Moreover, dibutyryl-cAMP impairs Akt phosphorylation induced by LY404039, indicating a relationship between mGluR3-reduced cAMP levels and PI3K/Akt pathway activation. We also demonstrated, by co-immunoprecipitation followed by western-blot, that the mGluR3 agonists not only induce per se survival-linked interaction between members of the NF-ΞΊB family p65 and c-Rel, but also impede reduction of levels of p65-c-Rel dimers caused by NO, suggesting a possible anti-apoptotic role for p65-c-Rel. All together, these data suggest that mGluR3 agonists may regulate cAMP/Akt/p65-c-Rel pathway, which would contribute to the protective effect of mGluR3 against NO challenge in astrocytes. Our results widen the knowledge about mechanisms of action of mGluR3, potential targets for the treatment of neurodegenerative disorders where a pathophysiological role for NO has been established
The Mechanism of Antifungal Action of Essential Oil from Dill (Anethum graveolens L.) on Aspergillus flavus
The essential oil extracted from the seeds of dill (Anethum graveolens L.) was demonstrated in this study as a potential source of an eco-friendly antifungal agent. To elucidate the mechanism of the antifungal action further, the effect of the essential oil on the plasma membrane and mitochondria of Aspergillus flavus was investigated. The lesion in the plasma membrane was detected through flow cytometry and further verified through the inhibition of ergosterol synthesis. The essential oil caused morphological changes in the cells of A. flavus and a reduction in the ergosterol quantity. Moreover, mitochondrial membrane potential (MMP), acidification of external medium, and mitochondrial ATPase and dehydrogenase activities were detected. The reactive oxygen species (ROS) accumulation was also examined through fluorometric assay. Exposure to dill oil resulted in an elevation of MMP, and in the suppression of the glucose-induced decrease in external pH at 4 Β΅l/ml. Decreased ATPase and dehydrogenase activities in A. flavus cells were also observed in a dose-dependent manner. The above dysfunctions of the mitochondria caused ROS accumulation in A. flavus. A reduction in cell viability was prevented through the addition of L-cysteine, which indicates that ROS is an important mediator of the antifungal action of dill oil. In summary, the antifungal activity of dill oil results from its ability to disrupt the permeability barrier of the plasma membrane and from the mitochondrial dysfunction-induced ROS accumulation in A. flavus
Intravenously Administered Alphavirus Vector VA7 Eradicates Orthotopic Human Glioma Xenografts in Nude Mice
VA7 is a neurotropic alphavirus vector based on an attenuated strain of Semliki Forest virus. We have previously shown that VA7 exhibits oncolytic activity against human melanoma xenografts in immunodeficient mice. The purpose of this study was to determine if intravenously administered VA7 would be effective against human glioma.In vitro, U87, U251, and A172 human glioma cells were infected and killed by VA7-EGFP. In vivo, antiglioma activity of VA7 was tested in Balb/c nude mice using U87 cells stably expressing firefly luciferase in subcutaneous and orthotopic tumor models. Intravenously administered VA7-EGFP completely eradicated 100% of small and 50% of large subcutaneous U87Fluc tumors. A single intravenous injection of either VA7-EGFP or VA7 expressing Renilla luciferase (VA7-Rluc) into mice bearing orthotopic U87Fluc tumors caused a complete quenching of intracranial firefly bioluminescence and long-term survival in total 16 of 17 animals. In tumor-bearing mice injected with VA7-Rluc, transient intracranial and peripheral Renilla bioluminescence was observed. Virus was well tolerated and no damage to heart, liver, spleen, or brain was observed upon pathological assessment at three and ninety days post injection, despite detectable virus titers in these organs during the earlier time point.VA7 vector is apathogenic and can enter and destroy brain tumors in nude mice when administered systemically. This study warrants further elucidation of the mechanism of tumor destruction and attenuation of the VA7 virus
A Mitochondria-Dependent Pathway Mediates the Apoptosis of GSE-Induced Yeast
Grapefruit seed extract (GSE), which has powerful anti-fungal activity, can induce apoptosis in S. cerevisiae. The yeast cells underwent apoptosis as determined by testing for apoptotic markers of DNA cleavage and typical chromatin condensation by Terminal Deoxynucleotidyl Transferaseβmediated dUTP Nick End Labeling (TUNEL) and 4,6β²-diaminidino-2-phenylindole (DAPI) staining and electron microscopy. The changes of ΞΞ¨mt (mitochondrial transmembrane potential) and ROS (reactive oxygen species) indicated that the mitochondria took part in the apoptotic process. Changes in this process detected by metabonomics and proteomics revealed that the yeast cells tenaciously resisted adversity. Proteins related to redox, cellular structure, membrane, energy and DNA repair were significantly increased. In this study, the relative changes in the levels of proteins and metabolites showed the tenacious resistance of yeast cells. However, GSE induced apoptosis in the yeast cells by destruction of the mitochondrial 60 S ribosomal protein, L14-A, and prevented the conversion of pantothenic acid to coenzyme A (CoA). The relationship between the proteins and metabolites was analyzed by orthogonal projections to latent structures (OPLS). We found that the changes of the metabolites and the protein changes had relevant consistency
GLI1 Confers Profound Phenotypic Changes upon LNCaP Prostate Cancer Cells That Include the Acquisition of a Hormone Independent State
The GLI (GLI1/GLI2) transcription factors have been implicated in the development
and progression of prostate cancer although our understanding of how they
actually contribute to the biology of these common tumours is limited. We
observed that GLI reporter activity was higher in normal (PNT-2) and
tumourigenic (DU145 and PC-3) androgen-independent cells compared to
androgen-dependent LNCaP prostate cancer cells and, accordingly, GLI mRNA levels
were also elevated. Ectopic expression of GLI1 or the constitutively active
ΞNGLI2 mutant induced a distinct cobblestone-like morphology in LNCaP cells
that, regarding the former, correlated with increased GLI2 as well as expression
of the basal/stem-like markers CD44, Ξ²1-integrin, ΞNp63 and BMI1, and
decreased expression of the luminal marker AR (androgen receptor). LNCaP-GLI1
cells were viable in the presence of the AR inhibitor bicalutamide and gene
expression profiling revealed that the transcriptome of LNCaP-GLI1 cells was
significantly closer to DU145 and PC-3 cells than to control LNCaP-pBP (empty
vector) cells, as well as identifying LCN2/NGAL as a highly induced transcript
which is associated with hormone independence in breast and prostate cancer.
Functionally, LNCaP-GLI1 cells displayed greater clonal growth and were more
invasive than control cells but they did not form colonies in soft agar or
prostaspheres in suspension suggesting that they do not possess inherent stem
cell properties. Moreover, targeted suppression of GLI1 or GLI2 with siRNA did
not reverse the transformed phenotype of LNCaP-GLI1 cells nor did double
GLI1/GLI2 knockdowns activate AR expression in DU145 or PC-3 cells. As such,
early targeting of the GLI oncoproteins may hinder progression to a hormone
independent state but a more detailed understanding of the mechanisms that
maintain this phenotype is required to determine if their inhibition will
enhance the efficacy of anti-hormonal therapy through the induction of a luminal
phenotype and increased dependency upon AR function
Multizone Paper Platform for 3D Cell Cultures
In vitro 3D culture is an important model for tissues in
vivo. Cells in different locations of 3D tissues are
physiologically different, because they are exposed to different concentrations
of oxygen, nutrients, and signaling molecules, and to other environmental
factors (temperature, mechanical stress, etc). The majority of high-throughput
assays based on 3D cultures, however, can only detect the
average behavior of cells in the whole 3D construct.
Isolation of cells from specific regions of 3D cultures is possible, but relies
on low-throughput techniques such as tissue sectioning and micromanipulation.
Based on a procedure reported previously (βcells-in-gels-in-paperβ
or CiGiP), this paper describes a simple method for culture of arrays of thin
planar sections of tissues, either alone or stacked to create more complex 3D
tissue structures. This procedure starts with sheets of paper patterned with
hydrophobic regions that form 96 hydrophilic zones. Serial spotting of cells
suspended in extracellular matrix (ECM) gel onto the patterned paper creates an
array of 200 micron-thick slabs of ECM gel (supported mechanically by cellulose
fibers) containing cells. Stacking the sheets with zones aligned on top of one
another assembles 96 3D multilayer constructs. De-stacking the layers of the 3D
culture, by peeling apart the sheets of paper, βsectionsβ all 96
cultures at once. It is, thus, simple to isolate 200-micron-thick
cell-containing slabs from each 3D culture in the 96-zone array. Because the 3D
cultures are assembled from multiple layers, the number of cells plated
initially in each layer determines the spatial distribution of cells in the
stacked 3D cultures. This capability made it possible to compare the growth of
3D tumor models of different spatial composition, and to examine the migration
of cells in these structures
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