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Find video protocols related to scientific articles indexed in Pubmed.
Enhanced production of nitric oxide in A549 cells through activation of TRPA1 ion channel by cold stress.
Nitric Oxide
PUBLISHED: 04-19-2014
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The respiratory epithelium is exposed to the external environment, and inhalation of cold air is common during the season of winter. In addition, the lung is a major source of nitric oxide (NO). However, the effect of cold stress on the production of NO is still unclear. In the present work, We measured the change of NO in single cell with DACF-DA and the change in cytosolic Ca(2+) concentration ([Ca(2+)]c) in A549 cell. We observed that cold stress (from 20 °C to 5 °C) induced an increase of NO in A549 cell, which was completely abolished by applying an extracellular Ca(2+) free medium. Further experiments showed that cold-sensing transient receptor potential subfamily member 1 (TRPA1) channel agonist (allyl isothiocyanate, AITC) increased the production of NO and the level of [Ca(2+)]c in A549 cell. Additionally, TRPA1 inhibitor, Ruthenium red (RR) and camphor, significantly blocked the enhanced production of NO and the rise of [Ca(2+)]c induced by AITC or cold stimulation, respectively. Taken together, these data indicated that cold-induced TRPA1 activation was responsible for the enhanced production of NO in A549 cell.
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The Effect of Aerosol Polyethylenimine/Interferon-? Plasmid Complexes on Expression of Inflammatory Cytokines in Mouse Lung.
J Aerosol Med Pulm Drug Deliv
PUBLISHED: 06-22-2013
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Abstract Background: The expression of inflammatory cytokines in lung tissue plays an important role in immune function of the lung. In this study, we tested whether aerosol delivery of the gene of interferon-? (IFN?) could affect inflammatory cytokine expression in mouse lung. Methods: Murine IFN?-expressing plasmids (pcDNA-IFN?) complexed with polyethylenimine (PEI) (PEI/pcDNA-IFN?) were constructed, and their transfection efficiency was assessed in vivo using real-time quantitative RT-PCR and enzyme-linked immunosorbent assay. After aerosol administration of the plasmid complexes and confirmation of the IFN? plasmid location in lung tissue, we measured mRNA levels of the inflammatory cytokines interleukin-1 (IL-1), IL-6, IL-10, tumor necrosis factor-? (TNF-?), and granulocyte-macrophage colony-stimulating factor (GM-CSF) on days 1 to 7 in mouse lung tissues using real-time RT-PCR. Results: IFN? mRNA expression in mouse lung was significantly increased 24?hr after a single aerosol administration of PEI/pcDNA-IFN? and gradually decreased over the next 5 days, whereas the mRNA expressions of IL-1, IL-6, and GM-CSF were markedly decreased, but not those of IL-10 and TNF-?. Conclusions: PEI/IFN? gene therapy delivered by aerosol has immune-regulating potential by suppressing lung cytokine mRNA expression, and therefore may alleviate lung disease.
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Nitric oxide induces apoptosis associated with TRPV1 channel-mediated Ca(2+) entry via S-nitrosylation in osteoblasts.
Eur. J. Pharmacol.
PUBLISHED: 05-02-2013
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The high-level production of nitric oxide (NO) induced by inflammatory cytokines has been shown to play a key role in the pathogenesis of inflammation-mediated osteoporosis. In the present work, we observed that 1mM of the NO donor sodium nitroprusside (SNP) induced an increase of the cytosolic calcium concentration ([Ca(2+)]c) in osteoblasts, which was completely abolished by applying an extracellular Ca(2+)-free buffer. Further experiments showed that the SNP-induced [Ca(2+)]c increase was specifically blocked by potent antagonists of the transient receptor potential vanilloid subtype 1 (TRPV1) channel: capsazepine, ruthenium red, and La(3+) in Ca(2+)-containing buffer. However, nifedipine, an L-type voltage sensitive Ca(2+)-channel blocker, failed to suppress the [Ca(2+)]c elevation caused by SNP. Additionally, 1mM SNP induced osteoblast apoptosis, which was largely inhibited by the blockers of TRPV1, capsazepine and ruthenium red. Interestingly, our data showed that the SNP-induced [Ca(2+)]c increase was significantly inhibited by N-ethylmaleimide, the blocker of S-nitrosylation modification, instead of inhibitors of the NO-cGMP-PKG pathway. Taken together, our data clearly demonstrated that the NO donor SNP resulted in apoptosis associated with TRPV1 channel-mediated Ca(2+) entry via S-nitrosylation in osteoblasts.
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Hypo-osmotic stress enhances the uptake of polyethylenimine/oligonucleotide complexes in A549 cells via Ca(2+) mobilization from intracellular stores.
Oligonucleotides
PUBLISHED: 01-19-2010
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To determine the mechanism of osmolarity involved in polyethylenimine (PEI)/oligonucleotide (ON) complex transfection in cells, we measured the fluorescence intensities of fluorescein isothiocyanate-labeled ONs complexed with PEI and the changes in cytosolic Ca(2+) concentration ([Ca(2+)](c)) in A549 cells, and we found that uptake of PEI/ON complexes was improved in the cells along with a rise of [Ca(2+)](c) in A549 cells challenged by 50% hypotonic medium. Further experiments showed that the enhanced uptake efficiency and the rise in [Ca(2+)](c) in A549 cells were almost completely abolished from cells loaded with the intracellular calcium chelator 1,2-bis(2-aminophenoxy)-N,N,N,N-tetraacetic acid-acetoxymethyl ester. 2-Aminoethoxydiphenyl borate or 8-(N,N-diethylamino) octyl-3,4,5-trimethoxybenzoate, two potent antagonists of inositol 1,4,5-trisphosphate-mediated Ca(2+) release that blunt [Ca(2+)](c) elevation via Ca(2+) release from endoplasmic reticulum, inhibited the enhanced uptake of PEI/ON complexes induced by Ca(2+)-free hypo-osmotic stress. In summary, the results strongly suggest that calcium-dependent transfection is responsible for the uptake of PEI/ON complexes into A549 cells under hypotonic conditions.
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What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

How does it work?

We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.