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Adenovirus-mediated delivery and expression of a cAMP-dependent protein kinase inhibitor gene to BEAS-2B epithelial cells abolishes the anti-inflammatory effects of rolipram, salbutamol, and prostaglandin E-2: A comparison with H-89
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UNSPECIFIED (2004) Adenovirus-mediated delivery and expression of a cAMP-dependent protein kinase inhibitor gene to BEAS-2B epithelial cells abolishes the anti-inflammatory effects of rolipram, salbutamol, and prostaglandin E-2: A comparison with H-89. JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS, 309 (2). pp. 833-844. doi:10.1124/jpet.103.060020 ISSN 0022-3565.
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Official URL: http://dx.doi.org/10.1124/jpet.103.060020
Abstract
cAMP-elevating drugs are thought to mediate their biological effects by activating the cAMP/cAMP-dependent protein kinase (PKA) cascade. However, this hypothesis is difficult to confirm due to a lack of selective inhibitors. Here, we have probed the role of PKA in mediating inhibitory effects of several cAMP-elevating drugs in BEAS-2B epithelial cells using an adenovirus vector encoding a PKA inhibitor protein (PKIalpha) and have compared it to H-89, a commonly used small molecule PKA inhibitor. Initial studies established efficient gene transfer and confirmed functionality of PKIalpha 48 h after virus infection. All cAMP-elevating drugs tested promoted the phosphorylation of cAMP response element-binding protein (CREB), activated a cAMP response element (CRE)-driven luciferase reporter gene, and suppressed both granulocyte/macrophage colony-stimulating factor (GM-CSF) generation and [H-3] arachidonic acid (AA) release in response to interleukin-1beta and monocyte chemotactic protein (MCP)-1, respectively. These effects were abolished by PKIalpha. In contrast, H-89 behaved unpredictably under the same conditions. Thus, although CREB phosphorylation evoked by a range of cAMP-elevating drugs was abolished by H-89, neither activation of the CRE-dependent luciferase reporter gene construct nor the inhibition of GM-CSF generation was inhibited. Paradoxically, H-89 antagonized MCP-1-induced [H-3] AA release and enhanced the inhibitory effect of submaximal concentrations of rolipram and 8-bromo-cAMP. We suggest that expression of PKIalpha in susceptible cells provides a simple and unambiguous way to assess the role of PKA in cAMP signaling and to probe the mechanism of action of other drugs and cAMP-dependent responses where the participation of PKA is equivocal. Furthermore, these data suggest that H-89 is not a selective inhibitor of PKA and should be avoided.
Item Type: | Journal Article | ||||
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Subjects: | R Medicine > RS Pharmacy and materia medica | ||||
Journal or Publication Title: | JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS | ||||
Publisher: | AMER SOC PHARMACOLOGY EXPERIMENTAL THERAPEUTICS | ||||
ISSN: | 0022-3565 | ||||
Official Date: | 1 May 2004 | ||||
Dates: |
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Volume: | 309 | ||||
Number: | 2 | ||||
Number of Pages: | 12 | ||||
Page Range: | pp. 833-844 | ||||
DOI: | 10.1124/jpet.103.060020 | ||||
Publication Status: | Published |
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