1. Inhibitor-κB kinase-β regulates LPS-induced TNF-α production in cardiac myocytes through modulation of NF-κB p65 subunit phosphorylation.
- Author
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Hall, Gentzon, Singh, Ishwar S., Hester, Lisa, Hasday, Jeffery D., and Rogers, Terry B.
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TUMOR necrosis factors , *CORONARY disease , *NF-kappa B , *MUSCLE cells , *PHOSPHORYLATION - Abstract
TNF-α is recognized as a significant contributor to myocardial dysfunction. Although several studies suggest that members of the NF-κB family of transcription factors are essential regulators of myocardial TNF-α gene expression, recent developments in our understanding of the modulation of NF-κB activity through posttranslational modification of NF-κB subunits suggest that the present view of NF-κB-dependent cytokine expression in heart is incomplete. Therefore, the goal of the present study was to examine the role of p65 subunit phosphorylation in the regulation of TNF-α production in cultured neonatal ventricular myocytes. Bacterial LPS-induced TNF-α production is accompanied by a 12-fold increase in phosphorylation of p65 at Ser536, a modification associated with enhancement of p65 transactivation potential. Pharmacological inhibition of IKK-β reduced LPS-induced TNF-α production 38-fold, TNF-α mRNA levels 6-fold, and IκB-α phosphorylation 5-fold and degraded IκB-α 2-fold and p65 phosphorylation 6-fold. Overexpression of dominant-negative p65 reduced TNF-α production 3.5-fold, whereas overexpression of dominant-negative IKK-β reduced LPS-induced TNF-α production 2-fold and p65 phosphorylation 2-fold. Overexpression of dominant-negative IKK-α had no effect on p65 phosphorylation or TNF-α production, revealing that IKK-β, not IKK-α, plays a central role in regulation of p65 phosphorylation at Ser536 and TNF-α production in heart. Finally, we demonstrated, using a chromatin immunoprecipitation assay, that LPS stimulates recruitment of Ser536-phosphorylated p65 to the TNF-α gene promoter in cardiac myocytes. Taken together, these data provide compelling evidence for the role of NF-κB signaling in TNF-α gene expression in heart and highlight the importance of this proinflammatory gene-regulatory pathway as a potential therapeutic target in the management of cytokine-induced myocardial dysfunction. [ABSTRACT FROM AUTHOR]
- Published
- 2005
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