Selective activation and functional significance of p38α mitogen-activated protein kinase in lipopolysaccharide-stimulated neutrophils
Jerry A. Nick, … , Gary L. Johnson, G. Scott Worthen
Jerry A. Nick, … , Gary L. Johnson, G. Scott Worthen
Published March 15, 1999
Citation Information: J Clin Invest. 1999;103(6):851-858. https://doi.org/10.1172/JCI5257.
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Article

Selective activation and functional significance of p38α mitogen-activated protein kinase in lipopolysaccharide-stimulated neutrophils

Abstract

Activation of leukocytes by proinflammatory stimuli selectively initiates intracellular signal transduction via sequential phosphorylation of kinases. Lipopolysaccharide (LPS) stimulation of human neutrophils is known to result in activation of p38 mitogen-activated protein kinase (MAPk); however, the upstream activator(s) of p38 MAPk is unknown, and consequences of p38 MAPk activation remain largely undefined. We investigated the MAPk kinase (MKK) that activates p38 MAPk in response to LPS, the p38 MAPk isoforms that are activated as part of this pathway, and the functional responses affected by p38 MAPk activation. Although MKK3, MKK4, and MKK6 all activated p38 MAPk in experimental models, only MKK3 was found to activate recombinant p38 MAPk in LPS-treated neutrophils. Of p38 MAPk isoforms studied, only p38α and p38δ were detected in neutrophils. LPS stimulation selectively activated p38α. Specific inhibitors of p38α MAPk blocked LPS-induced adhesion, nuclear factor-kappa B (NF-κB) activation, and synthesis of tumor necrosis factor-α (TNF-α). Inhibition of p38α MAPk resulted in a transient decrease in TNF-α mRNA accumulation but persistent loss of TNF-α synthesis. These findings support a pathway by which LPS stimulation of neutrophils results in activation of MKK3, which in turn activates p38α MAPk, ultimately regulating adhesion, NF-κB activation, enhanced gene expression of TNF-α, and regulation of TNF-α synthesis.

Authors

Jerry A. Nick, Natalie J. Avdi, Scott K. Young, Lisa A. Lehman, Patrick P. McDonald, S. Courtney Frasch, Marcella A. Billstrom, Peter M Henson, Gary L. Johnson, G. Scott Worthen

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Figure 4

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Effect of p38α MAPk inhibition on LPS-induced rapid neutrophil responses...
Effect of p38α MAPk inhibition on LPS-induced rapid neutrophil responses. Neutrophils suspended in SK&F86002 (open bars) or SB203580 (filled bars) over a range of concentrations for 60 min at 37°C were then stimulated with LPS (100 ng/ml) at 37°C. (a) Effect of in vivo inhibition of p38α MAPk on actin assembly. The -fold increase in relative fluorescence index (RFI) of LPS-stimulated neutrophils (5 min) compared with unstimulated cells was plotted for each concentration of SB203580. The plot represents mean activity and SEM for three independent experiments. The relationship between SB203580 concentration and inhibition of actin assembly was not statistically significant (P = 0.55). (b) Effect of in vivo inhibition of p38α MAPk on neutrophil adhesion. The -fold increase in adhesion of LPS-stimulated neutrophils (30 min) compared with unstimulated cells was plotted for each concentration of inhibitor. The plot represents mean values and SEM for six experiments. The relationship between the concentrations of SK&F86002 or SB203580 and inhibition of adhesion is significant (P < 0.0001). (c) Effect of p38α MAPk inhibition on LPS-induced release of TNF-α. The quantity of TNF-α released per 106 neutrophils stimulated with LPS (120 min) was plotted for each concentration of inhibitor. The plot represents mean values and SEM of three consecutive experiments. The relationship between the concentrations of SK&F86002 or SB203580 and inhibition of TNF-α release is significant (P < 0.0001). TNF, tumor necrosis factor.