Abstract
Barbiturates are known to suppress protective immunity, and their therapeutic use is associated with nosocomial infections. Although barbiturates inhibit T cell proliferation, differentiation, and cytokine synthesis, only thiobarbiturates markedly reduce the activation of immune regulatory transcription factors such as nuclear factor-κB and nuclear factor of activated T cells. In this study, we investigated barbiturate-mediated effects on the regulation of the transcription factor activator protein 1 (AP-1) in primary T lymphocytes. We show that both thiobarbiturates and their oxy-analogs inhibit AP-1-dependent gene expression and AP-1 complex formation at clinically relevant doses. Furthermore, mitogen-activated protein (MAP) kinase activity, which transcriptionally and posttranslationally regulates AP-1 complex formation, is suppressed by most barbiturates. CD3/CD28- or phorbol 12-myristate 13-acetate (PMA)/ionomycin-induced p38 and extracellular signal-regulated kinase 1/2 phosphorylation or c-jun NH2-terminal kinase (JNK) 1/2 kinase activity was significantly diminished by pentobarbital, thiamylal, secobarbital, or methohexital treatment. These barbiturates also inhibited the initiators of the MAP kinase cascade, the small G proteins ras and rac-1, and prevented binding to their partners raf-1 and PAK, respectively. Thiopental, unlike the other barbiturates, only reduced ras and JNK activity upon direct CD3/CD28 receptor engagement. Contrarily, upon PMA/ionomycin stimulation, thiopental blocked AP-1-dependent gene expression independently of the small G protein ras and MAP kinases, thus suggesting an additional, unknown mechanism of AP-1 regulation. In conclusion, our results contribute to the explanation of a clinically manifested immune suppression in barbiturate-treated patients and support the idea of a MAP kinase-independent regulation of AP-1 by PKC and calcium in human T cells.
Footnotes
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This study was supported by departmental funding and by grants from the Else Kroener-Fresenius-Stiftung and the Deutsche Forschungsgemeinschaft (Bonn, Heidelberg) to B.H.J.P. (Heisenberg-Stipends DFG PA 533/3-1 and 3-2).
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.104.071332.
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ABBREVIATIONS: MAP kinase, mitogen-activated protein kinase; AP-1, activator protein 1; ERK, extracellular signal-regulated kinase; JNK, c-jun NH2-terminal kinase; NFAT, nuclear factor of activated T cells; NF-κB, nuclear factor-κB; PMA, phorbol 12-myristate 13-acetate; EMSA, electrophoretic mobility shift assay; PMSF, phenylmethylsulfonyl fluoride; RBD, Rho-binding domain of mouse Rhotekin; GST-PAK-CD, Cdc4/Rac interactive binding domain, corresponding to the Rac and Cdc 42-binding domain of human PAK1B; GTPγS, guanosine 5′-3-O-(thio)triphosphate.
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↵1 These authors contributed equally to this work.
- Received May 13, 2004.
- Accepted July 19, 2004.
- The American Society for Pharmacology and Experimental Therapeutics
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