Received for publication May 14, 2004.
Revised July 16, 2004.
Accepted for publication July 19, 2004.

Inhibition of AP-1 by barbiturates is mediated by differential effects on MAP-kinases and the small G proteins Ras and Rac-1

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-B (NF-B) and nuclear factor of activated T cells (NFAT). 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-analogues inhibit AP-1 dependent gene expression and AP-1 complex formation at clinically relevant doses. Furthermore, MAP-kinase activity, which transcriptionally and posttranslationally regulates AP-1 complex formation is suppressed by most barbiturates. CD3/CD28 or PMA/ionomycin induced p38 and ERK1/2 phosphorylation or JNK1/2 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.

Key words: ERK1/2, G proteins, JNK, activator protein 1, p38, thiopental

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