Barbiturates directly inhibit the calmodulin/calcineurin complex: a novel mechanism of inhibition of nuclear factor of activated T cells.
Barbiturates are frequently used for the treatment of intracranial hypertension after brain injury but their application is associated with a profound increase in the infection rate. The mechanism of barbiturate-induced failure of protective immunity is still unknown. We provide evidence that nuclear factor of activated T cells (NFAT), an essential transcription factor in T cell activation, is a target of barbiturate-mediated immunosuppression in human T lymphocytes. Treatment of primary CD3+ lymphocytes with barbiturates inhibited the PMA and ionomycin induced increase in DNA binding of NFAT, whereas the activity of other transcription factors, such as Oct-1, SP-1, or the cAMP response element-binding protein, remained unaffected. Moreover, barbiturates suppressed the expression of a luciferase reporter gene under control of NFAT (stably transfected Jurkat T cells), and of the cytokine genes interleukin-2 and interferon-gamma that contain functional binding motifs for NFAT within their regulatory promotor domains (human peripheral blood CD3+ lymphocytes). Neither GABA receptor-initiated signaling nor direct interactions of barbiturates with nuclear proteins affected the activity of NFAT. In contrast, barbiturates suppressed the calcineurin-dependent dephosphorylation of NFAT in intact T cells and also inhibited the enzymatic activity of calcineurin in a cell-free system, excluding upstream regulation. Thus, our results demonstrate a novel mechanism of direct inhibition of the calcineurin/calmodulin complex that may explain some of the known immunosuppressive effects associated with barbiturate treatment.
Humar
Matjaz
M
Department of Anesthesiology and Critical Care Medicine, University Hospital Freiburg, Freiburg, Germany.
Pischke
Soeren E
SE
Loop
Torsten
T
Hoetzel
Alexander
A
Schmidt
Rene
R
Klaas
Christoph
C
Pahl
Heike L
HL
Geiger
Klaus K
KK
Pannen
Benedikt H J
BH
D001463:Barbiturates / Q000494:pharmacology*; D019703:Calcineurin / Q000378:metabolism; D065095:Calcineurin Inhibitors; D002147:Calmodulin / Q000037:antagonists & inhibitors* / Q000378:metabolism; D016207:Cytokines / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D019169:Jurkat Cells; D008214:Lymphocytes / Q000187:drug effects / Q000378:metabolism; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D014157:Transcription Factors / Q000037:antagonists & inhibitors* / Q000378:metabolism
D001463:Barbiturates; D065095:Calcineurin Inhibitors; D002147:Calmodulin; D016207:Cytokines; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D014157:Transcription Factors; D019703:Calcineurin