Ketamine inhibits tumor necrosis factor-alpha and interleukin-6 gene expressions in lipopolysaccharide-stimulated macrophages through suppression of toll-like receptor 4-mediated c-Jun N-terminal kinase phosphorylation and activator protein-1 activation.
Our previous study showed that ketamine, an intravenous anesthetic agent, has anti-inflammatory effects. In this study, we further evaluated the effects of ketamine on the regulation of tumor necrosis factor-alpha (TNF-alpha) and interlukin-6 (IL-6) gene expressions and its possible signal-transducing mechanisms in lipopolysaccharide (LPS)-activated macrophages. Exposure of macrophages to 1, 10, and 100 microM ketamine, 100 ng/ml LPS, or a combination of ketamine and LPS for 1, 6, and 24 h was not cytotoxic to macrophages. A concentration of 1000 microM of ketamine alone or in combined treatment with LPS caused significant cell death. Administration of LPS increased cellular TNF-alpha and IL-6 protein levels in concentration- and time-dependent manners. Meanwhile, treatment with ketamine concentration- and time-dependently alleviated the enhanced effects. LPS induced TNF-alpha and IL-6 mRNA syntheses. Administration of ketamine at a therapeutic concentration (100 microM) significantly inhibited LPS-induced TNF-alpha and IL-6 mRNA expressions. Application of toll-like receptor 4 (TLR4) small interfering (si)RNA into macrophages decreased cellular TLR4 levels. Co-treatment of macrophages with ketamine and TLR4 siRNA decreased the LPS-induced TNF-alpha and IL-6 productions more than alone administration of TLR4 siRNA. LPS stimulated phosphorylation of c-Jun N-terminal kinase and translocation of c-Jun and c-Fos from the cytoplasm to nuclei. However, administration of ketamine significantly decreased LPS-induced activation of c-Jun N-terminal kinase and translocation of c-Jun and c-Fos. LPS increased the binding of nuclear extracts to activator protein-1 consensus DNA oligonucleotides. Administration of ketamine significantly ameliorated LPS-induced DNA binding activity of activator protein-1. Therefore, a clinically relevant concentration of ketamine can inhibit TNF-alpha and IL-6 gene expressions in LPS-activated macrophages. The suppressive mechanisms occur through suppression of TLR4-mediated sequential activations of c-Jun N-terminal kinase and activator protein-1.
Wu
Gone-Jhe
GJ
Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan.
Chen
Ta-Liang
TL
Ueng
Yune-Fang
YF
Chen
Ruei-Ming
RM
D001711:Biotransformation / Q000187:drug effects; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D024202:Electrophoretic Mobility Shift Assay; D018691:Excitatory Amino Acid Antagonists / Q000633:toxicity*; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D015850:Interleukin-6 / Q000096:biosynthesis* / Q000235:genetics; D048031:JNK Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors*; D007649:Ketamine / Q000633:toxicity*; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism*; D010766:Phosphorylation; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects; D051197:Toll-Like Receptor 4 / Q000037:antagonists & inhibitors*; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D015533:Transcriptional Activation / Q000187:drug effects; D014178:Translocation, Genetic / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis* / Q000235:genetics
D018691:Excitatory Amino Acid Antagonists; D015850:Interleukin-6; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D051197:Toll-Like Receptor 4; D018808:Transcription Factor AP-1; D014409:Tumor Necrosis Factor-alpha; D007649:Ketamine; D048031:JNK Mitogen-Activated Protein Kinases