The biphasic immunoregulation of pyrimidylpiperazine (Y-40138) is IL-10 sensitive and requires NF-kappa B targeting in the alveolar epithelium.
1. Pyrimidylpiperazine (Y-40138), a synthetic derivative of N-[1-(4-([4-(pyrimidin-2-yl)piperazin-1-yl]methyl)phenyl)cyclopropyl] acetamide, is a novel dual regulator of pro- and anti-inflammatory cytokines in vivo. The aim of the present study was to determine the signal transduction mechanisms implicated in vitro. 2. In alveolar epithelial cells, pre-treatment (30 min) with Y-40138 reduced LPS-induced biosynthesis of IL-1 beta, IL-6 and TNF-alpha, an effect paralleled by up-regulating an anti-inflammatory counter-loop mediated through IL-10. 3. This differential regulation of pro- and anti-inflammatory signals was accompanied by an inhibition of the nuclear localization of selective NF-kappa B subunits, particularly NF-kappa B(1) (p50), RelA (p65), the major transactivating member of the Rel family, RelB (p68) and c-Rel (p75). In addition, Y-40138 blockaded, in a dose-dependent manner, the LPS-induced nuclear activation of NF-kappa B. 4. Analysis of the upstream pathway involved in Y-40138-dependent retardation of LPS-induced NF-kappa B translocation/activation revealed the involvement of an I kappa B-alpha sensitive pathway. Pre-treatment with Y-40138 ameliorated LPS-induced degradation of I kappa B-alpha in the cytosolic compartment and retarded its phosphorylation, suggesting the involvement of an upstream kinase. 5. Recombinant IL-10 (0 -- 10 ng ml(-1)) blockaded, in a dose-dependent manner, LPS-induced biosynthesis of IL-1 beta, IL-6 and TNF-alpha. Furthermore, rhIL-10 reduced the DNA binding activity of NF-kappa B. Immunoneutralization of endogenous IL-10 by a polyclonal alpha IL-10 (5 microg ml(-1)) reversed the inhibitory effect of Y-40138 on pro-inflammatory cytokines and partially restored the DNA binding activity of NF-kappa B. 6. These results indicate that Y-40138 mediated dual immunoregulation of pro- and anti-inflammatory cytokines is IL-10 sensitive and mediated through the I kappa B-alpha/NF-kappa B signal transduction pathway.
Haddad
J J
JJ
Oxygen Signalling Group, Centre for Research into Human Development, Tayside Institute of Child Health, Faculty of Medicine, Ninewells Hospital and Medical School, University of Dundee, Dundee, DD1 9SY. j.j.haddad@dundee.ac.uk
Safieh-Garabedian
B
B
Saadé
N E
NE
Land
S C
SC
D000081:Acetamides / Q000276:immunology / Q000494:pharmacology*; D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D016207:Cytokines / Q000096:biosynthesis / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism; D020672:I-kappa B Proteins; D007106:Immune Sera / Q000276:immunology; D020128:Inhibitory Concentration 50; D016753:Interleukin-10 / Q000276:immunology / Q000494:pharmacology*; D008070:Lipopolysaccharides / Q000037:antagonists & inhibitors / Q000494:pharmacology; D008168:Lung / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008954:Models, Biological; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D010879:Piperazines / Q000276:immunology / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D021122:Protein Subunits; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects
D000081:Acetamides; D016207:Cytokines; D004268:DNA-Binding Proteins; D020672:I-kappa B Proteins; D007106:Immune Sera; D008070:Lipopolysaccharides; D016328:NF-kappa B; C000606204:NFKBIA protein, human; C000606206:Nfkbia protein, rat; D010879:Piperazines; D021122:Protein Subunits; C406663:Y 39041; D016753:Interleukin-10; D000072000:NF-KappaB Inhibitor alpha; D004247:DNA