Selective inhibitors of Kv11.1 regulate IL-6 expression by macrophages in response to TLR/IL-1R ligands.
The mechanism by which the platelet-endothelial cell adhesion molecule PECAM-1 regulates leukodiapedesis, vascular endothelial integrity, and proinflammatory cytokine expression in vivo is not known. We recently identified PECAM-1 as a negative regulator of Kv11.1, a specific voltage-gated potassium channel that functioned in human macrophages to reset a resting membrane potential following depolarization. We demonstrate here that dofetilide (DOF), a selective inhibitor of the Kv11.1 current, had a profound inhibitory effect on neutrophil recruitment in mice following TLR/IL-1R-elicited peritonitis or intrascrotal injection of IL-1 Beta, but had no effect on responses seen with TNF alpha. Furthermore, inhibitors of Kv11.1 (DOF, E4031, and astemizole), but not Kv1.3 (margatoxin), suppressed the expression of IL-6 and MCP-1 cytokines by murine resident peritoneal macrophages, while again having no effect on TNF alpha. In contrast, IL-6 expression by peritoneal mesothelial cells was unaffected. Using murine P388 cells, which lack endogenous C/EBP Beta expression and are unresponsive to LPS for the expression of both IL-6 and MCP-1, we observed that DOF inhibited LPS-induced expression of IL-6 mRNA following ectopic expression of wild-type C/EBP Beta, but not a serine-64 point mutant. Finally, DOF inhibited the constitutive activation of cdk2 in murine peritoneal macrophages; cdk2 is known to phosphorylate C/EBP Beta at serine-64. Taken together, our results implicate a potential role for Kv11.1 in regulating cdk2 and C/EBP Beta activity, where robust transactivation of both IL-6 and MCP-1 transcription is known to be dependent on serine-64 of C/EBP Beta. Our data might also explain the altered phenotypes displayed by PECAM-1 knockout mice in several disease models.
Hunter
Cheryl
C
MRC Centre for Inflammation Research, Queen's Medical Research Institute, College of Medicine and Veterinary Medicine, University of Edinburgh. CHunter@regentec.net
Kadakia
Tejas B
TB
Cooper
Dianne
D
Perretti
Mauro
M
Schwartz
Richard C
RC
Brown
Simon B
SB
D000818:Animals; D015153:Blotting, Western; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000378:metabolism; D002478:Cells, Cultured; D000072237:ERG1 Potassium Channel; D051638:Ether-A-Go-Go Potassium Channels / Q000037:antagonists & inhibitors* / Q000378:metabolism; D005260:Female; D015850:Interleukin-6 / Q000235:genetics / Q000378:metabolism*; D008024:Ligands; D008264:Macrophages / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism; D010538:Peritonitis / Q000378:metabolism / Q000517:prevention & control; D010627:Phenethylamines / Q000494:pharmacology*; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000235:genetics / Q000378:metabolism; D026902:Potassium Channel Blockers / Q000494:pharmacology; D017472:Receptors, Interleukin-1 / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology*; D051193:Toll-Like Receptors / Q000378:metabolism
D022782:CCAAT-Enhancer-Binding Protein-beta; D000072237:ERG1 Potassium Channel; D051638:Ether-A-Go-Go Potassium Channels; D015850:Interleukin-6; C000606913:KCNH2 protein, human; D008024:Ligands; D010627:Phenethylamines; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D026902:Potassium Channel Blockers; D017472:Receptors, Interleukin-1; D013449:Sulfonamides; D051193:Toll-Like Receptors; C063533:dofetilide