Toll-like receptors differentially regulate CC and CXC chemokines in skeletal muscle via NF-kappaB and calcineurin.
Immunologically active molecules such as cytokines and chemokines have been implicated in skeletal muscle weakness during sepsis as well as recovery from muscle injury. In sepsis, Toll-like receptors (TLRs) act as key sentinel molecules of the innate immune system. Here we determined skeletal muscle cell responses of two prototypical CC and CXC chemokine genes (monocyte chemoattractant protein 1 [MCP-1] and KC, respectively), to stimulation with specific TLR ligands. In addition, we examined whether NF-kappaB and calcineurin signaling are involved in these responses. Differentiated myotubes and intact whole muscles expressed TLR2, TLR4, TLR5, and TLR9. Stimulation with ligands for TLR2 (peptidoglycan) or TLR4 (LPS) elicited robust and equivalent levels of MCP-1 and KC mRNA expression, whereas stimulation of TLR5 (by flagellin) required gamma interferon priming to induce similar effects. Although both TLR2 and TLR4 ligands activated the NF-kappaB pathway, NF-kappaB reporter activity was approximately 20-fold greater after TLR4 stimulation than after TLR2 stimulation. Inhibitory effects of NF-kappaB blockade on TLR-mediated chemokine gene expression, by either pharmacological (pyrrolidine dithiocarbamate) or molecular (IKKbeta dominant-negative transfection) methods, were also more pronounced during TLR4 stimulation. In contrast, inhibitory effects on TLR-mediated chemokine expression of calcineurin blockade (by FK506) were greater for TLR2 than for TLR4 stimulation. MCP-1 and KC mRNA levels also demonstrated differential responses to NF-kappaB and calcineurin blockade during stimulation with specific TLR ligands. We conclude that skeletal muscle cells differentially utilize the NF-kappaB and calcineurin pathways in a TLR-specific manner to enable complex regulation of CC and CXC chemokine gene expression.
Boyd
John H
JH
Respiratory Division, Room L411, Royal Victoria Hospital, 687 Pine Avenue West, Montreal, Quebec H3A 1A1, Canada.
Divangahi
Maziar
M
Yahiaoui
Linda
L
Gvozdic
Dusanka
D
Qureshi
Salman
S
Petrof
Basil J
BJ
D000818:Animals; D019703:Calcineurin / Q000378:metabolism*; D065095:Calcineurin Inhibitors; D018932:Chemokine CCL2 / Q000235:genetics; D054360:Chemokine CXCL1; D019742:Chemokines, CC / Q000235:genetics*; D019743:Chemokines, CXC / Q000235:genetics*; D005408:Flagellin / Q000494:pharmacology; D005786:Gene Expression Regulation; D007166:Immunosuppressive Agents / Q000494:pharmacology; D008024:Ligands; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D018482:Muscle, Skeletal / Q000166:cytology / Q000276:immunology*; D016328:NF-kappa B / Q000378:metabolism*; D010457:Peptidoglycan / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis / Q000378:metabolism; D016559:Tacrolimus / Q000494:pharmacology; D051193:Toll-Like Receptors / Q000819:agonists / Q000502:physiology*
D065095:Calcineurin Inhibitors; C491768:Ccl2 protein, mouse; D018932:Chemokine CCL2; D054360:Chemokine CXCL1; D019742:Chemokines, CC; D019743:Chemokines, CXC; C475895:Cxcl1 protein, mouse; D007166:Immunosuppressive Agents; D008024:Ligands; D008070:Lipopolysaccharides; D016328:NF-kappa B; D010457:Peptidoglycan; D012333:RNA, Messenger; D051193:Toll-Like Receptors; D005408:Flagellin; D019703:Calcineurin; D016559:Tacrolimus