Synergism of toll-like receptor 2 (TLR2), TLR4, and TLR6 ligation on the production of tumor necrosis factor (TNF)-alpha in a spontaneous arthritis animal model of interleukin (IL)-1 receptor antagonist-deficient mice.
The aim of this study was to determine whether stimulation of toll-like receptor 2 (TLR2), TLR4, and TLR6 by their specific ligands induces the production of tumor necrosis factor-alpha (TNF-alpha) in fibroblast-like synoviocytes (FLS) from interleukin-1 receptor antagonist (IL-1Ra)-deficient mice. FLS were isolated from synovial tissues from IL-1Ra-deficient mice and stimulated with various ligands of TLRs. The concentrations of TNF-alpha, interleukin (IL)-1beta, and IL-10 in the culture supernatants of spleen cells were measured by ELISA, and mRNA levels were assessed by real-time PCR. The expression of TLR2, TLR4, TLR6, and TNF-alpha in the synovial tissue was quantified by immunohistochemistry. Cytokine production and TLR expression were measured in FLS stimulated in the presence of the TLR2 ligand PAM3, the TLR4 ligand lipopolysaccharide (LPS), and the TLR6 ligand zymosan, with and without blocking antibody to TNF-alpha and IL-1beta. Stimulation of TLR2, TLR4, and TLR6 by their specific ligands increased the production of TNF-alpha in FLS from IL-1Ra-deficient mice. The stimulatory effect of these TLR ligands showed a dose-dependent pattern. The combination of TLR2, TLR4, and TLR6 synergistically increased the production of TNF-alpha, IL-1beta, TLR2, TLR4, and TLR6. Addition of blocking antibodies to TNF-alpha and IL-1beta abrogated the stimulatory effect of the ligands of TLR2, TLR4, and TLR6 on the production of TNF-alpha, IL-1beta, TLR2, TLR4, and TLR6. These data show that TLR2, TLR4, and TLR6 ligation synergistically stimulates the production of TNF-alpha and IL-1beta in IL-1Ra-deficient mice and suggest that TLRs contribute to the perpetuation of spontaneous arthritis in this animal model.
Division of Rheumatology, Department of Internal Medicine, Hallym University Kang-Nam Sacred Heart Hospital, Seoul, Republic of Korea.
Young Ok
YO
Jung
Mi-La
ML
Cho
Seon-Yeong
SY
Lee
Hye-Jwa
HJ
Oh
Jin-Sil
JS
Park
Mi-Kyung
MK
Park
Min-Jung
MJ
Park
Ji-Hyeon
JH
Ju
Sung-Il
SI
Kim
Sung-Hwan
SH
Park
Ho-Youn
HY
Kim
Jun-Ki
JK
Min
D000818:Animals; D001169:Arthritis, Experimental / Q000276:immunology* / Q000378:metabolism / Q000473:pathology; D004195:Disease Models, Animal; D005347:Fibroblasts / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D053590:Interleukin 1 Receptor Antagonist Protein / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D016753:Interleukin-10 / Q000096:biosynthesis / Q000276:immunology; D053583:Interleukin-1beta / Q000096:biosynthesis / Q000276:immunology; D007719:Knee Joint / Q000276:immunology / Q000378:metabolism / Q000473:pathology; D008024:Ligands; D008070:Lipopolysaccharides / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D018345:Mice, Knockout; D051195:Toll-Like Receptor 2 / Q000819:agonists* / Q000276:immunology / Q000378:metabolism; D051197:Toll-Like Receptor 4 / Q000819:agonists* / Q000276:immunology / Q000378:metabolism; D051216:Toll-Like Receptor 6 / Q000819:agonists* / Q000276:immunology / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis* / Q000276:immunology; D015054:Zymosan / Q000494:pharmacology
D053590:Interleukin 1 Receptor Antagonist Protein; D053583:Interleukin-1beta; D008024:Ligands; D008070:Lipopolysaccharides; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051216:Toll-Like Receptor 6; D014409:Tumor Necrosis Factor-alpha; D016753:Interleukin-10; D015054:Zymosan