TLR4 signaling in effector CD4+ T cells regulates TCR activation and experimental colitis in mice
José M. González-Navajas, … , Jongdae Lee, Eyal Raz
José M. González-Navajas, … , Jongdae Lee, Eyal Raz
Published January 4, 2010
Citation Information: J Clin Invest. 2010;120(2):570-581. https://doi.org/10.1172/JCI40055.
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TLR4 signaling in effector CD4+ T cells regulates TCR activation and experimental colitis in mice

Abstract

TLRs sense various microbial products. Their function has been best characterized in DCs and macrophages, where they act as important mediators of innate immunity. TLR4 is also expressed on CD4+ T cells, but its physiological function on these cells remains unknown. Here, we have shown that TLR4 triggering on CD4+ T cells affects their phenotype and their ability to provoke intestinal inflammation. In a model of spontaneous colitis, Il10–/–Tlr4–/– mice displayed accelerated development of disease, with signs of overt colitis as early as 8 weeks of age, when compared with Il10–/– and Il10–/–Tlr9–/– mice, which did not develop colitis by 8 months. Similar results were obtained in a second model of colitis in which transfer of naive Il10–/–Tlr4–/– CD4+ T cells into Rag1–/– recipients sufficient for both IL-10 and TLR4 induced more aggressive colitis than the transfer of naive Il10–/– CD4+ T cells. Mechanistically, LPS stimulation of TLR4-bearing CD4+ T cells inhibited ERK1/2 activation upon subsequent TCR stimulation via the induction of MAPK phosphatase 3 (MKP-3). Our data therefore reveal a tonic inhibitory role for TLR4 signaling on subsequent TCR-dependent CD4+ T cell responses.

Authors

José M. González-Navajas, Sean Fine, Jason Law, Sandip K. Datta, Kim P. Nguyen, Mandy Yu, Maripat Corr, Kyoko Katakura, Lars Eckman, Jongdae Lee, Eyal Raz

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Figure 6

TLR4 modulates TCR-dependent MAPK phosphatase activation.

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TLR4 modulates TCR-dependent MAPK phosphatase activation. (A) Immunoblot...
(A) Immunoblot analysis of the indicated phosphatases after LPS stimulation of CD4+ cells at different time points. (B) Expression of MKPs and p-SHIP1 in freshly isolated, unstimulated CD4+ cells from Il10–/– and Il10–/–Tlr4–/– mice. (C) Immunoblotting of protein extracts from CD4+ T cells stimulated with LPS or left untreated before stimulation with anti-CD3/28 Abs for different periods of time. (D) Analysis of the siRNA knockdown in CD4+ T cells 24 hours after transfection with either MKP-3 or control siRNA (Ctrl). (E) TCR-dependent phosphorylation of ERK1/2 24 hours after transfection with MKP-3 or control siRNA. Numbers within histograms denote percentage of p-ERK1/2–positive cells in stimulated cells when compared with control cells. (F) Cytokine levels measured from 24-hour supernatants of cells treated as in E. Data represent mean ± SEM. (G) p-ERK expression in freshly isolated CD4+ cells from Il10–/– mice treated with MKP inhibitor in vivo (NSC 95397) or vehicle. (H) Cytokine levels after 24 hours of anti-CD3/28 Ab stimulation of MLN-derived CD4+ T cells isolated from Il10–/– mice 3 days after injection of NSC 95397 or vehicle. Data represent mean ± SEM. Data are representative of at least 2 independent experiments. *P < 0.05. All experiments were carried out with FACS-sorted MLN-derived CD4+ cells from Il10–/– or Il10–/–Tlr4–/– mice (purity, >98%). Stimulation was carried out with 100 ng/ml LPS in all experiments. The lanes shown in B were run on the same gel but were noncontiguous.