Toll-like receptor 3 signaling converts tumor-supporting myeloid cells to tumoricidal effectors - PubMed (original) (raw)

Toll-like receptor 3 signaling converts tumor-supporting myeloid cells to tumoricidal effectors

Hiroaki Shime et al. Proc Natl Acad Sci U S A. 2012.

Abstract

Smoldering inflammation often increases the risk of progression for malignant tumors and simultaneously matures myeloid dendritic cells (mDCs) for cell-mediated immunity. PolyI:C, a dsRNA analog, is reported to induce inflammation and potent antitumor immune responses via the Toll-like receptor 3/Toll-IL-1 receptor domain-containing adaptor molecule 1 (TICAM-1) and melanoma differentiation-associated protein 5/IFN-β promoter stimulator 1 (IPS-1) pathways in mDCs to drive activation of natural killer cells and cytotoxic T lymphocytes. Here, we found that i.p. or s.c. injection of polyI:C to Lewis lung carcinoma tumor-implant mice resulted in tumor regression by converting tumor-supporting macrophages (Mfs) to tumor suppressors. F4/80(+)/Gr1(-) Mfs infiltrating the tumor respond to polyI:C to rapidly produce inflammatory cytokines and thereafter accelerate M1 polarization. TNF-α was increased within 1 h in both tumor and serum upon polyI:C injection into tumor-bearing mice, followed by tumor hemorrhagic necrosis and growth suppression. These tumor responses were abolished in TNF-α(-/-) mice. Furthermore, F4/80(+) Mfs in tumors extracted from polyI:C-injected mice sustained Lewis lung carcinoma cytotoxic activity, and this activity was partly abrogated by anti-TNF-α Ab. Genes for supporting M1 polarization were subsequently up-regulated in the tumor-infiltrating Mfs. These responses were completely abrogated in TICAM-1(-/-) mice, and unaffected in myeloid differentiation factor 88(-/-) and IPS-1(-/-) mice. Thus, the TICAM-1 pathway is not only important to mature mDCs for cross-priming and natural killer cell activation in the induction of tumor immunity, but also critically engaged in tumor suppression by converting tumor-supporting Mfs to those with tumoricidal properties.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.

Fig. 1.

Antitumor activity of polyI:C against 3LL tumor cells is mediated by the TICAM-1 pathway in vivo. (A) Representative photographs of 3LL tumors excised from WT, TNF-α−/−, TICAM-1−/−, and IPS-1−/− mice. Whole tumor (Upper) and bisected tumor (Lower) are shown. (B–D) On day 0, 3LL tumor cells (3 × 106) were s.c. implanted into B6 WT (B–D), TNF-α−/− (B), TICAM-1−/− (C), and IPS-1−/− (D) mice. PolyI:C i.p. injection was started on the day indicated by arrow, then repeated every 4 d. Data are shown as tumor average size ± SE; n = 3–4 mice per group. *P < 0.05; **P < 0.001. N.S., not significant. A representative experiment of two with similar outcomes is shown.

Fig. 2.

Fig. 2.

TNF-α production in tumor and serum of polyI:C-injected 3LL tumor-bearing mice. Mice bearing 3LL tumor were i.p. injected with 200 μg polyI:C. Tumor (A) and serum (B) were collected at 0, 1, 2, and 3 h after polyI:C injection, and TNF-α concentration was determined by ELISA. TNF-α level in tumor is presented as [TNF-α protein (pg)/tumor weight (g)]. (C) Tumors were isolated from polyI:C-injected tumor-bearing WT, TICAM-1−/−, and IPS-1−/− mice, and TNF-α mRNA was measured by quantitative PCR; n = 3. Data are shown as average ± SD. A representative experiment of two with similar outcomes is shown.

Fig. 3.

Fig. 3.

F4/80+ cells are responsible for the polyI:C-induced elevation of TNF-α production in tumor. Mice bearing 3LL tumors were i.p. injected with 200 μg polyI:C. TNF-α–producing cells in tumors of polyI:C- or PBS-injected mice were examined by immunohistochemical staining and flow cytometry to determine intracellular cytokine expression profiles of CD45+ cells (A), F4/80+ cells (B), and Gr1+ cells (C). CD45+ cells in tumor were gated and are shown in B and C. A representative experiment of two with similar outcomes is shown. TNF-α+ gating squares are shown in red (positive) and green (negative).

Fig. 4.

Fig. 4.

PolyI:C enhances TNF-α production and cytotoxicity of F4/80+ cells in tumor. PolyI:C (200 μg) or PBS was i.p. injected into 3LL tumor-bearing WT mice. After 30 min, F4/80+ cells isolated from tumor were cultured for 24 h and TNF-α concentration in the conditioned medium was determined by ELISA (A). In parallel, the cytotoxicity of tumor-infiltrating F4/80+ cells against 3LL tumor cells was measured by 51Cr-release assay (B). Anti–TNF-α neutralization antibody or control antibody was added (10 μg/mL) to mixed culture of isolated tumor-infiltrating F4/80+ cells and 3LL tumor cells (C). (D) Cytotoxic activity of TNF-α against 3LL tumor cells. Recombinant TNF-α was added to 51Cr-labeled 3LL tumor cell culture at various concentrations. After 20 h, cytotoxicity was measured; n = 3. Data are shown as average ± SD. *P < 0.05, **P < 0.001. A representative experiment of three with similar outcomes is shown.

Fig. 5.

Fig. 5.

PolyI:C induces M1 polarization of TAMs. F4/80+ cells were isolated from 3LL tumor and stimulated with polyI:C (50 μg/mL) for 4 h. Total RNA was extracted and used to analyze the transcript expression levels of M1 (A) and M2 (B and C) markers; n = 3. Data are shown as average ± SD. A representative experiment of two with similar outcomes is shown.

Comment in

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