Activin A secretion by muscle-repairing macrophages induces heterotopic ossification in mice
Wenqiang Yin, Kazuo Okamoto, Asuka Terashima, Warunee Pluemsakunthai, Takehito Ono, Taku Ito-Kureha, Shizuo Akira, Yoshinobu Hashizume, Roland Baron, Satoshi Ueha, Kouji Matsushima, Martin M. Matzuk, Yuji Mishina, Hiroshi Takayanagi
Wenqiang Yin, Kazuo Okamoto, Asuka Terashima, Warunee Pluemsakunthai, Takehito Ono, Taku Ito-Kureha, Shizuo Akira, Yoshinobu Hashizume, Roland Baron, Satoshi Ueha, Kouji Matsushima, Martin M. Matzuk, Yuji Mishina, Hiroshi Takayanagi
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Research Article Bone biology Immunology Muscle biology

Activin A secretion by muscle-repairing macrophages induces heterotopic ossification in mice

Abstract

The immune system is not only essential for host defense, but it is also involved in tissue maintenance and disease pathogenesis. Macrophages play a key role in tissue repair, fibrosis, and tumorigenesis, but the mechanisms underlying their multifunctionality have not been fully explored. Here, we identified Mrep (Ly6ChiCX3CR1loPDPN+CD9+) as a crucial subset of macrophages for muscle regeneration after muscle injury. Muscle regeneration required Mrep-derived activin A, which was produced via the TLR4/TIR domain–containing adapter-inducing interferon-β/TANK-binding kinase 1/interferon regulatory factor 3/7 signaling pathway in response to muscle injury. Mrep exerted pathological effects by secreting activin A in a model of genetically induced heterotopic ossification (HO), which was suppressed by TLR4 inhibition. Thus, this study elucidates the context-dependent functions of macrophages and the link between injury and HO, suggesting that Mrep is a potential therapeutic target for regenerating muscles and suppressing HO.

Authors

Wenqiang Yin, Kazuo Okamoto, Asuka Terashima, Warunee Pluemsakunthai, Takehito Ono, Taku Ito-Kureha, Shizuo Akira, Yoshinobu Hashizume, Roland Baron, Satoshi Ueha, Kouji Matsushima, Martin M. Matzuk, Yuji Mishina, Hiroshi Takayanagi

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Figure 7

Activin A expression in Mrep is induced via the TRIF/TBK1/IRF3/7 signaling pathway.

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Activin A expression in Mrep is induced via the TRIF/TBK1/IRF3/7 signali...
(A and B) RT-qPCR results showing the relative Inhba expression in muscles of indicated mice at 1 dpi (A) and 4 dpi (B). Uninjured (n = 9); 1 dpi: WT (n = 6), Myd88–/– (n = 4), Ticam1–/– (n = 5); 4 dpi: WT (n = 3), Myd88–/– (n = 5), Ticam1–/– (n = 9). (C) RT-qPCR results showing relative Inhba expression in the Mrep cells sorted from WT (n = 9) and Ticam1–/– (n = 6) mice at 1 dpi. (D) RT-qPCR showing relative Inhba mRNA expression in muscles at 1 dpi in WT mice treated with vehicle (n = 7), TBK1 inhibitor (n = 5), or NF-κB inhibitor (n = 4). (E) RT-qPCR results showing relative Inhba mRNA expression in BMDMs derived from WT (n = 5), Irf3–/– (n = 4), or Irf7–/– mice (n = 4) stimulated with LPS for 1 day. (F) ChIP-seq (GEO GSE38377) showing RNA polymerase II (Pol II) and H3K4me3 occupancy in untreated and LPS-stimulated BMDMs at 24 hours. The Inhba gene is shown as a blue line with 3 exons indicated by blue boxes. Upon LPS treatment, a prominent RNA Pol II peak appeared approximately 2 kb upstream of the transcription start site (TSS), indicating the promoter. H3K4me3 enrichment in the same region indicates active chromatin. The promoter contains an interferon-stimulated responsive element (ISRE) motif recognized by IRF3 and IRF7, with its sequence and location shown in the inset. The P values were calculated using unpaired 2-tailed t test (C) and 1-way ANOVA with Tukey’s multiple-comparison test (A, B, D, and E). A P value < 0.05 was considered significant. Data are shown as the mean ± SEM, and symbols represent individual mice (A–E).