Osteogenic CpG Oligodeoxynucleotide, iSN40, Inhibits Osteoclastogenesis in a TLR9-Dependent Manner
A CpG oligodeoxynucleotide (CpG-ODN), iSN40, was originally identified as promoting the mineralization and differentiation of osteoblasts, independent of Toll-like receptor 9 (TLR9). Since CpG ODNs are often recognized by TLR9 and inhibit osteoclastogenesis, this study investigated the TLR9 dependen...
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2024-11-01
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| author | Rena Ikeda Chihaya Kimura Yuma Nihashi Koji Umezawa Takeshi Shimosato Tomohide Takaya |
| author_facet | Rena Ikeda Chihaya Kimura Yuma Nihashi Koji Umezawa Takeshi Shimosato Tomohide Takaya |
| author_sort | Rena Ikeda |
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| description | A CpG oligodeoxynucleotide (CpG-ODN), iSN40, was originally identified as promoting the mineralization and differentiation of osteoblasts, independent of Toll-like receptor 9 (TLR9). Since CpG ODNs are often recognized by TLR9 and inhibit osteoclastogenesis, this study investigated the TLR9 dependence and anti-osteoclastogenic effect of iSN40 to validate its potential as an osteoporosis drug. The murine monocyte/macrophage cell line RAW264.7 was treated with the receptor activator of nuclear factor-κB ligand (RANKL) to induce osteoclast differentiation, then the effect of iSN40 on was quantified by tartrate-resistant acid phosphatase (TRAP) staining and real-time RT-PCR. iSN40 completely inhibited RANKL-induced differentiation into TRAP<sup>+</sup> multinucleated osteoclasts by suppressing osteoclastogenic genes and inducing anti-/non-osteoclastogenic genes. Treatment with a TLR9 inhibitor, E6446, or a mutation in the CpG motif of iSN40 abolished the intracellular uptake and anti-osteoclastogenic effect of iSN40. These results demonstrate that iSN40 is subcellularly internalized and is recognized by TLR9 via its CpG motif, modulates RANKL-dependent osteoclastogenic gene expression, and ultimately inhibits osteoclastogenesis. Finally, iSN40 was confirmed to inhibit the osteoclastogenesis of RAW264.7 cells cocultured with the murine osteoblast cell line MC3T3-E1, presenting a model of bone remodeling. This study demonstrates that iSN40, which exerts both pro-osteogenic and anti-osteoclastogenic effects, may be a promising nucleic acid drug for osteoporosis. |
| format | Article |
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| issn | 2075-1729 |
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| publishDate | 2024-11-01 |
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| spelling | doaj-art-7fb2e5acfc8646979396bf789c3adf642025-08-20T02:53:44ZengMDPI AGLife2075-17292024-11-011412157210.3390/life14121572Osteogenic CpG Oligodeoxynucleotide, iSN40, Inhibits Osteoclastogenesis in a TLR9-Dependent MannerRena Ikeda0Chihaya Kimura1Yuma Nihashi2Koji Umezawa3Takeshi Shimosato4Tomohide Takaya5Department of Agriculture, Graduate School of Science and Technology, Shinshu University, 8304 Minami-minowa, Kami-ina, Nagano 399-4598, JapanDepartment of Agriculture, Graduate School of Science and Technology, Shinshu University, 8304 Minami-minowa, Kami-ina, Nagano 399-4598, JapanDepartment of Science and Technology, Graduate School of Medicine, Science and Technology, Shinshu University, 8304 Minami-minowa, Kami-ina, Nagano 399-4598, JapanDepartment of Science and Technology, Graduate School of Medicine, Science and Technology, Shinshu University, 8304 Minami-minowa, Kami-ina, Nagano 399-4598, JapanDepartment of Agriculture, Graduate School of Science and Technology, Shinshu University, 8304 Minami-minowa, Kami-ina, Nagano 399-4598, JapanDepartment of Agriculture, Graduate School of Science and Technology, Shinshu University, 8304 Minami-minowa, Kami-ina, Nagano 399-4598, JapanA CpG oligodeoxynucleotide (CpG-ODN), iSN40, was originally identified as promoting the mineralization and differentiation of osteoblasts, independent of Toll-like receptor 9 (TLR9). Since CpG ODNs are often recognized by TLR9 and inhibit osteoclastogenesis, this study investigated the TLR9 dependence and anti-osteoclastogenic effect of iSN40 to validate its potential as an osteoporosis drug. The murine monocyte/macrophage cell line RAW264.7 was treated with the receptor activator of nuclear factor-κB ligand (RANKL) to induce osteoclast differentiation, then the effect of iSN40 on was quantified by tartrate-resistant acid phosphatase (TRAP) staining and real-time RT-PCR. iSN40 completely inhibited RANKL-induced differentiation into TRAP<sup>+</sup> multinucleated osteoclasts by suppressing osteoclastogenic genes and inducing anti-/non-osteoclastogenic genes. Treatment with a TLR9 inhibitor, E6446, or a mutation in the CpG motif of iSN40 abolished the intracellular uptake and anti-osteoclastogenic effect of iSN40. These results demonstrate that iSN40 is subcellularly internalized and is recognized by TLR9 via its CpG motif, modulates RANKL-dependent osteoclastogenic gene expression, and ultimately inhibits osteoclastogenesis. Finally, iSN40 was confirmed to inhibit the osteoclastogenesis of RAW264.7 cells cocultured with the murine osteoblast cell line MC3T3-E1, presenting a model of bone remodeling. This study demonstrates that iSN40, which exerts both pro-osteogenic and anti-osteoclastogenic effects, may be a promising nucleic acid drug for osteoporosis.https://www.mdpi.com/2075-1729/14/12/1572CpG oligodeoxynucleotide (CpG-ODN)osteoclastosteoclastogenesisosteogenetic oligodeoxynucleotide (osteoDN)Toll-like receptor 9 (TLR9) |
| spellingShingle | Rena Ikeda Chihaya Kimura Yuma Nihashi Koji Umezawa Takeshi Shimosato Tomohide Takaya Osteogenic CpG Oligodeoxynucleotide, iSN40, Inhibits Osteoclastogenesis in a TLR9-Dependent Manner Life CpG oligodeoxynucleotide (CpG-ODN) osteoclast osteoclastogenesis osteogenetic oligodeoxynucleotide (osteoDN) Toll-like receptor 9 (TLR9) |
| title | Osteogenic CpG Oligodeoxynucleotide, iSN40, Inhibits Osteoclastogenesis in a TLR9-Dependent Manner |
| title_full | Osteogenic CpG Oligodeoxynucleotide, iSN40, Inhibits Osteoclastogenesis in a TLR9-Dependent Manner |
| title_fullStr | Osteogenic CpG Oligodeoxynucleotide, iSN40, Inhibits Osteoclastogenesis in a TLR9-Dependent Manner |
| title_full_unstemmed | Osteogenic CpG Oligodeoxynucleotide, iSN40, Inhibits Osteoclastogenesis in a TLR9-Dependent Manner |
| title_short | Osteogenic CpG Oligodeoxynucleotide, iSN40, Inhibits Osteoclastogenesis in a TLR9-Dependent Manner |
| title_sort | osteogenic cpg oligodeoxynucleotide isn40 inhibits osteoclastogenesis in a tlr9 dependent manner |
| topic | CpG oligodeoxynucleotide (CpG-ODN) osteoclast osteoclastogenesis osteogenetic oligodeoxynucleotide (osteoDN) Toll-like receptor 9 (TLR9) |
| url | https://www.mdpi.com/2075-1729/14/12/1572 |
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