論文 - 詳細
| RRC ID | 63509 |
|---|---|
| 著者 | Hiraku Y, Guo F, Ma N, Yamada T, Wang S, Kawanishi S, Murata M. |
| タイトル | Multi-walled carbon nanotube induces nitrative DNA damage in human lung epithelial cells via HMGB1-RAGE interaction and Toll-like receptor 9 activation. |
| ジャーナル | Part Fibre Toxicol |
| Abstract |
BACKGROUND:Carbon nanotube (CNT) is used for various industrial purposes, but exhibits carcinogenic effects in experimental animals. Chronic inflammation in the respiratory system may participate in CNT-induced carcinogenesis. 8-Nitroguanine (8-nitroG) is a mutagenic DNA lesion formed during inflammation. We have previously reported that multi-walled CNT (MWCNT) induced 8-nitroG formation in lung epithelial cells and this process involved endocytosis. To clarify the mechanism of CNT-induced carcinogenesis, we examined the role of Toll-like receptor (TLR) 9, which resides in endosomes and lysosomes, in 8-nitroG formation in human lung epithelial cell lines. METHODS:We performed immunocytochemistry to examine 8-nitroG formation in A549 and HBEpC cells treated with MWCNT with a length of 1-2 μm (CNT-S) or 5-15 μm (CNT-L) and a diameter of 20-40 nm. We examined inhibitory effects of endocytosis inhibitors, small interfering RNA (siRNA) for TLR9, and antibodies against high-mobility group box-1 (HMGB1) and receptor for advanced glycation end-products (RAGE) on 8-nitroG formation. The release of HMGB1 and double-stranded DNA (dsDNA) into the culture supernatant from MWCNT-treated cells was examined by ELISA and fluorometric analysis, respectively. The association of these molecules was examined by double immunofluorescent staining and co-immunoprecipitation. RESULTS:CNT-L significantly increased 8-nitroG formation at 0.05 μg/ml in A549 cells and its intensity reached a maximum at 1 μg/ml. CNT-L tended to induce stronger cytotoxicity and 8-nitroG formation than CNT-S. Endocytosis inhibitors, TLR9 siRNA and antibodies against HMGB1 and RAGE largely reduced MWCNT-induced 8-nitroG formation. MWCNT increased the release of HMGB1 and dsDNA from A549 cells into culture supernatant. The culture supernatant of MWCNT-exposed cells induced 8-nitroG formation in fresh A549 cells. Double immunofluorescent staining and co-immunoprecipitation showed that TLR9 was associated with HMGB1 and RAGE in lysosomes of MWCNT-treated cells. CONCLUSIONS:MWCNT induces injury or necrosis of lung epithelial cells, which release HMGB1 and DNA into the extracellular space. The HMGB1-DNA complex binds to RAGE on neighboring cells and then CpG DNA is recognized by TLR9 in lysosomes, leading to generation of nitric oxide and 8-nitroG formation. This is the first study demonstrating that TLR9 and related molecules participate in MWCNT-induced genotoxicity and may contribute to carcinogenesis. |
| 巻・号 | 13 |
| ページ | 16 |
| 公開日 | 2016-3-29 |
| DOI | 10.1186/s12989-016-0127-7 |
| PII | 10.1186/s12989-016-0127-7 |
| PMID | 27026438 |
| PMC | PMC4812657 |
| MeSH | Acute Lung Injury / chemically induced* Acute Lung Injury / genetics Acute Lung Injury / metabolism Acute Lung Injury / pathology Antibodies / pharmacology Cell Line, Tumor CpG Islands DNA Breaks, Double-Stranded* Dose-Response Relationship, Drug Endocytosis / drug effects Epithelial Cells / drug effects* Epithelial Cells / metabolism Epithelial Cells / pathology Guanine / analogs & derivatives Guanine / metabolism HMGB1 Protein / antagonists & inhibitors HMGB1 Protein / immunology HMGB1 Protein / metabolism* Humans Lung / drug effects* Lung / metabolism Lung / pathology Lysosomes / drug effects Lysosomes / metabolism Nanotubes, Carbon / toxicity* Necrosis Nitric Oxide / metabolism Particle Size RNA Interference Reactive Nitrogen Species / metabolism* Receptor for Advanced Glycation End Products / antagonists & inhibitors Receptor for Advanced Glycation End Products / immunology Receptor for Advanced Glycation End Products / metabolism* Risk Assessment Signal Transduction / drug effects Time Factors Toll-Like Receptor 9 / genetics Toll-Like Receptor 9 / metabolism* Transfection |
| IF | 7.546 |
| リソース情報 | |
| ヒト・動物細胞 | A549(RCB0098) |