NETs activate the GAS6-AXL-NLRP3 axis in macrophages to drive morphine tolerance
Abstract Background The development of morphine tolerance presents a major clinical challenge in the effective management of severe pain. This study aims to explore the mechanisms underlying morphine tolerance from a novel perspective, with the ultimate goal of uncovering new insights and identifyin...
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BMC
2025-04-01
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| Series: | Cell Communication and Signaling |
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| Online Access: | https://doi.org/10.1186/s12964-025-02181-4 |
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| author | Qingyan Tian Haiyue Guo Mengyao Zhang Kunmao Jiang Fan Hu Yan Xu Li Wan Xiaokai Zhou Yinbing Pan Wentao Liu Chun-Yi Jiang |
| author_facet | Qingyan Tian Haiyue Guo Mengyao Zhang Kunmao Jiang Fan Hu Yan Xu Li Wan Xiaokai Zhou Yinbing Pan Wentao Liu Chun-Yi Jiang |
| author_sort | Qingyan Tian |
| collection | DOAJ |
| description | Abstract Background The development of morphine tolerance presents a major clinical challenge in the effective management of severe pain. This study aims to explore the mechanisms underlying morphine tolerance from a novel perspective, with the ultimate goal of uncovering new insights and identifying promising therapeutic targets for its treatment. Methods C57BL/6J mice were used in the tail-flick test to evaluate morphine tolerance. Neutrophils derived from mouse bone marrow were employed to investigate the mechanisms underlying morphine-induced NETs formation. Bone marrow-derived macrophages (BMDMs) were harvested from the femur and tibia to study the role of NETs-induced inflammation in analgesic tolerance. Proinflammatory cytokines were measured using Western blotting and real-time PCR. The levels of NETs and the TLR7/9-NLRP3-related signaling pathway were assessed through Western blotting, real-time PCR, and ELISA. Confocal laser scanning microscopy was utilized to visualize NETs in the dorsal root ganglion (DRG) and in cells. Results Our experiments demonstrated that the levels of NETs in the plasma of patients using morphine for analgesia, as well as in morphine-tolerant animals, were significantly elevated. Genetic elimination of Pad4, neutrophil depletion, and treatment with DNase 1 and RNase A to disrupt NETs formation all effectively alleviated morphine tolerance. These findings indicate that NETs play a critical role in the development of morphine tolerance. Mechanistically, we discovered that morphine-induced NETs can be engulfed by macrophages through the GAS6-AXL axis, which subsequently triggers the activation of the TLR7/TLR9-mediated NLRP3 inflammasome, leading to significantly increased levels of IL-1β and IL-18, and ultimately contributing to tolerance. Deletion of Axl, Gas6, or Nlrp3 each significantly improved morphine tolerance. Furthermore, in the murine model, treatment with the IL-1 receptor antagonist anakinra and the IL-18 decoy receptor IL-18BP prevented the development of morphine tolerance. Conclusions This study identifies morphine-induced NETs as a key contributor to morphine tolerance, with the GAS6-AXL-TLR7/9 axis emerging as a potential therapeutic target. Strategies focused on disrupting NETs and modulating this axis may offer a promising approach to combat morphine tolerance. |
| format | Article |
| id | doaj-art-a625debfeddb439c8e5ea21d8b329ed5 |
| institution | DOAJ |
| issn | 1478-811X |
| language | English |
| publishDate | 2025-04-01 |
| publisher | BMC |
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| series | Cell Communication and Signaling |
| spelling | doaj-art-a625debfeddb439c8e5ea21d8b329ed52025-08-20T03:06:55ZengBMCCell Communication and Signaling1478-811X2025-04-0123111810.1186/s12964-025-02181-4NETs activate the GAS6-AXL-NLRP3 axis in macrophages to drive morphine toleranceQingyan Tian0Haiyue Guo1Mengyao Zhang2Kunmao Jiang3Fan Hu4Yan Xu5Li Wan6Xiaokai Zhou7Yinbing Pan8Wentao Liu9Chun-Yi Jiang10Jiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical UniversityJiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical UniversityJiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical UniversityJiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical UniversityJiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical UniversityDepartment of Pain, The First People’s Hospital of Changzhou, Soochow UniversityJiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical UniversityDepartment of Anesthesiology, The First Affiliated Hospital of Nanjing Medical UniversityDepartment of Anesthesiology, The First Affiliated Hospital of Nanjing Medical UniversityJiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical UniversityJiangsu Key Laboratory of Neurodegeneration, Department of Pharmacology, Nanjing Medical UniversityAbstract Background The development of morphine tolerance presents a major clinical challenge in the effective management of severe pain. This study aims to explore the mechanisms underlying morphine tolerance from a novel perspective, with the ultimate goal of uncovering new insights and identifying promising therapeutic targets for its treatment. Methods C57BL/6J mice were used in the tail-flick test to evaluate morphine tolerance. Neutrophils derived from mouse bone marrow were employed to investigate the mechanisms underlying morphine-induced NETs formation. Bone marrow-derived macrophages (BMDMs) were harvested from the femur and tibia to study the role of NETs-induced inflammation in analgesic tolerance. Proinflammatory cytokines were measured using Western blotting and real-time PCR. The levels of NETs and the TLR7/9-NLRP3-related signaling pathway were assessed through Western blotting, real-time PCR, and ELISA. Confocal laser scanning microscopy was utilized to visualize NETs in the dorsal root ganglion (DRG) and in cells. Results Our experiments demonstrated that the levels of NETs in the plasma of patients using morphine for analgesia, as well as in morphine-tolerant animals, were significantly elevated. Genetic elimination of Pad4, neutrophil depletion, and treatment with DNase 1 and RNase A to disrupt NETs formation all effectively alleviated morphine tolerance. These findings indicate that NETs play a critical role in the development of morphine tolerance. Mechanistically, we discovered that morphine-induced NETs can be engulfed by macrophages through the GAS6-AXL axis, which subsequently triggers the activation of the TLR7/TLR9-mediated NLRP3 inflammasome, leading to significantly increased levels of IL-1β and IL-18, and ultimately contributing to tolerance. Deletion of Axl, Gas6, or Nlrp3 each significantly improved morphine tolerance. Furthermore, in the murine model, treatment with the IL-1 receptor antagonist anakinra and the IL-18 decoy receptor IL-18BP prevented the development of morphine tolerance. Conclusions This study identifies morphine-induced NETs as a key contributor to morphine tolerance, with the GAS6-AXL-TLR7/9 axis emerging as a potential therapeutic target. Strategies focused on disrupting NETs and modulating this axis may offer a promising approach to combat morphine tolerance.https://doi.org/10.1186/s12964-025-02181-4Morphine toleranceNETsGAS6AXLNLRP3 |
| spellingShingle | Qingyan Tian Haiyue Guo Mengyao Zhang Kunmao Jiang Fan Hu Yan Xu Li Wan Xiaokai Zhou Yinbing Pan Wentao Liu Chun-Yi Jiang NETs activate the GAS6-AXL-NLRP3 axis in macrophages to drive morphine tolerance Cell Communication and Signaling Morphine tolerance NETs GAS6 AXL NLRP3 |
| title | NETs activate the GAS6-AXL-NLRP3 axis in macrophages to drive morphine tolerance |
| title_full | NETs activate the GAS6-AXL-NLRP3 axis in macrophages to drive morphine tolerance |
| title_fullStr | NETs activate the GAS6-AXL-NLRP3 axis in macrophages to drive morphine tolerance |
| title_full_unstemmed | NETs activate the GAS6-AXL-NLRP3 axis in macrophages to drive morphine tolerance |
| title_short | NETs activate the GAS6-AXL-NLRP3 axis in macrophages to drive morphine tolerance |
| title_sort | nets activate the gas6 axl nlrp3 axis in macrophages to drive morphine tolerance |
| topic | Morphine tolerance NETs GAS6 AXL NLRP3 |
| url | https://doi.org/10.1186/s12964-025-02181-4 |
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