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Macrophage-Derived Extracellular DNA Initiates Heterotopic Ossification

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Abstract

Heterotopic ossification (HO) severely affects people's lives; however, its pathological mechanism remains poorly understood. Although extracellular DNA (ecDNA) has been shown to play important roles in pathological calcification, its effects in HO development and progression remain unknown. The in vivo rat Achilles tendon injury model and in vitro collagen I calcification model were used to evaluate the effects of ecDNA in the ectopic calcifications and the main cell types involved in those pathological process. Histology, immunofluorescent staining, reverse transcriptase-polymerase chain reaction analysis and micro-computed tomography were used to identify the distribution of macrophage-derived ecDNA and elucidate their roles in HO. The results showed that the amount of ecDNA and ectopic calcification increased significantly and exhibited a strong correlation in the injured tendons of HO model compared with those of the controls, which was accompanied by a significantly increased number of M2 macrophages in the injured tendon. During in vitro co-culture experiments, M2 macrophages calcified the reconstituted type I collagen and ectopic bone collected from the injured tendons of HO rats, while those effects were inhibited by deoxyribonuclease. More importantly, deoxyribonuclease reversed the pathological calcification in the injured rat tendon HO model. The present study showed that ecDNA from M2 macrophages initiates pathological calcification in HO, and the elimination of ecDNA might be developed into a clinical strategy to prevent ectopic mineralization diseases. The use of deoxyribonuclease for the targeted degradation of ecDNA at affected tissue sites provides a potential solution to treat diseases associated with ectopic mineralization.

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DATA AVAILABILITY

The datasets for this study can be found in the the article/Supplementary Material, and further inquiries can be directed to the corresponding authors.

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ACKNOWLEDGEMENTS

The authors would like to thank all of the participants for their time and effort.

Funding

This work was supported by grants 81870787, 82170978, and 81870805 from the National Nature Science Foundation of China, Distinguished Young Scientists Funds of Shannxi Province (2021JC-34), grant 2020TD-033 from the Shaanxi Key Scientific and Technological Innovation Team.

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Author notes

  1. Xiaoxiao Han, Changhe Gao and Weicheng Lu are contributed equally to this work.

Authors and Affiliations

  1. Department of Stomatology, Tangdu Hospital, The Fourth Military Medical University, Xi’an, Shaanxi, China

    Xiaoxiao Han, Changhe Gao, Weicheng Lu, Jianfei Yan, Haoqing Xu, Zhenxing Guo, Wenpin Qin, Jialu Gao, Weiwei Zhu & Kai Jiao

  2. State Key Laboratory of Oral & Maxillofacial Reconstruction and Regeneration & National Clinical Research Center for Oral Diseases & Shaanxi Key Laboratory of Stomatology, School of Stomatology & Shaanxi Key Laboratory of Stomatology, School of Stomatology, The Fourth Military Medical University, Xi’an, Shaanxi, China

    Xiaoxiao Han, Changhe Gao, Weicheng Lu, Jianfei Yan, Haoqing Xu, Zhenxing Guo, Wenpin Qin, Jialu Gao, Weiwei Zhu, Yutong Fu & Kai Jiao

  3. The College of Life Science, Northwest University, Xi’an, Shaanxi, China

    Xiaoxiao Han, Haoqing Xu, Weiwei Zhu & Yutong Fu

  4. The Third Affiliated Hospital of Xinxiang Medical University, Xinxiang, China

    Changhe Gao

  5. Department of Neurobiology, The Fourth Military Medical University, Xi’an, Shaanxi, China

    Naining Lu

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Contributions

XH and CG contributed equally to the experimental performing, data acquisition and analysis and manuscript drafting. WL and JY contributed to animal experiments. WQ and HX contributed to data analysis and interpretation. JG and NL contributed to immunofluorescence experiments. ZG, WZ and YF contributed to data interpretation. KJ contributed to the study conception and design, data interpretation and manuscript revision. All authors have read and approved the current version of the manuscript.

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Correspondence to Kai Jiao.

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Animal experiments were performed in accordance with National Animal Protection and Use Guidelines and were approved by the Institutional Animal Care and Use Committee of the Fourth Military Medical University (ethics approval number: 20220906).

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Han, X., Gao, C., Lu, W. et al. Macrophage-Derived Extracellular DNA Initiates Heterotopic Ossification. Inflammation 46, 2225–2240 (2023). https://doi.org/10.1007/s10753-023-01873-8

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