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ORIGINAL RESEARCH ARTICLE

Bone organoid–derived extracellular vesicles promote angiogenesis and osteogenesis in vitro

Zhenning Dai1† ,  Ding Li2† ,  Hui Lu3† ,  Suiqing Huang4† ,  Jiarui Liu5 ,  Jingyuan Huang5 ,  Ruoqi Ma5 ,  Wenqiang Zhou6 ,  Weiwei Su7* ,  Wengang Liu1* ,  Shiyu Li5* ,  Guobing Chen5*
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1 Guangdong Provincial Key Laboratory of Research and Development in Traditional Chinese Medicine, Guangdong Provincial Second Hospital of Traditional Chinese Medicine, Guangzhou, Guangdong , China
2 School of Fashion, Guangdong Polytechnic, Foshan, Guangdong , China
3 Institute of Translational Medicine, Shanghai University, Shanghai , China
4 Department of Cardiac Surgery, First Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong , China
5 Department of Immunology, School of Medicine, Jinan University, Guangzhou, Guangdong , China
6 Department of Intensive Care Unit, The First Affiliated Hospital of Jinan University, Guangzhou, Guangdong , China
7 Department of Minimally Invasive Spinal Surgery, The Affiliated Hospital of Putian University, Putian, Fujian , China
†These authors contributed equally to this work.
OR, 026140021 https://doi.org/10.36922/OR026140021
Received: 2 April 2026 | Revised: 21 July 2026 | Accepted: 5 August 2026 | Published online: 18 September 2026
© 2026 by the Author(s). This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution 4.0 International License ( https://creativecommons.org/licenses/by/4.0/ )
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Abstract

Bone organoids (BOs) demonstrate excellent therapeutic effects in bone defect transplantation, but their high cultivation difficulty and cost limit widespread adoption, prompting us to focus on their derivatives. Whether BO-derived extracellular vesicles (BOEVs) can promote bone regeneration and whether they offer greater advantages than stem cell-derived extracellular vesicles (EVs) remains unclear. In this study, we isolated rat bone marrow mesenchymal stem cells (BMSCs) and induced them into BOs. As culture duration increased, BO size gradually enlarged to 203.71 ± 45.30 μm, but the number of spheroids progressively decreased. Messenger RNA (mRNA) sequencing revealed that after seven days of culture, BOs exhibited hypoxic alterations, leading to increased expression of cytokines, including hypoxia-inducible factor-1α (HIF-1α). Subsequently, BOEVs were isolated from BO culture medium at corresponding time points. Both BOEVs cultured for seven days (BOEV-7d) and 14 days (BOEV-14d) were enriched with bone morphogenetic protein 2 (BMP-2), transforming growth factor beta 1 (TGF-β1), HIF-1α, and vascular endothelial growth factor (VEGF). Co-culturing BOEVs with human umbilical vein endothelial cells and hFOB1.19 cells revealed that BOEV-7d significantly promoted tube length and branching while upregulating VEGF and CCN2 mRNA, demonstrating superior effects compared to BOEV-1d and BOEV-14d. Concurrently, BOEV-7d enhanced alizarin red S staining levels in hFOB1.19 cells, while the expression of osteogenesis-related genes Runx2, Col1a1, and Ocn was positively correlated with culture duration. Notably, BOEVs demonstrated significantly greater osteogenic and angiogenic effects than BMSC-derived EVs in 2D culture. This study demonstrates, in vitro, that BOEVs play dual roles in bone and vascular regeneration and have the potential to modulate the vascularization–osteogenesis pathway in bone defects. It offers insights into bone defect treatment and inspires research on organoid-derived EVs.

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Keywords
Bone organoid
Extracellular vesicles
Hypoxia
Angiogenesis
Osteogenic differentiation
Funding
This work was supported by the National Natural Science Foundation of China (82300018 to SL), the Guangdong Basic and Applied Basic Research Foundation (2025A1515012604 to SL), the Fujian Provincial Natural Science Foundation of China (2025J08345 to WS), the Medical Research Foundation of Putian University (2024114 to WS), the National Center for Translational Medicine (Shanghai) SHU Branch of Shanghai University (SUITM-202503 to HL), and the Youth S&T Talent Support Programme of Guangdong Provincial Association for Science and Technology (to SL).
Conflict of interest
The authors declare that there are no conflicts of interest.
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