Self-organizing vascularized subchondral bone organoids from stromal vascular fraction enable functional osteochondral interface regeneration

Abstract

Osteoarthritis (OA) is closely associated with subchondral bone (SCB) degeneration; however, current models fail to adequately mimic its complex microenvironment. Here, we developed a self-organizing subchondral bone organoid (SSBO) by co-culturing stromal vascular fraction (SVF) cells with decellularized cartilage extracellular matrix (CECM). SVF provided cellular heterogeneity, including adipose-derived stem cells (ADSCs), endothelial cells, pericytes, and macrophages, while CECM served as a native scaffold with tissue-specific cues. SSBO exhibited spontaneous spheroid formation, active cellular infiltration, and dynamic matrix remodeling. Compared to ADSC-only controls, SSBO showed enhanced cell viability, vascularization, collagen remodeling, and spatial organization. Immunostaining and qPCR analyses confirmed an endochondral ossification-like process, characterized by the sequential expression of SOX9, COL2A1, RUNX2, COL1A1, and OCN. In vivo implantation into immunodeficient mice demonstrated robust angiogenesis, bone-like tissue formation, and integration with host vasculature. Furthermore, in a mouse osteochondral defect model, SSBO significantly promoted repair, with improved bone volume, trabecular architecture, and cartilage regeneration. Collectively, this study presents a novel strategy for constructing vascularized, immunomodulatory, and osteogenic SCB organoids, offering a promising platform for regenerative medicine and bone-cartilage interface repair.

Keywords

SVF / Organoids / Tissue engineering / Stromal vascular fraction / Subchondral bone

Acknowledgments

None.

Funding

This study was supported by the National Key Research and Development Program of China (Grant No. 2024YFA1108600).

Conflict of interest

The authors declare no competing financial interests.

Author contributions

Conceptualization: Jiazhou Wu, Zexian Liu, Aiyuan Wang,Hongyu Jiang, Yun Bai

Formal analysis: Tao Qian, Jiazhou Wu, Zexian Liu, Junli Wang, Biao Ma, Jialiang You

Investigation: Zexian Liu, Aiyuan Wang, Yanbin Wu, Zhengrui Zhou, Cheng Huang

Methodology: Tao Qian, Yanbin Wu, Junli Wang, Hongyu Jiang, Zhengrui Zhou, Endong Luo

Writing-original draft: Tao Qian, Jiazhou Wu, Cheng Huang, Junming Zhang, Dingkai Wang

Writing-review & editing: Tao Qian, Yazhou Li, Ying He, Jiang Peng

Ethics approval and consent to participate

This study involving human participants was approved by the Ethics Committee of the Chinese PLA General Hospital (Approval No. 2023KY088-KS001). Adipose tissue samples were collected from 9 patients who underwent liposuction at the Fourth Medical Center of the Chinese PLA General Hospital. All participants provided written informed consent prior to inclusion in the study. None of the participants had a history of metabolic or of medications affecting glucose or lipid metabolism. All procedures involving human participants were conducted in accordance with the Declaration of Helsinki and the relevant institutional and national ethical guidelines.

Consent for publication

Written informed consent was obtained from all participants prior to their inclusion in the study. Each participant explicitly agreed to the use of their clinical samples for research purposes. No personally identifiable information or images of human subjects are included in the manuscript.

Availability of data

The data that support the findings of this study are available from the corresponding author upon reasonable request.

References

[1] Bijlsma JW, Berenbaum F, Lafeber FP. Osteoarthritis: an update with relevance for clinical practice. Lancet (London, England). Jun 18 2011; 377(9783):2115-2126. doi: 10.1016/s0140-6736(11)60243-2

[2] GBD 2017 Disease and Injury Incidence and Prevalence Collaborators. Global, regional, and national incidence, prevalence, and years lived with disability for 354 diseases and injuries for 195 countries and territories, 1990-2017: a systematic analysis for the Global Burden of Disease Study 2017. Lancet (London, England). Nov 10 2018;392(10159):1789-1858. doi: 10.1016/s0140-6736(18)32279-7

[3] Reichenbach S, Felson DT, Hincapié CA, et al. Effect of Biomechanical Footwear on Knee Pain in People With Knee Osteoarthritis: The BIOTOK Randomized Clinical Trial. Jama. May 12 2020; 323(18):1802-1812. doi: 10.1001/jama.2020.3565

[4] Martel-Pelletier J, Barr AJ, Cicuttini FM, et al. Osteoarthritis. Nature reviews Disease primers. Oct 13 2016;2:16072. doi: 10.1038/nrdp.2016.72

[5] Grynpas MD, Alpert B, Katz I, Lieberman I, Pritzker KP. Subchondral bone in osteoarthritis. Calcified tissue international. Jul 1991; 49(1):20-26. doi: 10.1007/bf02555898

[6] Burr DB, Gallant MA. Bone remodelling in osteoarthritis. Nature reviews Rheumatology. Nov 2012; 8(11):665-673. doi: 10.1038/nrrheum.2012.130

[7] Suri S, Walsh DA. Osteochondral alterations in osteoarthritis. Bone. Aug 2012; 51(2):204-211. doi: 10.1016/j.bone.2011.10.010

[8] Goldring SR, Goldring MB. Changes in the osteochondral unit during osteoarthritis: structure, function and cartilage-bone crosstalk. Nature reviews Rheumatology. Nov 2016; 12(11):632-644. doi: 10.1038/nrrheum.2016.148

[9] Karsdal MA, Bay-Jensen AC, Lories RJ, et al. The coupling of bone and cartilage turnover in osteoarthritis: opportunities for bone antiresorptives and anabolics as potential treatments? Annals of the rheumatic diseases. Feb 2014; 73(2):336-348. doi: 10.1136/annrheumdis-2013-204111

[10] Zhang H, Wang L, Cui J, et al. Maintaining hypoxia environment of subchondral bone alleviates osteoarthritis progression. Science advances. Apr 5 2023; 9(14):eabo7868. doi: 10.1126/sciadv.abo7868

[11] Chen Z, Bo Q, Wang C, Xu Y, Fei X, Chen R. Single BMSC-derived cartilage organoids for gradient heterogeneous osteochondral regeneration by leveraging native vascular microenvironment. Journal of nanobiotechnology. Apr 29 2025;23:325. doi: 10.1186/s12951-025-03403-0

[12] Lyu X, Wang J, Su J. Intelligent Manufacturing for Osteoarthritis Organoids. Cell proliferation. Jul 2025; 58(7):e70043. doi: 10.1111/cpr.70043

[13] Day JS, Ding M, van der Linden JC, Hvid I, Sumner DR, Weinans H. A decreased subchondral trabecular bone tissue elastic modulus is associated with pre-arthritic cartilage damage. Journal of orthopaedic research: official publication of the Orthopaedic Research Society. Sep 2001; 19(5):914-918. doi: 10.1016/s0736-0266(01)00012-2

[14] Hu W, Chen Y, Dou C, Dong S. Microenvironment in subchondral bone: predominant regulator for the treatment of osteoarthritis. Annals of the rheumatic diseases. Apr 2021; 80(4):413-422. doi: 10.1136/annrheumdis-2020-218089

[15] Hu Y, Chen X, Wang S, Jing Y, Su J. Subchondral bone microenvironment in osteoarthritis and pain. Bone research. Mar 17 2021; 9(1):20. doi: 10.1038/s41413-021-00147-z

[16] Song H, Li X, Zhao Z, et al. Reversal of Osteoporotic Activity by Endothelial Cell-Secreted Bone Targeting and Biocompatible Exosomes. Nano letters. May 8 2019; 19(5):3040-3048. doi: 10.1021/acs.nanolett.9b00287

[17] Castañeda S, Roman-Blas JA, Largo R, Herrero-Beaumont G. Subchondral bone as a key target for osteoarthritis treatment. Biochemical pharmacology. Feb 1 2012; 83(3):315-323. doi: 10.1016/j.bcp.2011.09.018

[18] Li G, Yin J, Gao J, et al. Subchondral bone in osteoarthritis: insight into risk factors and microstructural changes. Arthritis research & therapy. 2013; 15(6):223. doi: 10.1186/ar4405

[19] Henrotin Y, Pesesse L, Sanchez C. Subchondral bone and osteoarthritis: biological and cellular aspects. Osteoporosis international. Dec 2012; 23 (Suppl 8):847-851. doi: 10.1007/s00198-012-2162-z

[20] Kim W, Gwon Y, Park S, Kim H, Kim J. Therapeutic strategies of three-dimensional stem cell spheroids and organoids for tissue repair and regeneration. Bioactive materials. Jan 2023;19:50-74. doi: 10.1016/j.bioactmat.2022.03.039

[21] Hofer M, Lutolf MP. Engineering organoids. Nature reviews Materials. 2021; 6(5):402-420. doi: 10.1038/s41578-021-00279-y

[22] Takebe T, Sekine K, Enomura M, et al. Vascularized and functional human liver from an iPSC-derived organ bud transplant. Nature. Jul 25 2013; 499(7459):481-484. doi: 10.1038/nature12271

[23] Lancaster MA, Renner M, Martin CA, et al. Cerebral organoids model human brain development and microcephaly. Nature. Sep 19 2013; 501(7467):373-379. doi: 10.1038/nature12517

[24] Takasato M, Er PX, Chiu HS, et al. Kidney organoids from human iPS cells contain multiple lineages and model human nephrogenesis. Nature. Oct 22 2015; 526(7574):564-568. doi: 10.1038/nature15695

[25] Zhang C, Jing Y, Wang J, et al. Skeletal organoids. Biomaterials translational. 2024; 5(4):390-410. doi: 10.12336/biomatertransl.2024.04.005

[26] Zakhari JS, Zabonick J, Gettler B, Williams SK. Vasculogenic and angiogenic potential of adipose stromal vascular fraction cell populations in vitro. In vitro cellular & developmental biology Animal. Jan 2018; 54(1):32-40. doi: 10.1007/s11626-017-0213-7

[27] Reid G, Cerino G, Melly L, et al. Harnessing the angiogenic potential of adipose-derived stromal vascular fraction cells with perfusion cell seeding. Stem cell research & therapy. May 1 2025;16:220. doi: 10.1186/s13287-025-04286-6

[28] Moreira HR, Rodrigues DB, Freitas-Ribeiro S, et al. Spongy-like hydrogels prevascularization with the adipose tissue vascular fraction delays cutaneous wound healing by sustaining inflammatory cell influx. Materials today Bio. Dec 15 2022;17:100496. doi: 10.1016/j.mtbio.2022.100496

[29] Liu W, Jiang H, Chen J, et al. High paracrine activity of hADSCs cartilage microtissues inhibits extracellular matrix degradation and promotes cartilage regeneration. Materials today Bio. Feb 2025; 30:101372. doi: 10.1016/j.mtbio.2024.101372

[30] Kim YS, Majid M, Melchiorri AJ, Mikos AG. Applications of decellularized extracellular matrix in bone and cartilage tissue engineering. Bioengineering & translational medicine. Jan 2019; 4(1):83-95. doi: 10.1002/btm2.10110

[31] Morris AH, Stamer DK, Kyriakides TR. The host response to naturally-derived extracellular matrix biomaterials. Seminars in immunology. Feb 2017;29:72-91. doi: 10.1016/j.smim.2017.01.002

[32] Liu C, Pei M, Li Q, Zhang Y. Decellularized extracellular matrix mediates tissue construction and regeneration. Frontiers of medicine. Feb 2022; 16(1):56-82. doi: 10.1007/s11684-021-0900-3

[33] Zhang X, Chen X, Hong H, Hu R, Liu J, Liu C. Decellularized extracellular matrix scaffolds: Recent trends and emerging strategies in tissue engineering. Bioactive materials. Apr 2022;10:15-31. doi: 10.1016/j.bioactmat.2021.09.014

[34] Guo X, Liu B, Zhang Y, et al. Decellularized extracellular matrix for organoid and engineered organ culture. Journal of tissue engineering. Jan-Dec 2024;15:20417314241300386. doi: 10.1177/20417314241300386

[35] Pan J, Zhou X, Li W, Novotny JE, Doty SB, Wang L. In situ measurement of transport between subchondral bone and articular cartilage. Journal of orthopaedic research. Oct 2009; 27(10):1347-1352. doi: 10.1002/jor.20883

[36] Verdugo-Avello F, Wychowaniec JK, Villacis-Aguirre CA, D’Este M, Toledo JR. Bone microphysiological models for biomedical research. Lab on a chip. Feb 25 2025; 25(5):806-836. doi: 10.1039/d4lc00762j

[37] Wang J, Wu Y, Li G, et al. Engineering Large-Scale Self-Mineralizing Bone Organoids with Bone Matrix- Inspired Hydroxyapatite Hybrid Bioinks. Advanced materials (Deerfield Beach, Fla). Jul 2024; 36(30):e2309875. doi: 10.1002/adma.202309875

[38] Zhang X, Jiang W, Wu X, et al. Divide-and-conquer strategy with engineered ossification center organoids for rapid bone healing through developmental cell recruitment. Nature communications. Jul 4 2025;16:6200. doi: 10.1038/s41467-025-61619-y

[39] Bora P, Majumdar AS. Adipose tissue-derived stromal vascular fraction in regenerative medicine: a brief review on biology and translation. Stem cell research & therapy. Jun 15 2017; 8(1):145. doi: 10.1186/s13287-017-0598-y

[40] Rehman J, Traktuev D, Li J, et al. Secretion of angiogenic and antiapoptotic factors by human adipose stromal cells. Circulation. Mar 16 2004; 109(10):1292-1298 doi: 10.1161/01.Cir.0000121425.42966.F1

[41] Wu J, He Y, Qian T, et al. Stromal vascular fraction self-assembles vascularized osteogenic organoids with immunomodulatory functions. Bioactive materials. 2026/03/01/ 2026;57:323-343. doi: 10.1016/j.bioactmat.2025.10.030

[42] Ahmad N, Anker A, Klein S, et al. Autologous Fat Grafting-A Panacea for Scar Tissue Therapy? Cells. Aug 20 2024; 13(16) doi: 10.3390/cells13161384

[43] Airuddin SS, Halim AS, Wan Sulaiman WA, Kadir R, Nasir NAM. Adipose-Derived Stem Cell: “Treat or Trick”. Biomedicines. Nov 5 2021; 9(11) doi: 10.3390/biomedicines9111624

[44] Guan F, Wang R, Yi Z, et al. Tissue macrophages: origin, heterogenity, biological functions, diseases and therapeutic targets. Signal transduction and targeted therapy. Mar 7 2025; 10(1):93. doi: 10.1038/s41392-025-02124-y

[45] Uribe-Querol E, Rosales C. Phagocytosis: Our Current Understanding of a Universal Biological Process. Frontiers in immunology. 2020;11:1066. doi: 10.3389/fimmu.2020.01066

[46] Blanchard L, Girard JP. High endothelial venules (HEVs) in immunity, inflammation and cancer. Angiogenesis. Nov 2021; 24(4):719-753. doi: 10.1007/s10456-021-09792-8

[47] Mohan SP, Priya SP, Tawfig N, et al. The Potential Role of Adipose-Derived Stem Cells in Regeneration of Peripheral Nerves. Neurology international. Feb 6 2025; 17(2) doi: 10.3390/neurolint17020023

[48] Xiong J, Qiang H, Li T, et al. Human adipose-derived stem cells promote seawater-immersed wound healing via proangiogenic effects. Aging. Mar 26 2021; 13(13):17118-17136. doi: 10.18632/aging.202773

[49] Li J, Liu Y, Zhang R, et al. Insights into the role of mesenchymal stem cells in cutaneous medical aesthetics: from basics to clinics. Stem cell research & therapy. Jun 18 2024; 15(1):169. doi: 10.1186/s13287-024-03774-5

[50] Vanderstichele S, Vranckx JJ. Anti-fibrotic effect of adipose-derived stem cells on fibrotic scars. World journal of stem cells. Feb 26 2022; 14(2):200-213. doi: 10.4252/wjsc.v14.i2.200