Abstract
Heterotopic ossification (HO), the abnormal formation of bone in soft tissues, is a debilitating complication that occurs after severe burns, trauma, and joint replacement surgery. Current diagnostic methods detect HO only after substantial progression, limiting opportunities for early intervention. In this study, we show that circulating mesenchymal progenitor cells (cMPCs), isolated using a microfluidic iChip from patients undergoing hip replacement and a mouse model of traumatic HO, exhibit HO-associated gene expression as early as 6 hours post-injury—41 days before radiographic detection. RNA sequencing of cMPCs enabled development of a liquid biopsy-based HO risk prediction model, achieving up to 90% sensitivity and 100% specificity in human subjects. Furthermore, the model detected significant reductions in HO risk following prophylactic treatment, correlating with decreased HO volume. These findings establish a noninvasive platform for early detection and monitoring of HO and suggest broader utility for diseases involving aberrant mesenchymal cell fate. This approach enables high-throughput screening of at-risk patients and real-time assessment of therapeutic efficacy.
Data availability
Flow cytometry data was deposited to the Flow Repository and is available under Repository ID FR-FCM-Z7CJ. RNAseq data have been deposited to GEO and is available under accession codes GSE267142, GSE267143GSE267144, and GSE316836. The previously published sequencing data [13] used in this work is available via accession number GSE126060. Source data are provided with this paper.
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Acknowledgments
We are grateful to the MGH Nextgen Sequencing, MGH Flow and Mass Cytometry Cores for technical assistance with Flow Cytometry and RNA sequencing. We thank the following orthopedic surgeons at UTSW that contributed to this work: Drs. Robert Weinschenk, Varatharaj Mounasamy, Alexandra Callan, Michael Huo, Garen Collett, Ishvinder Grewal, Antonia Chen. We thank the technical assistance from Drs. Avanish Mishra, Jon Edd, Ezgi Antmen, Qin Tao, and Sweta Munagapati. This work was supported by funding from Shriners Hospital for Children, MGH ECOR, Orphan Disease Center (to N.M.K.). Dr. Levi is grateful for support from the NIH (R61AR078072/R33AR078072 and R01AR078324). We also appreciate support from Surgical Partners in Research (SPAR) at UTSW, including Jarrett Hubbard and Katie Naumann.
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J.H.N. performed animal experiments; M.H. performed iChip enrichment, data analysis, machine learning; S.K. performed bioinformatic analysis; H.K., F.L., and H.S. performed iChip enrichment and RNA-seq experiments; C.P. developed experimental design and performed animal experiments; A.S. performed bioinformatic analysis; E.C.J. developed experimental design and animal experiments; M.E., A.C., and A.F.C developed clinical design and supported patient enrollment; S.N. performed flow cytometry; R.T. designed bioinformatic approach; J.C. performed animal experiments and microCT analysis, H.K. performed animal experiments; C.A. provided mouse model expertise; M.T. contributed methodology; B.L. and N.M.K. conceived the study, directed the study, contributed methodology, and coordinated the collaborative work. N.M.K. and B.L. wrote the paper. All the authors commented on the manuscript.
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Massachusetts General Hospital has filed a patent application for the use of rare cell isolation technology in heterotopic ossification described in this publication (PCT Application No. PCT/US2024/055005). The remaining authors declare no competing interests.
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Nunez, J., Holtz, M., Korlakunta, S. et al. Early detection of aberrant cell fate and repair using circulating progenitor cells in patients with heterotopic ossification. Nat Commun (2026). https://doi.org/10.1038/s41467-026-68857-8
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DOI: https://doi.org/10.1038/s41467-026-68857-8
