Human skeletal development and regeneration are shaped by functional diversity of stem cells across skeletal sites

Thomas H Ambrosi¹, Sahar Taheri², Kun Chen³, Rahul Sinha⁴, Yuting Wang⁵, Ethan J Hunt³, L Henry Goodnough⁶, Matthew P Murphy⁷, Holly M Steininger⁸, Malachia Y Hoover⁸, Franco Felix⁴, Kelly C Weldon³, Lauren S Koepke⁸, Jan Sokol⁸, Daniel Dan Liu⁴, Liming Zhao⁹, Stephanie D Conley⁴, Wan-Jin Lu⁴, Maurizio Morri¹⁰, Norma F Neff¹⁰, Noelle L Van Rysselberghe⁶, Erika E Wheeler¹¹, Yongheng Wang¹², J Kent Leach¹¹, Augustine Saiz³, Aijun Wang¹³, George P Yang¹⁴, Stuart Goodman⁶, Julius A Bishop⁶, Michael J Gardner⁶, Derrick C Wan¹⁵, Irving L Weissman¹⁶, Michael T Longaker¹⁷, Debashis Sahoo¹⁸, Charles K F Chan¹⁹

Affiliations

¹ Department of Orthopaedic Surgery, UC Davis Health, Sacramento, CA 95817, USA; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA. Electronic address: thambrosi@ucdavis.edu.
² Department of Computer Science and Engineering, Jacob's School of Engineering, University of California, San Diego, La Jolla, CA 92093, USA.
³ Department of Orthopaedic Surgery, UC Davis Health, Sacramento, CA 95817, USA.
⁴ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA.
⁵ Department of Orthopaedic Surgery, UC Davis Health, Sacramento, CA 95817, USA; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Orthopaedic Surgery, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
⁶ Department of Orthopaedic Surgery, Stanford Hospital and Clinics, Stanford, CA 94063, USA.
⁷ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA; Blond McIndoe Laboratories, Division of Cell Matrix Biology and Regenerative Medicine, School of Biological Sciences, Faculty of Biology, Medicine and Health, The University of Manchester, Manchester Academic Health Science Centre, Manchester, UK.
⁸ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA.
⁹ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Orthopaedic Surgery, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
¹⁰ Chan Zuckerberg BioHub, San Francisco, CA 94158, USA.
¹¹ Department of Orthopaedic Surgery, UC Davis Health, Sacramento, CA 95817, USA; Department of Biomedical Engineering, University of California, Davis, Davis, CA 95616, USA.
¹² Department of Biomedical Engineering, University of California, Davis, Davis, CA 95616, USA.
¹³ Department of Biomedical Engineering, University of California, Davis, Davis, CA 95616, USA; Department of Surgery, University of California Davis Health, Sacramento, CA 95817, USA; Institute for Pediatric Regenerative Medicine, Shriners Hospitals for Children, Sacramento, CA 95817, USA.
¹⁴ Department of Surgery, University of Alabama at Birmingham, Birmingham, AL 35233, USA.
¹⁵ Hagey Laboratory for Pediatric Regenerative Medicine, Stanford University School of Medicine, Stanford University, Stanford, CA 94305, USA.
¹⁶ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Ludwig Center for Cancer Stem Cell Biology and Medicine at Stanford University, Stanford, CA 94305, USA.
¹⁷ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA; Hagey Laboratory for Pediatric Regenerative Medicine, Stanford University School of Medicine, Stanford University, Stanford, CA 94305, USA. Electronic address: longaker@stanford.edu.
¹⁸ Department of Computer Science and Engineering, Jacob's School of Engineering, University of California, San Diego, La Jolla, CA 92093, USA; Department of Pediatrics, University of California, San Diego, La Jolla, CA 92093, USA. Electronic address: dsahoo@ucsd.edu.
¹⁹ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA; Hagey Laboratory for Pediatric Regenerative Medicine, Stanford University School of Medicine, Stanford University, Stanford, CA 94305, USA.

PMID: 40118065
PMCID: PMC12048286 (available on 2026-05-01)
DOI: 10.1016/j.stem.2025.02.013

Human skeletal development and regeneration are shaped by functional diversity of stem cells across skeletal sites

Thomas H Ambrosi et al. Cell Stem Cell. 2025.

. 2025 May 1;32(5):811-823.e11.

doi: 10.1016/j.stem.2025.02.013. Epub 2025 Mar 20.

Authors

Affiliations

¹ Department of Orthopaedic Surgery, UC Davis Health, Sacramento, CA 95817, USA; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA. Electronic address: thambrosi@ucdavis.edu.
² Department of Computer Science and Engineering, Jacob's School of Engineering, University of California, San Diego, La Jolla, CA 92093, USA.
³ Department of Orthopaedic Surgery, UC Davis Health, Sacramento, CA 95817, USA.
⁴ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA.
⁵ Department of Orthopaedic Surgery, UC Davis Health, Sacramento, CA 95817, USA; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Orthopaedic Surgery, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
⁶ Department of Orthopaedic Surgery, Stanford Hospital and Clinics, Stanford, CA 94063, USA.
⁷ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA; Blond McIndoe Laboratories, Division of Cell Matrix Biology and Regenerative Medicine, School of Biological Sciences, Faculty of Biology, Medicine and Health, The University of Manchester, Manchester Academic Health Science Centre, Manchester, UK.
⁸ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA.
⁹ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Orthopaedic Surgery, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
¹⁰ Chan Zuckerberg BioHub, San Francisco, CA 94158, USA.
¹¹ Department of Orthopaedic Surgery, UC Davis Health, Sacramento, CA 95817, USA; Department of Biomedical Engineering, University of California, Davis, Davis, CA 95616, USA.
¹² Department of Biomedical Engineering, University of California, Davis, Davis, CA 95616, USA.
¹³ Department of Biomedical Engineering, University of California, Davis, Davis, CA 95616, USA; Department of Surgery, University of California Davis Health, Sacramento, CA 95817, USA; Institute for Pediatric Regenerative Medicine, Shriners Hospitals for Children, Sacramento, CA 95817, USA.
¹⁴ Department of Surgery, University of Alabama at Birmingham, Birmingham, AL 35233, USA.
¹⁵ Hagey Laboratory for Pediatric Regenerative Medicine, Stanford University School of Medicine, Stanford University, Stanford, CA 94305, USA.
¹⁶ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Ludwig Center for Cancer Stem Cell Biology and Medicine at Stanford University, Stanford, CA 94305, USA.
¹⁷ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA; Hagey Laboratory for Pediatric Regenerative Medicine, Stanford University School of Medicine, Stanford University, Stanford, CA 94305, USA. Electronic address: longaker@stanford.edu.
¹⁸ Department of Computer Science and Engineering, Jacob's School of Engineering, University of California, San Diego, La Jolla, CA 92093, USA; Department of Pediatrics, University of California, San Diego, La Jolla, CA 92093, USA. Electronic address: dsahoo@ucsd.edu.
¹⁹ Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA; Department of Surgery, Stanford University School of Medicine, Stanford, CA 94305, USA; Hagey Laboratory for Pediatric Regenerative Medicine, Stanford University School of Medicine, Stanford University, Stanford, CA 94305, USA.

PMID: 40118065
PMCID: PMC12048286 (available on 2026-05-01)
DOI: 10.1016/j.stem.2025.02.013

Abstract

The skeleton is one of the most structurally and compositionally diverse organ systems in the human body, depending on unique cellular dynamisms. Here, we integrate prospective isolation of human skeletal stem cells (hSSCs; CD45^-CD235a^-TIE2^-CD31^-CD146^-PDPN⁺CD73⁺CD164⁺) from ten skeletal sites with functional assays and single-cell RNA sequencing (scRNA-seq) analysis to identify chondrogenic, osteogenic, stromal, and fibrogenic subtypes of hSSCs during development and their linkage to skeletal phenotypes. We map the distinct composition of hSSC subtypes across multiple skeletal sites and demonstrate their unique in vivo clonal dynamics. We find that age-related changes in bone formation and regeneration disorders stem from a pathological fibroblastic shift in the hSSC pool. Utilizing a Boolean algorithm, we uncover gene regulatory networks that dictate differences in the ability of hSSCs to generate specific skeletal tissues. Importantly, hSSC lineage dynamics are pharmacologically malleable, providing a new strategy to treat aberrant hSSC diversity central to aging and skeletal maladies.

Keywords: Boolean relationships; bone aging; fibrous dysplasia; fracture healing; gene regulatory networks; human skeletal stem cell; nonunion; skeletal development.

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Conflict of interest statement

Declaration of interests The authors (T.H.A., R.S., I.L.W., M.T.L., D.S., and C.K.F.C.) have filed an invention disclosure (USPTO Application #63/676,773; filed on July 29, 2024) related to this work entitled: “Compositions and methods for re-activation of dysfunctional skeletal stem cells.”

References

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Human skeletal development and regeneration are shaped by functional diversity of stem cells across skeletal sites

Affiliations

Human skeletal development and regeneration are shaped by functional diversity of stem cells across skeletal sites

Authors

Affiliations

Abstract

Conflict of interest statement

References

MeSH terms

Grants and funding

LinkOut - more resources

Full Text Sources

Medical

Miscellaneous