Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Multiple pathways of CD34+ cell differentiation during embryogenesis

Cell Death & Differentiation (2026) Cite this article

Subjects

Abstract

CD34 has long been defined as a canonical marker for endothelial progenitors as well as hematopoietic stem cells, implicating its role in vascular development and hematopoiesis. However, the precise developmental hierarchy and lineage potential of CD34+ cells remain controversial. In this study, we integrated inducible genetic lineage tracing techniques, proteomics and single-cell RNA-seq (scRNA-seq) analyses to elucidate the dynamic developmental trajectory of CD34+ cells during various embryonic periods in both humans and mice. Remarkably, our analyses indicated that the progeny of CD34+ cells marked distinct, spatiotemporally restricted progenitor waves with divergent fates, at which point cells adopted endothelial, hematopoietic and fibroblastic fates, respectively. During gastrulation (E6.5–E8.5), an initial wave of CD34+ progenitors predominantly orchestrates vasculogenesis via a Kdr-dependent mechanism. Subsequently, from E9.5 to E14.5, cell cycle activation serves as a molecular switch, facilitating the endothelial-to-hematopoietic transition (EHT) of CD34+ progenitors. Unexpectedly, we identify a wave of CD34+ progenitors in late embryogenesis that gives rise to fibroblasts, distinct from earlier endothelial or hematopoietic lineages. Furthermore, because umbilical cord blood is a valuable source of different circulating stem/progenitor cells, we distinguish circulating endothelial progenitors from fibroblast progenitors in human cord blood by unique molecular signatures, with GFPT2 specifically marking the fibroblast progenitors. Collectively, our study provides a high-resolution spatiotemporal atlas of CD34+ cells during embryogenesis, redefining the temporal shifts of CD34+ cells in cell states and offering a precise framework for manipulating CD34+ cells in regenerative medicine.

The alternative text for this image may have been generated using AI.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to the full article PDF.

USD 39.95

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: The proteomics unveiled the dynamic changes of nascent human embryos from PCW6 to PCW12.
The alternative text for this image may have been generated using AI.
Fig. 2: Time-resolved single-cell transcriptome profiling of Cd34+ cells uncovered the developmental potential.
The alternative text for this image may have been generated using AI.
Fig. 3: CD34+ cells intricately orchestrated endothelial differentiation throughout embryogenesis.
The alternative text for this image may have been generated using AI.
Fig. 4: Ablation of Kdr in CD34+ cells resulted in stalled vessel development during the stage spanning E6.5 to E8.5.
The alternative text for this image may have been generated using AI.
Fig. 5: CD34+ cells gave rise to fibroblasts in E18.5 embryos.
The alternative text for this image may have been generated using AI.
Fig. 6: Ablation of Pdgfra in CD34+ cells stalled late-stage cardiovascular development.
The alternative text for this image may have been generated using AI.
Fig. 7: GFPT2, a candidate biomarker for distinguishing circulating fibroblast progenitor cells from endothelial progenitor cells.
The alternative text for this image may have been generated using AI.

Similar content being viewed by others

Data availability

The publicly available datasets are provided in the supplementary material. Single-cell RNA-sequencing data generated in this study will be publicly accessible from the NCBI Gene Expression Omnibus (GSE317390). The human sequence data reported in this article are available in the Genome Sequence Archive in the National Genomics Data Center, China National Center for Bioinformation/Beijing Institute of Genomics, Chinese Academy of Sciences (HRA016338). Any additional information reported in this paper is available upon reasonable request.

References

  1. Asahara T, Masuda H, Takahashi T, Kalka C, Pastore C, Silver M, et al. Bone marrow origin of endothelial progenitor cells responsible for postnatal vasculogenesis in physiological and pathological neovascularization. Circ Res. 1999;85:221–8.

    Article  CAS  PubMed  Google Scholar 

  2. Wu H, Yang X, Chen T, Yu B, Chen M, Wang T, et al. Aneurysm is restricted by CD34+ cell-formed fibrous collars through the PDGFRb-PI3K Axis. Adv Sci. 2025;12:e2408996.

  3. Jiang L, Chen T, Sun S, Wang R, Deng J, Lyu L, et al. Non-bone marrow CD34 + cells are crucial for endothelial repair of injured arteries. Circ Res. 2021;129:e146–e165.

    Article  CAS  PubMed  Google Scholar 

  4. Krause D, Ito T, Fackler M, Smith O, Collector M, Sharkis S, et al. Characterization of murine CD34, a marker for hematopoietic progenitor and stem cells. Blood. 1994;84:691–701.

    Article  CAS  PubMed  Google Scholar 

  5. Sidney LE, Branch MJ, Dunphy SE, Dua HS, Hopkinson A. Concise review: evidence for CD34 as a common marker for diverse progenitors. Stem Cells. 2014;32:1380–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Brown J, Greaves MF, Molgaard HV. The gene encoding the stem cell antigen, CD34, is conserved in mouse and expressed in haemopoietic progenitor cell lines, brain, and embryonic fibroblasts. Int Immunol. 1991;3:175–84.

    Article  CAS  PubMed  Google Scholar 

  7. Kojima Y, Tam OH, Tam PPL. Timing of developmental events in the early mouse embryo. Semin Cell Dev Biol. 2014;34:65–75.

    Article  CAS  PubMed  Google Scholar 

  8. Tam PPL, Loebel DAF. Gene function in mouse embryogenesis: get set for gastrulation. Nat Rev Genet. 2007;8:368–81.

    Article  CAS  PubMed  Google Scholar 

  9. DeLaughter DM, Bick AG, Wakimoto H, McKean D, Gorham JM, Kathiriya IS, et al. Single-cell resolution of temporal gene expression during heart development. Dev Cell. 2016;39:480–90.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Cao J, Spielmann M, Qiu X, Huang X, Ibrahim DM, Hill AJ, et al. The single-cell transcriptional landscape of mammalian organogenesis. Nature. 2019;566:496–502.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Wood HB, May G, Healy L, Enver T, Morriss-Kay GM. CD34 expression patterns during early mouse development are related to modes of blood vessel formation and reveal additional sites of hematopoiesis. Blood. 1997;90:2300–11.

    Article  CAS  PubMed  Google Scholar 

  12. The International Mouse Phenotyping Consortium, Dickinson ME, Flenniken AM, Ji X, Teboul L, Wong MD, et al. High-throughput discovery of novel developmental phenotypes. Nature. 2016;537:508–14.

    Article  Google Scholar 

  13. Kingsley PD, Malik J, Fantauzzo KA, Palis J. Yolk sac–derived primitive erythroblasts enucleate during mammalian embryogenesis. Blood. 2004;104:19–25.

    Article  CAS  PubMed  Google Scholar 

  14. Maltsev VA, Wobus AM, Rohwedel J, Bader M, Hescheler J. Cardiomyocytes differentiated in vitro from embryonic stem cells developmentally express cardiac-specific genes and ionic currents. Circ Res. 1994;75:233–44.

    Article  CAS  PubMed  Google Scholar 

  15. Coscia F, Lengyel E, Duraiswamy J, Ashcroft B, Bassani-Sternberg M, Wierer M, et al. Multi-level proteomics identifies CT45 as a chemosensitivity mediator and immunotherapy target in ovarian cancer. Cell. 2018;175:159–170.e16.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Krinidis S, Chatzis V. A robust fuzzy local information C-Means clustering algorithm. IEEE Trans Image Process. 2010;19:1328–37.

    Article  PubMed  Google Scholar 

  17. Bezdek JC, Ehrlich R, Full W. FCM: The fuzzy c-means clustering algorithm. Comput Geosci. 1984;10:191–203.

  18. Chan MM, Smith ZD, Grosswendt S, Kretzmer H, Norman TM, Adamson B, et al. Molecular recording of mammalian embryogenesis. Nature. 2019;570:77–82.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Dong J, Hu Y, Fan X, Wu X, Mao Y, Hu B, et al. Single-cell RNA-seq analysis unveils a prevalent epithelial/mesenchymal hybrid state during mouse organogenesis. Genome Biol. 2018;19:31.

    Article  PubMed  PubMed Central  Google Scholar 

  20. Ren H, Zhou X, Yang J, Kou K, Chen T, Pu Z, et al. Single-cell RNA sequencing of murine hearts for studying the development of the cardiac conduction system. Sci Data. 2023;10:577.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Shalaby F, Rossant J, Yamaguchi TP, Gertsenstein M, Wu XF, Breitman ML, et al. Failure of blood-island formation and vasculogenesis in Flk-1-deficient mice. Nature. 1995;376:62–6.

    Article  CAS  PubMed  Google Scholar 

  22. Susaki EA, Tainaka K, Perrin D, Kishino F, Tawara T, Watanabe TM, et al. Whole-brain imaging with single-cell resolution using chemical cocktails and computational analysis. Cell. 2014;157:726–39.

    Article  CAS  PubMed  Google Scholar 

  23. Lazarov T, Loyher PL, Yang H, Choo ZN, Deng Z, Nowotschin S, et al. Characterization of the mammalian prodefinitive angio-hematopoietic lineage. Sci Immunol. 2025;10:eadt6616.

    Article  CAS  PubMed  Google Scholar 

  24. Kobayashi M, Wei H, Yamanashi T, Azevedo Portilho N, Cornelius S, Valiente N, et al. HSC-independent definitive hematopoiesis persists into adult life. Cell Rep. 2023;42:112239.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Komorowska K, Doyle A, Wahlestedt M, Subramaniam A, Debnath S, Chen J, et al. Hepatic leukemia factor maintains quiescence of hematopoietic stem cells and protects the stem cell pool during regeneration. Cell Rep. 2017;21:3514–23.

    Article  CAS  PubMed  Google Scholar 

  26. Zhou F, Li X, Wang W, Zhu P, Zhou J, He W, et al. Tracing haematopoietic stem cell formation at single-cell resolution. Nature. 2016;533:487–92.

    Article  CAS  PubMed  Google Scholar 

  27. Iwasaki H, Arai F, Kubota Y, Dahl M, Suda T. Endothelial protein C receptor-expressing hematopoietic stem cells reside in the perisinusoidal niche in fetal liver. Blood. 2010;116:544–53.

    Article  CAS  PubMed  Google Scholar 

  28. Benz C, Copley MR, Kent DG, Wohrer S, Cortes A, Aghaeepour N, et al. Hematopoietic stem cell subtypes expand differentially during development and display distinct lymphopoietic programs. Cell Stem Cell. 2012;10:273–83.

    Article  CAS  PubMed  Google Scholar 

  29. Ivanovs A, Rybtsov S, Anderson RA, Turner ML, Medvinsky A. Identification of the niche and phenotype of the first human hematopoietic stem cells. Stem Cell Rep. 2014;2:449–56.

    Article  Google Scholar 

  30. Wang Y, Yao F, Wang L, Li Z, Ren Z, Li D, et al. Single-cell analysis of murine fibroblasts identifies neonatal to adult switching that regulates cardiomyocyte maturation. Nat Commun. 2020;11:2585.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Zhang W, Bouchard G, Yu A, Shafiq M, Jamali M, Shrager JB, et al. GFPT2 -expressing cancer-associated fibroblasts mediate metabolic reprogramming in human lung adenocarcinoma. Cancer Res. 2018;78:3445–57.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Wang L, Yang Y, Ma H, Xie Y, Xu J, Near D, et al. Single-cell dual-omics reveals the transcriptomic and epigenomic diversity of cardiac non-myocytes. Cardiovasc Res. 2021;118:1548–63.

    Article  Google Scholar 

  33. Sun X, Wang T, Gong H, Qiu Y, Zhang Y, Chen M, et al. Circulating CD34+ fibroblast progenitors engaged in heart fibrosis of the allograft. Circ Res. 2026;138:e326558.

  34. Rosa FF, Pires CF, Kurochkin I, Ferreira AG, Gomes AM, Palma LG, et al. Direct reprogramming of fibroblasts into antigen-presenting dendritic cells. Sci Immunol. 2018;3:eaau4292.

    Article  PubMed  Google Scholar 

  35. Civin CI, Strauss LC, Brovall C, Fackler MJ, Schwartz JF, Shaper JH. Antigenic analysis of hematopoiesis. III. A hematopoietic progenitor cell surface antigen defined by a monoclonal antibody raised against KG-1a cells. J Immunol. 1984;133:157–65.

    Article  CAS  PubMed  Google Scholar 

  36. Young P, Baumhueter S, Lasky L. The sialomucin CD34 is expressed on hematopoietic cells and blood vessels during murine development. Blood. 1995;85:96–105.

    Article  CAS  PubMed  Google Scholar 

  37. Joe AWB, Yi L, Natarajan A, Le Grand F, So L, Wang J, et al. Muscle injury activates resident fibro/adipogenic progenitors that facilitate myogenesis. Nat Cell Biol. 2010;12:153–63.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Shalaby F, Ho J, Stanford WL, Fischer KD, Schuh AC, Schwartz L, et al. A requirement for Flk1 in primitive and definitive hematopoiesis and vasculogenesis. Cell. 1997;89:981–90.

    Article  CAS  PubMed  Google Scholar 

  39. Zhu Q, Gao P, Tober J, Bennett L, Chen C, Uzun Y, et al. Developmental trajectory of prehematopoietic stem cell formation from endothelium. Blood. 2020;136:845–56.

    Article  PubMed  PubMed Central  Google Scholar 

  40. Fennie C, Cheng J, Dowbenko D, Young P, Lasky L. CD34+ endothelial cell lines derived from murine yolk sac induce the proliferation and differentiation of yolk sac CD34+ hematopoietic progenitors. Blood. 1995;86:4454–67.

    Article  CAS  PubMed  Google Scholar 

  41. Dzierzak E. The emergence of definitive hematopoietic stem cells in the mammal. Curr Opin Hematol. 2005;12:197–202.

    Article  PubMed  Google Scholar 

  42. Du L, Sun X, Gong H, Wang T, Jiang L, Huang C, et al. Single-cell and lineage tracing studies reveal the impact of CD34+ cells on myocardial fibrosis during heart failure. Stem Cell Res Ther. 2023;14:33.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Xie J, Jiang L, Wang J, Yin Y, Wang R, Du L, et al. Multilineage contribution of CD34+ cells in cardiac remodeling after ischemia/reperfusion injury. Basic Res Cardiol. 2023;118:17.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Pu X, Zhu P, Zhou X, He Y, Wu H, Du L, et al. CD34+ cell atlas of main organs implicates its impact on fibrosis. Cell Mol Life Sci. 2022;79:576.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Asahara T, Murohara T, Sullivan A, Silver M, van der Zee R, Li T, et al. Isolation of putative progenitor endothelial cells for angiogenesis. Science. 1997;275:964–6.

    Article  CAS  PubMed  Google Scholar 

  46. Chen T, Sun X, Gong H, Chen M, Li Y, Wang T, et al. Host CD34+ cells are replacing donor endothelium of transplanted heart. J Heart Lung Transpl. 2023;42:1651–65.

    Article  Google Scholar 

  47. Rotmans JI, Heyligers JMM, Verhagen HJM, Velema E, Nagtegaal MM, de Kleijn DPV, et al. In vivo cell seeding with anti-CD34 antibodies successfully accelerates endothelialization but stimulates intimal hyperplasia in porcine arteriovenous expanded polytetrafluoroethylene grafts. Circulation. 2005;112:12–8.

    Article  CAS  PubMed  Google Scholar 

  48. Aoki J, Serruys PW, van Beusekom H, Ong ATL, McFadden EP, Sianos G, et al. Endothelial progenitor cell capture by stents coated with an antibody against CD34. J Am Coll Cardiol. 2005;45:1574–9.

    Article  CAS  PubMed  Google Scholar 

  49. Otis EM, Brent R. Equivalent ages in mouse and human embryos. Anat Rec. 1954;120:33–63.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

The authors thank Dr. Yanhua Hu for the significant contribution to the construction of laboratory animal models. The authors are grateful to pathologists Buyi Zhang and Fei Hu at the Second Affiliated Hospital, Zhejiang University School of Medicine, for technical assistance in human embryo sample collection and preparation. We thank the technical assistant of Oebiotech Company, Novogene, Ribobio and Pioneer in Proteomics Biolabs. We thank Yuanyuan Lyu and Yun Lyu from the core facilities, Zhejiang University School of Medicine, for their technical support.

Funding

This study is in part supported by the National Natural Science Foundation of China W2541023 (QBX), U24A20799 (WM), 31830039 (QBX), 82400490 (SSS), 82400575 (KC), 82200479 (LJJ); National Key Research and Development Program 2023YFC3606201 (WM), Central guidance for local scientific and technological development funding projects 2024ZY01016 (WM), “Pioneer” and “Leading Goose” R&D Program of Zhejiang 2025C02147 (WM), Natural Science Foundation of Zhejiang Province LMS25H020009 (HG), The Fundamental Research Funds for the Central Universities 226-2025-00184 (KC).

Author information

Author notes

  1. These authors contributed equally: Ting Wang, Hui Gong, Guoguo Ye.

Authors and Affiliations

  1. Department of Cardiology, the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China

    Ting Wang, Hui Gong, Guoguo Ye, Bohuan Zhang, Liujun Jiang, Yuesheng Zhang & Qingbo Xu

  2. Nanhu Brain-computer Interface Institute, Hangzhou, China

    Hui Gong

  3. Department of Vascular Surgery, the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China

    Ruihan Chen, Xueyin Huang & Kai Chen

  4. Department of Cardiology, Institute for Developmental and Regenerative Cardiovascular Medicine, Xinhua Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China

    Shasha Sun

  5. Department of Cardiology, Affiliated Zhejiang Hospital, Zhejiang University School of Medicine and Zhejiang Key Laboratory of Integrative Chinese and Western Medicine for Diagnosis and Treatment of Circulatory Diseases, Hangzhou, China

    Tingting Chen, Yuqing Pan & Wei Mao

  6. Department of Pathology, the Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China

    Jinghong Xu

  7. Department of Reproductive Medicine, the Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China

    Min Jin

Authors
  1. Ting Wang
    View author publications

    Search author on:PubMed Google Scholar

  2. Hui Gong
    View author publications

    Search author on:PubMed Google Scholar

  3. Guoguo Ye
    View author publications

    Search author on:PubMed Google Scholar

  4. Ruihan Chen
    View author publications

    Search author on:PubMed Google Scholar

  5. Shasha Sun
    View author publications

    Search author on:PubMed Google Scholar

  6. Xueyin Huang
    View author publications

    Search author on:PubMed Google Scholar

  7. Bohuan Zhang
    View author publications

    Search author on:PubMed Google Scholar

  8. Liujun Jiang
    View author publications

    Search author on:PubMed Google Scholar

  9. Yuesheng Zhang
    View author publications

    Search author on:PubMed Google Scholar

  10. Tingting Chen
    View author publications

    Search author on:PubMed Google Scholar

  11. Yuqing Pan
    View author publications

    Search author on:PubMed Google Scholar

  12. Jinghong Xu
    View author publications

    Search author on:PubMed Google Scholar

  13. Min Jin
    View author publications

    Search author on:PubMed Google Scholar

  14. Kai Chen
    View author publications

    Search author on:PubMed Google Scholar

  15. Wei Mao
    View author publications

    Search author on:PubMed Google Scholar

  16. Qingbo Xu
    View author publications

    Search author on:PubMed Google Scholar

Contributions

Conceptualization: QBX, TW, and HG. Methodology: TW, GGY, RHC, and MTY. Visualization: GGY and SSS. Supervision: HG, GGY, RHC, KC, WM, and QBX. Writing—original draft: TW and QBX. Writing—review and editing: TW, HG, GGY, RHC, SSS, XYH, BHZ, LJJ, YSZ, TTC, YQP, JHX, MJ, KC, WM, and QBX.

Corresponding authors

Correspondence to Kai Chen, Wei Mao or Qingbo Xu.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Wang, T., Gong, H., Ye, G. et al. Multiple pathways of CD34+ cell differentiation during embryogenesis. Cell Death Differ (2026). https://doi.org/10.1038/s41418-026-01735-4

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Version of record:

  • DOI: https://doi.org/10.1038/s41418-026-01735-4

Search

Advanced search

Quick links