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3D hierarchically aligned nanofiber scaffolds promote cell migration for tissue regeneration

Nature Protocols (2026)Cite this article

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Abstract

During tissue regeneration, cells are recruited from surrounding tissue to the defect site. However, when the defect site is large and morphologically complex, cell recruitment often fails to match healthy tissue morphology, resulting in a dysfunctional repair. The integration of bioscaffolds can help to direct the repair process. Here, we present a protocol that integrates electrospinning, weaving, thermal fixation and modified gas-foaming technologies to fabricate 3D hierarchically aligned nanofiber scaffolds. The scaffolds exhibit high porosity, controlled fiber alignment and diverse configurations (uniaxial, bidirectional, radial and gradient alignments), creating effective ‘cell highways’ for promoting collective cell migration. Applications include hemostatic materials, skin and bone regeneration, hernia repair and biomedical swabs. Both in vitro and in vivo, the highly porous and directionally arranged 3D nanofiber scaffolds markedly enhance cell migration, accelerating the reconstruction of defective tissues. This protocol resolves challenges in production scalability, facilitating the wider adoption of these scaffolds, with a procedure intended for users with expertise in biomaterials and regenerative medicine. The 3D nanofiber scaffolds require 1 d to synthesize and result in improved cell migration during in situ tissue regeneration.

Key points

  • The 3D aligned nanofiber scaffolds provide spatial support for the repair of trauma-induced tissue defects, where the hierarchical alignment of the nanofibers create pathways for the migrating cells, facilitating their rapid and large-scale transportation from the surrounding tissues to the wound.

  • The scaffolds provide topological stimulation to enhance the functional activities of the recruited cells, thus facilitating the repair process.

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Fig. 1: The preparation of different 3D hierarchical nanofiber scaffolds with controlled alignment.
Fig. 2: The application of 3D hierarchical nanofiber scaffolds with controlled alignment in the biomedical field.
Fig. 3: Comparison of manufacturing methods for 3D nanofiber scaffolds.
Fig. 4: Quantification of nanofiber alignment by using ImageJ.
Fig. 5: Evaluation of the quality of nanofiber mat cutting.
Fig. 6: The 3D cell-migration model in vitro.
Fig. 7: The cell-recruitment effects of the 3D RAS and VAS by using the subcutaneous implantation model.
Fig. 8: The cell-recruitment effects of the 3D RAS and VAS in critical-size cranial bone defects.

Data availability

The data from this study are available from the corresponding author upon reasonable request.

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Acknowledgements

This work was funded by the National Key R&D program of China (2024YFA1108403, to S.C.), the National Natural Science Foundation of China (82472551, to S.C.) and the Zhejiang Provincial Natural Science Foundation of China (LR24H150001, to S.C.).

Author information

Author notes

  1. These authors contributed equally: Hao Pan, Jiebing Zhao, Ruyi Fan.

Authors and Affiliations

  1. Zhejiang Engineering Research Center for Tissue Repair Materials, Wenzhou Institute, University of Chinese Academy of Sciences, Wenzhou, China

    Hao Pan, Jiebing Zhao, Ruyi Fan & Shixuan Chen

  2. Department of Orthopaedic Surgery, the First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China

    Hao Pan

  3. Global Medical Affairs, Merz Aesthetics, Raleigh, NC, USA

    Alec D. McCarthy

  4. Department of Orthopaedic Surgery, the Second Affiliated Hospital, Jiangxi Medical College, Nanchang University, Nanchang, China

    Wenbing Wan

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  1. Hao Pan
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  4. Alec D. McCarthy
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Contributions

H.P., J.Z. and R.F designed and performed the experimental protocol and wrote the draft of this protocol. A.D.M. helped to analyze the fiber alignment and polish the manuscript. W.W. and S.C. revised the manuscript and approved the final version of this protocol.

Corresponding authors

Correspondence to Wenbing Wan or Shixuan Chen.

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Competing interests

The authors declare no competing interests.

Peer review

Peer review information

Nature Protocols thanks Linhao Li, Dong-Woo Cho and Alberto Sensini for their contribution to the peer review of this work.

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Key references

Chen, S. et al. Biomaterials 179, 46–59 (2018): https://doi.org/10.1016/j.biomaterials.2018.06.031

Chen, S. et al. Nano Lett. 19, 2059–2065 (2019): https://doi.org/10.1021/acs.nanolett.9b00217

Chen, S. et al. Sci. Adv. 7, eabg3089 (2021): https://doi.org/10.1126/sciadv.abg3089

Chen, S. et al. Appl. Phys. Rev. 7, 021406 (2020): https://doi.org/10.1063/1.5144808

Yuan, J. et al. Bioact. Mater. 39, 582–594 (2024): https://doi.org/10.1016/j.bioactmat.2024.04.025

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Pan, H., Zhao, J., Fan, R. et al. 3D hierarchically aligned nanofiber scaffolds promote cell migration for tissue regeneration. Nat Protoc (2026). https://doi.org/10.1038/s41596-026-01339-9

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