Novel Sodium Deoxycholate-Based Chemical Decellularization Method for Peripheral Nerve
Abstract
Decellularized peripheral nerve has been proven to be an effective clinical intervention for peripheral nerve repair and a preclinical cell carrier after spinal cord injury. However, there are currently a lack of decellularization methods for peripheral nerve that remove cells and maintain matrix similar to the previously established, clinically translated technique (the Hudson method) that relies on the discontinued Triton X-200 detergent. Therefore, the aim of this study was to optimize a novel chemical decellularization method for peripheral nerves based on the currently available anionic detergent sodium deoxycholate. Sprague Dawley rat sciatic nerves were isolated, frozen in buffered solution, and then subject to sequential washes in water, salt buffer, zwitterionic detergents sulfobetaines -10 and -16, and varying concentrations of sodium deoxycholate (SDC). To optimize DNA removal after SDC decellularization, nerves were subjected to deoxyribonuclease (DNase) incubation and salt buffer washes. Immunohistochemical results demonstrated that utilization of 3% SDC in the decellularization process preserved extracellular matrix (ECM) components and structure while facilitating significantly better removal of Schwann cells, axons, and myelin compared with the Hudson method. The addition of a 3-h DNase incubation to the 3% SDC decellularization process significantly removed cellular debris compared with the Hudson method. Proteomic analysis demonstrated that our novel decellularization method based on 3% SDC +3-h DNase used in conjunction with zwitterionic detergents, and salt buffers (new decellularization method using 3% SDC + 3-h DNase, zwitterionic detergents, and salt buffers [SDD method]) produced a similar proteomic profile compared with the Hudson method and had significantly fewer counts of cellular proteins. Finally, cytotoxicity analysis demonstrated that the SDD decellularized scaffolds do not contain significant cytotoxic residuals as eluted media supported metabolically active Schwann cells in vitro . Overall, this study demonstrates that SDD decellularization represents a novel alternative utilizing currently commercially available chemical reagents. Impact Statement: Decellularized nerves are clinically relevant materials that can be used for a variety of regenerative applications such as peripheral nerve and spinal cord injury repair. However, discontinuation of key detergents used in a proven chemical decellularization process necessitates the optimization of an equivalent or better method. This research presents the field with a novel chemical decellularization method to replace the previous validated standard. Scaffolds generated from this method provide an extracellular matrix-rich material that can be used in a variety of in vitro applications to understand cellular behavior and in vivo applications to facilitate regeneration after neural injury.
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Prolonged exposure to nucleases can modify ECM structure and compromise its mechanical stability.
“However, prolonged exposure to nucleases can modify ECM structure and compromise its mechanical stability.”
From Bioactive scaffolds for tissue engineering: A review of decellularized extracellular matrix applications and innovations (2025) · cites this paper as [47]
DNase is typically applied after chemical detergents to solubilize cell membranes and dissociate their inner structure, enhancing DNase infiltration and efficiency within tissues.
“Deoxyribonuclease (DNase), a common nuclease, is typically applied after chemical detergents to solubilize cell membranes and dissociate their inner structure, enhancing DNase infiltration and efficiency within tissues [[152], [153], [154]].”
From Liver tissue engineering using decellularized scaffolds: Current progress, challenges, and opportunities (2024) · cites this paper as [153]
Prolonged exposure to nucleases used in research settings, which may contain impurities, may negatively impact ECM structure and constituents, affecting the mechanical stability.
“Prolonged exposure to nucleases used in research settings, which may contain impurities, may negatively impact ECM structure and constituents, affecting the mechanical stability [152,153].”
From Liver tissue engineering using decellularized scaffolds: Current progress, challenges, and opportunities (2024) · cites this paper as [153]
DNase may exert detrimental influences on ECM composition, as prolongation of DNase treatment time is indicated to deplete major basal lamina constituents, laminin, and collagen IV.
“DNase may also exert detrimental influences on ECM composition, as prolongation of DNase treatment time is indicated to deplete major basal lamina constituents, laminin, and collagen IV [98].”
From Immunogenicity of decellularized extracellular matrix scaffolds: a bottleneck in tissue engineering and regenerative medicine (2023) · cites this paper as [98]
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Self-citation
- Authors
- McCrary, M. W., Vaughn, N. E., Hlavac, N., Song, Y. H., Wachs, R. A., & Schmidt, C. E.
- Affiliations
- University of Florida (US)
- University of Nebraska–Lincoln (US)
- Corresponding author
- Schmidt, Christine E.
- ORCID
- Venue
- Tissue Engineering Part C: Methods
- Publisher
- SAGE Publications
- Published
- 2020-01-01
- Volume
- 26
- Issue
- 1
- Pages
- 23-36
- ISSN
- 1937-3384 (ISSN-L)
- 1937-3392
- OpenAlex
- W2986814501
- Cited by
- 61
- References
- 49
- Citations by year
- 2026: 7 · 2025: 8 · 2024: 11 · 2023: 13 · 2022: 10 · 2021: 6 · 2020: 6
- FWCI
- 5.26
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Tissue Engineering Part C Methods (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Health Sciences › Medicine › Surgery › Tissue Engineering and Regenerative Medicine
- Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Nerve injury and regeneration
- Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Neuroscience and Neural Engineering
- Keywords
- sodium deoxycholate, extracellular matrix preservation, zwitterionic detergents, sulfobetaine, sciatic nerve, spinal cord injury, peripheral nerve repair, cytotoxicity, proteomic profile, cell-free scaffolds
- MeSH terms
- Animals, Cell Survival, Cholagogues and Choleretics, Deoxycholic Acid, Extracellular Matrix, Male, Peripheral Nerves, Rats, Sprague-Dawley, Proteome, Tissue Engineering, Rats, Tissue Scaffolds
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