Low-detergent sonication decellularization preserves extracellular matrix architecture and enhances cell infiltration in human acellular dermal matrix.

INTRODUCTION

Acellular dermal matrix (ADM) scaffolds support tissue regeneration by providing structural integrity, biological cues, and a permissive microenvironment; optimal ADMs integrate with native tissue while preserving extracellular matrix (ECM) composition, microarchitecture, and biomechanics. Decellularization is a critical step in ADM development, as incomplete cellular removal can induce inflammation, whereas excessive chemical treatment can disrupt ECM structure and impair scaffold biofunctionality.

METHODS

Here, we optimized human dermal decellularization to develop ADMs that retain ECM instruction while enabling deep host-cell infiltration and effective recellularization. Two low-detergent protocols were evaluated, Protocol A (sequential hypotonic and hypertonic solutions) and Protocol B (0.1% Triton X-100 in a hypotonic solution), both incorporating controlled ultrasonication and benzonase treatment, and compared with a conventional high-detergent method (Protocol C).

RESULTS

All protocols effectively removed cellular material and nucleic acids, with substantial reductions in residual DNA content; however, Protocols A and B better preserved key ECM proteins, including collagen IV, elastin, and laminin, as confirmed by Raman spectroscopy and scanning electron microscopy. Functional assays demonstrated enhanced fibroblast adhesion and cell proliferation in Protocols A and B, whereas Protocol C showed limited cell growth. In ex vivo human skin wound models, Protocol B supported the highest cell infiltration and ECM deposition by day 21, followed by Protocol A, while Protocol C exhibited minimal integration and remodeling. Histological and immunohistochemical analyses consistently confirmed superior ECM preservation with low-detergent approaches.

CONCLUSIONS

Overall, combining controlled ultrasonication with minimal non-ionic detergent exposure represents a promising decellularization strategy that preserves ECM architecture, supports recellularization, and enhances scaffold integration potential for tissue engineering applications.