Development and Characterization of an Acellular Porcine Small Intestine Submucosa Scaffold for Use in Corneal Epithelium Tissue Engineering

ABSTRACT

Purpose: To produce an acellular small intestine submucosa (SIS) that would be a suitable scaffold for corneal epithelium tissue engineering.

Methods: The SIS was decellularized by immersion in 0.1% (wt/vol) sodium dodecyl sulfate (SDS). The efficacy of acellularization was confirmed by histological observation and DNA quantification. The mechanical properties were evaluated by uniaxial tensile testing. ELISA was performed to assess the growth factor contents. The cytotoxicity of SIS scaffolds and extracts to rabbit corneal epithelial cells was determined by CCK-8 assay. We also investigated the inflammatory reaction of SIS implanted subcutaneously in a rat. The biocompatibility was studied by rabbit interlamellar corneal transplantation and reseeding assay with cornea-derived cells. Immunofluorescent staining was used to detect the expression of CK3, ZO-1 and K13.

Results: Histological analyses showed that complete cell removal was achieved, and the DNA quantity, which reflects the presence of cellular materials, was significantly diminished in acellular SIS. Collagen fibers were properly preserved and appeared in an orderly fashion. The tissue structure, the mechanical properties and the growth factor contents within the acellular SIS were well retained. The CCK8 assay demonstrated that the acellular SIS scaffolds and extracts had no cytotoxicity to rabbit corneal epithelial cells. There was no sign that an immune reaction occurred with acellular SIS implanted subcutaneously in a rat. In fact, in vivo implantation to rabbit interlamellar stromal pockets showed good biocompatibility. We also observed that clusters of rabbit corneal epithelial cells were growing well on the surface of the SIS in vitro and the distinctive CK3, ZO-1 for corneal epithelial cells was detected.

Conclusions: The decellularized SIS retained the major structural components. The matrix is biocompatible with cornea-derived cells and might be a suitable scaffold for corneal epithelium tissue engineering.

KEYWORDS:

• Acellular small intestine submucosa
• tissue engineering
• scaffold
• corneal epithelium

Acknowledgments

The authors thank Feng Jinbo for selfless experimental assistance; Dr. Gao Wenjuan, The Key Laboratory of Cardiovascular Remodeling and Function Research, Chinese Ministry of Education and Chinese Ministry of Health, Qilu Hospital, Shandong University, for direction in cell culture technology.

Authors Contributions

Conceived and designed the experiments: LQD. Performed the experiments: FYW. Analyzed the data: FYW LQD QS XYW. Contributed reagents/materials/analysis tools: LQD. Contributed to the writing of the manuscript: FYW LQD.

Conflict of Interest

None of the authors has a conflict of interest with the submission.

Ethics approval

Approved by the Ethical Committee of Qilu Hospital at Shandong University.

Additional information

Funding

Publication of this article was supported by a grant of the Natural Science Foundation of Shandong Province (ZR2015HM050) and Key Research & Development Program of Shandong Province (2019GSF108085).