Tissue Engineering and Regenerative Medicine

, Volume 16, Issue 1, pp 39–50 | Cite as

A Comparative Study of the Effects of Different Decellularization Methods and Genipin-Cross-Linking on the Properties of Tracheal Matrices

  • Yi Zhong
  • Ai Jiang
  • Fei Sun
  • Yuanfan Xiao
  • Ying Gu
  • Lei Wu
  • Yujie Zhang
  • Hongcan ShiEmail author
Original Article



Different decellularization methods can affect the integrity and the biomechanical and biocompatible properties of the tracheal matrix. Natural cross-linking with genipin can be applied to improve those properties. The goals of this study were to evaluate the effects of different decellularization methods on the properties of genipin-cross-linked decellularized tracheal matrices in rabbits.


The tracheas of New Zealand rabbits were decellularized by the Triton-X 100-processed method (TPM) and the detergent-enzymatic method (DEM) and were then cross-linked with genipin. Mechanical tests, haematoxylin–eosin staining, Masson trichrome staining, Safranin O staining, DAPI staining, scanning electronic microscopy (SEM), and biocompatibility tests were used to evaluate the treatment. The bioengineered trachea and control trachea were then implanted into allogeneic rabbits for 30 days. The structural and functional analyses were performed after transplantation.


The biomechanical tests demonstrated that the biomechanical properties of the decellularized tracheas decreased and that genipin improved them (p < 0.05). The histological staining results revealed that most of the mucosal epithelial cells were removed and that the decellularized trachea had lower immunogenicity than the control group. The analysis of SEM revealed that the decellularized trachea retained the micro- and ultra-structural architectures of the trachea and that the matrices cross-linked with genipin were denser. The biocompatibility evaluation and in vivo implantation experiments showed that the decellularized trachea treated with the DEM had better biocompatibility than that treated with the TPM and that immunogenicity in the cross-linked tissues was lower than that in the uncross-linked tissues (p < 0.05).


Compared with the trachea treated with the TPM, the rabbit trachea processed by the DEM had better biocompatibility and lower immunogenicity, and its structural and mechanical characteristics were effectively improved after the genipin treatment, which is suitable for engineering replacement tracheal tissue.


Tissue engineering Trachea Decellularization Genipin Biological properties 



This study was supported by the National Natural Science Foundation of China (No. 81770018), the Foundation of Taizhou People’s Hospital (No. ZL201830) and the Jiangsu Students’ Innovation Training Program (No. 201811117035Z).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical statement

All animal experiments were approved in compliance with the rules and regulations of Laboratory Animals formulated by the National Ministry of Science (IACUC no. SYXK2016-0041).


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Copyright information

© The Korean Tissue Engineering and Regenerative Medicine Society and Springer Science+Business Media B.V., part of Springer Nature 2018

Authors and Affiliations

  1. 1.Medical College of Yangzhou UniversityYangzhouChina
  2. 2.Center of Translational MedicineYangzhou UniversityYangzhouChina
  3. 3.Peking University Third HospitalBeijingChina
  4. 4.Taizhou People’s HospitalTaizhouChina

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