Reply to Kleebayoon et al.

Dear Editor,

Thank you for sending us your comment on ‘Fabrication of a three-dimensional scaffold-free trachea with horseshoe-shaped hyaline cartilage: Comment’ [1]. First of all, thank you for taking an interest in our paper, ‘Fabrication of a three-dimensional scaffold-free trachea with horseshoe-shaped hyaline cartilage’ [2].

We have already conducted cartilage differentiation experiments using adipose-derived stem cells, and the results showed that bone marrow-derived stem cells produced better cartilage differentiation. We also plan to conduct experiments using stem cells derived from umbilical cord blood.

As you have pointed out, there is room for further investigation into growth factors and culture conditions. We have referred to previous reviews for guidance on selecting growth factors [3], and we have also started experiments using a drug delivery system [4] to focus more on efficiency in the future administration method.

We acknowledge the importance of studying the mechanical properties of the newly formed trachea. Mechanical tests have been performed on the structures fabricated using previous bio-three-dimensional printers [5]. Based on the results of grasping impressions and special staining tests, we judged that the structure fabricated in this study was sufficiently stronger than those previously fabricated.

We agree that the grafts may have been affected by the immune response in the transplantation experiments. However, the experiments were conducted in nude rats, which lack a fully functional immune system, and no evidence of graft destruction was observed grossly or microscopically. We are planning to conduct experiments using CD4 immunostaining in the future. As a long-term model, a 1-year long-term survival model has been created and confirmed in the past in other cartilage models. In almost all three-dimensional models, capillary formation to the structures and the presence of red blood cells have been confirmed in the early stages. However, we have also confirmed that vascular endothelial cells are required to form many vascular structures, and the question is how much to mix them.

We also believe that it is essential to investigate another long-term model to confirm the changes in the body, and we are planning to conduct experiments using large animals such as pigs and immunostaining experiments using CD31 in the future.

Conflict of interest: None declared.

REFERENCES