Formation of tissue engineered composite construct of cartilage and skin using high density polyethylene as inner scaffold in the shape of human helix.

BACKGROUND

Formation of external ear via tissue engineering has created interest amongst surgeons as an alternative for ear reconstruction in congenital microtia.

OBJECTIVE

To reconstruct a composite human construct of cartilage and skin in the shape of human ear helix in athymic mice.

METHODS

Six human nasal cartilages were used and digested with Collagenase II. Chondrocytes were passaged in 175 cm(2) culture flasks at a density of 10,000 cells/cm(2). Frozen human plasma was then mixed with human chondrocytes. Six human skin samples were cut into small pieces trypsinized and resuspended. The keratinocytes were plated in six-well plate culture dishes at a density of 2×105 cells per well. Dermis tissues were digested and the fibroblast cells resuspended in six-well plate at the density of 10,000 cells per well. Fibrin-fibroblast layer and fibrin-keratinocytes were formed by mixing with human plasma to create 6 bilayered human skin equivalent (BSE) constructs. The admixture of fibrin chondrocytes layers was wrapped around high density polyethylene (HDP), and implanted at the dorsum of the athymic mice. The construct was left for 4 weeks and after maturation the mice skin above the implanted construct was removed and replaced by BSE for another 4 weeks.

RESULTS

Haematoxylin and Eosin showed that the construct consists of fine arrangement and organized tissue structure starting with HDP followed by cartilage, dermis and epidermis. Safranin-O staining was positive for proteoglycan matrix production. Monoclonal mouse antihuman cytokeratin, 34βE12 staining displayed positive result for human keratin protein.

CONCLUSIONS

The study has shown the possibility to reconstruct ear helix with HDP and tissue engineered human cartilage and skin. This is another step to form a human ear and hopefully will be an alternative in reconstructive ear surgery.